![MP/ARF5 promoter activity at different developmental stages. (a)–(c) MP/ARF5 (TAIR: AT1G19850) promoter activity in the Arabidopsis embryo (heart (a), early torpedo (b), and mature (c) stage) visualized by a stably transformed pARF5:SV40-3xGFP reporter [60] (green). Cell walls are counterstained by FM4-64 (red). (d) MP/ARF5 promoter activity in the mature Arabidopsis stem visualized by a pARF5:ER-EYFP reporter (green). Cell walls are counterstained by propidium iodide (red). c, cambium; p, phloem; x, xylem. Scale bars: 50μm. Asterisks in (d) label the position of primary vascular bundles.](https://www.researchgate.net/publication/267816314/figure/fig2/AS:268884385202178@1441118441420/MP-ARF5-promoter-activity-at-different-developmental-stages-a-c-MP-ARF5-TAIR_Q320.jpg)
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(Pro)cambium formation and proliferation: Two sides of the same coin?
Abstract and Figures
(Pro)cambium cells give rise to vascular tissues and form a reticulate meristem pervading the whole plant body.•In the root, the procambium arises from oriented and coordinated cell divisions, controlled by a mutual interaction between auxin and cytokinin signaling.•Cambium proliferation is regulated by a system involving extensive intercellular communication.•Currently, established regulatory networks initiating procambium cell identity and controlling cambium proliferation are barely connected.
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... This then initiates the process of secondary growth, during which the cambial meristem continuously generates secondary xylem toward the inside of the stem and secondary phloem toward the outside of the stem, resulting in radial stem expansion. 2 Secondary growth induced woodiness not only provides sturdiness to stems, allowing plants to grow taller 1 but has also been linked to more harsh growth conditions requiring drought tolerance. 3,4 Secondary vasculature production is performed by cambial initials located at the center of the cambial zone that undergo two types of cell division: (1) periclinal cell division giving rise to proliferating derivatives called phloem and xylem mother cells, and (2) anticlinal cell division supporting the increase in circumference of the cambial zone. ...
... In contrast, directing the cell division rate of the stem cells of the vascular cambium will provide quantitative and qualitative control over the production of wood as biomaterial, and may also be used to enhance drought resistance in crops. 1,2,[4][5][6] Although previous studies have provided important insights into the molecular mechanisms regulating cambial activity, the current models are far from complete. 1,2,5,6 For example, the flowering time genes SOC1 and FUL that have been identified as clear repressors of cambial activity 19 are often not included. ...
... 1,2,[4][5][6] Although previous studies have provided important insights into the molecular mechanisms regulating cambial activity, the current models are far from complete. 1,2,5,6 For example, the flowering time genes SOC1 and FUL that have been identified as clear repressors of cambial activity 19 are often not included. ...
Plant secondary growth, which is the basis of wood formation, includes the production of secondary xylem, which is derived from meristematic cambium cells embedded in vascular tissue. Here, we identified an important role for the Arabidopsis thaliana (Arabidopsis) AT-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED 15 (AHL15) transcriptional regulator in controlling vascular cambium activity. The limited secondary xylem development in inflorescence stems of herbaceous Arabidopsis plants was significantly reduced in ahl15 loss-of-function mutants, whereas constitutive or vascular meristem-specific AHL15 overexpression produced woody inflorescence stems. AHL15 was required for enhanced secondary xylem formation in the woody suppressor of overexpression of constans 1 (soc1) fruitfull (ful) double loss-of-function mutant. Moreover, we found that AHL15 induces vascular cambium activity downstream of the repressing SOC1 and FUL transcription factors, most likely similar to how it enhances lateral branching by promoting biosynthesis of the hormone cytokinin. Our results uncover a novel pathway driving cambium development, in which AHL15 expression levels act in parallel to and are dependent on the well-established TDIF-PXY-WOX pathway to differentiate between herbaceous and woody stem growth.
... The MtLM1-like gene is a class I HD-ZIP gene and appears to be strongly linked to vascularization in M. truncatula. In A. thaliana procambium initiation, MONOPTEROS has a major role in translating auxin accumulation into procambium identity (Jouannet et al. 2015) in the globular and heart stage embryos. MONOPTEROS regulates a class III HD-ZIP gene AtHB8. ...
... MONOPTEROS regulates a class III HD-ZIP gene AtHB8. The detailed signaling network is fully discussed by Jouannet et al. (2015). WOX4 is also expressed in the procambium (Ji et al. 2010) with a major role in promoting cambial activity (Ji et al. 2010;Jouannet et al. 2015). ...
... The detailed signaling network is fully discussed by Jouannet et al. (2015). WOX4 is also expressed in the procambium (Ji et al. 2010) with a major role in promoting cambial activity (Ji et al. 2010;Jouannet et al. 2015). In M. truncatula we did not find strong WOX4 expression in the heart stage embryo, which is surprising, with its expression being more connected to ovule development. ...
Medicago truncatula has readily accessible ovules that can be obtained from pods with morphological characteristics that correlate with embryos at different stages of development. In this chapter the expression pattern of a number of major transcription factors is examined, for M. truncatula embryogenesis, in relation to the characteristic morphology of the developing M. truncatula embryo. The embryo stages considered are the initial asymmetric zygotic division and early cell divisions, the development of the globular stage and the development of the shoot, root and procambium meristems of the heart stage embryo. The transcription factors belong to the AP2/ERF, ARF, HD‐ZIP, KNOX, WOX, and WRKY gene families. While embryogenesis in M. truncatula and Arabidopsis thaliana have much in common, there are a number of M. truncatula genes that may have distinctive roles or different roles to their A. thaliana homologues. The significance of transcriptional regulation studies in somatic embryos (SE) for understanding zygotic embryogenesis (ZE) regulation is also considered.
... Our understanding of the development of the vascular cambium reinforces this idea; the main genes involved in vascular cambium differentiation are also pleiotropically hardwired to procambium formation, such as meristem development for other nonwoody tissues (Jouannet et al. 2015). Observing the differences in achieving arborescence between eudicots and monocots and considering that a vascular cambium and secondary growth are more likely ancestral to seed plants than independently acquired, we hypothesized that the lack of a vascular cambium in monocots are likely due to the loss of individual genes, while herbaceous eudicots may have merely lost the regulatory interactions between genes essential for woody growth, while the required genes remain intact and conserved. ...
... We constructed an early vascular cambium differentiation gene network for A. thaliana from literature (Demura and Fukuda 2007;Lucas et al. 2013;Jouannet et al. 2015;De Rybel et al. 2016;Smet and De Rybel 2016) and investigated the presence of orthologs of these genes in the eudicots and monocots, as well as in Am. trichopoda ( fig. 2 and supplementary table S7, Supplementary Material online). Although the network was constructed using data from A. thaliana, we found that all genes included in the network showed 100% ortholog conservation in all eudicot species investigated, indicating that this network of vascular cambium differentiation is highly conserved in the eudicots. ...
... We observed 100% conservation of orthologs in this network across the eudicots, supporting our hypothesis that herbaceous eudicots have never lost orthologs of these genes, but that changes in gene interactions likely led to the absence of secondary xylem and an herbaceous growth habit in nonarborescent species. This observation also supports our understanding of vascular development; that many genes involved in vascular cambial differentiation are also pleiotropically linked to other processes in eudicots such as procambial formation and meristem development (Jouannet et al. 2015). An example involves the homeobox genes REV and PHABULOSA (PHB) that are not only involved in shoot apical meristem differentiation but also in the development of leaf polarity, flower development as well as xylem differentiation in eudicots (McConnell and Barton 1998). ...
Woodiness (secondary xylem derived from vascular cambium) has been gained and lost multiple times in the angiosperms, but has been lost ancestrally in all monocots. Here, we investigate the conservation of genes involved in xylogenesis in fully sequenced angiosperm genomes, hypothesising that monocots have lost some essential orthologs involved in this process. We analysed the conservation of genes preferentially expressed in the developing secondary xylem of two eudicot trees in the sequenced genomes of 26 eudicot and seven monocot species, and the early-diverging angiosperm Amborella trichopoda. We also reconstructed a regulatory model of early vascular cambial cell identity and differentiation and investigated the conservation of orthologs across the angiosperms. Additionally, we analysed the genome of the aquatic seagrass Zostera marina for additional losses of genes otherwise essential to, especially, secondary cell wall formation. Despite almost complete conservation of orthology within the early cambial differentiation gene network, we show a clear pattern of loss of genes preferentially expressed in secondary xylem in the monocots that are highly conserved across eudicot species. Our study provides candidate genes that may have led to the loss of vascular cambium in the monocots, and, by comparing terrestrial angiosperms to an aquatic monocot, highlights genes essential to vasculature on land.
... Primary functionality of 73 the program is to simulate the evolution of production of relevant chemicals in a 74 cambium cell. 75 The software incorporates a feature that allows application of so-called "control 76 actions", which manually override the state of any chosen node at any time step. This 77 feature allows simulation of the effect of gene knockout. ...
... These efforts resulted in discovery of a number of genes responsible for 441 each process. However, as mutations affecting procambium formation would also impact 442 later cambium functions, the relevance of various sets of data for modeling cambium 443 regulation requires further clarification [76]. In the absence of unequivocal experimental 444 evidence to the contrary, it is reasonable to assume that these networks act in concert. ...
Rise of atmospheric CO 2 is one of the main causes of global warming. Catastrophic climate change can be avoided by reducing emissions and increasing sequestration of CO 2 . Trees are known to sequester CO 2 during photosynthesis, and then store it as wood biomass. Thus, breeding of trees with higher wood yield would mitigate global warming as well as augment production of renewable construction materials, energy, and industrial feedstock. Wood is made of cellulose-rich xylem cells produced through proliferation of a specialized stem cell niche called cambium. Importance of cambium in xylem cells production makes it an ideal target for the tree breeding programs; however our knowledge about control of cambium proliferation remains limited. The morphology and regulation of cambium differs from stem cell niches that control axial growth. For this reason, translating the knowledge about axial growth to radial growth has limited use. Furthermore, genetic approaches cannot be easily applied because overlaying tissues conceal cambium from direct observation and complicate identification of mutants. To overcome the paucity of experimental tools in cambium biology, we constructed a Boolean network CARENET (CAmbium Regulation gene NETwork) for modelling cambium activity, which includes the key transcription factors WOX4 and HD-ZIP III as well as their potential regulators. Our simulations revealed that: (1) auxin, cytokinin, gibberellin, and brassinosteroids act cooperatively in promoting transcription of WOX4 and HD-ZIP III ; (2) auxin and cytokinin pathways negatively regulate each other; (3) hormonal pathways act redundantly in sustaining cambium activity; (4) individual cells in the stem cell niches can have diverse molecular identities. CARENET can be extended to include components of other signalling pathways and be integrated with models of xylem and phloem differentiation. Such extended models would facilitate breeding trees with higher wood yield.
... NRT2.4 and NRT2.5, were highly expressed in epidermis and root cap cells, concomitant with the role of sensing NO 3changes in soil(Ho et al., 2009), and uptake function respectively(Lhamo and Luan, 2021). NPF1.1 and NPF1.2 were expressed in procambium cells, indicating that they could be participating in loading NO 3to phloem and xylem cells in developing roots(Jouannet et al., 2015; ...
A plant’s response to external and internal nitrogen signals/status relies on sensing and signaling mechanisms that operate across spatial and temporal dimensions. From a comprehensive systems biology perspective, this involves integrating nitrogen responses in different cell types and over long distances to ensure organ coordination in real time and yield practical applications. In this prospective review, we focus on novel aspects of nitrogen (N) sensing/signaling uncovered using temporal and spatial systems biology approaches, largely in the model Arabidopsis. The temporal aspects span: transcriptional responses to N-dose mediated by Michaelis-Menten kinetics; the role of the master NLP7 transcription factor as a nitrate sensor, its nitrate-dependent TF nuclear retention, its “hit-and-run” mode of target gene regulation and temporal transcriptional cascade identified by “Network Walking”. Spatial aspects of N-sensing/signaling have been uncovered in cell type-specific studies in roots and in root-to-shoot communication. We explore new approaches using single cell sequencing data, trajectory inference and pseudotime analysis as well as machine learning and artificial intelligence approaches. Finally, unveiling the mechanisms underlying the spatial dynamics of nitrogen sensing/signaling networks across species from model-to-crop could pave the way for translational studies to improve nitrogen-use efficiency in crops. Such outcomes could potentially reduce the detrimental effects of excessive fertilizer usage on groundwater pollution and greenhouse gas emissions.
... The basal plasma membrane accumulation of PIN3, PIN4, and PIN7 in xylem parenchyma cells in the main stem of Arabidopsis is enhanced by CK by an unknown mechanism (Kotov et al., 2021;Wu et al., 2021). According to the data obtained on Arabidopsis roots, CKs can increase IAA content in the areas of procambial bundle establishment and proto-and metaxylem differentiation through specific effects on the direction of auxin transport Jouannet et al., 2015). It can be said that CKs are essential regulators of auxin distribution (Box 1), limiting lateral auxin transport and enabling cell differentiation and elongation (Wu et al., 2021) ...
This article comments on:
Cammarata J, Roeder AHK, Scanlon MJ. 2023. The ratio of auxin to cytokinin controls leaf development and meristem initiation in Physcomitrium patens. Journal of Experimental Botany 74, 6541–6550.
... We searched for Ar. thaliana genes involved in secondary cell wall formation 69,73,[77][78][79] , and these genes are highly conserved across eudicots (Supplementary Data 11). However, the XTH16 (XYLOGLUCAN ENDOTRANSGLUCOSYLASE/ HYDROLASE 16), CEV1 (CONSTITUTIVE EXPRESSION OF VSP 1), and AIL6/ 7 (AINTEGUMENTA-LIKE 6/7) genes were lost in monocots (Supplementary Table 24, Supplementary Data 11). ...
Monocots are a major taxon within flowering plants, have unique morphological traits, and show an extraordinary diversity in lifestyle. To improve our understanding of monocot origin and evolution, we generate chromosome-level reference genomes of the diploid Acorus gramineusand the tetraploid Ac. calamus, the only two accepted species from the family Acoraceae, which form a sister lineage to all other monocots. Comparing the genomes of Ac. grami- neus and Ac. calamus, we suggest that Ac. gramineus is not a potential diploid progenitor of Ac. calamus, and Ac. calamus is an allotetraploid with two subgenomes A, and B, presenting asymmetric evolution and B subgenome dominance. Both the diploid genome of Ac. gramineus and the subgenomes A and B of Ac. calamus show clear evidence of whole-genome duplication (WGD), but Acoraceae does not seem to share an older WGD that is shared by most other monocots. We reconstruct an ancestral monocot karyotype and gene toolkit, and discuss scenarios that explain the complex history of the Acorus genome. Our analyses show that the ancestors of monocots exhibit mosaic genomic features, likely important for that appeared in early monocot evolution, providing fundamental insights into the origin, evolution, and diversification of monocots.
... Interestingly, this analysis revealed that a significant portion of TNL genes, including those encoding TNLs with known functions such as RPP1, RPP5, SNC1, and RPS6, were predominantly expressed in the procambium cells with either mock treatment or fungal infection (Fig. 2B). Procambium is a cell type from which primary phloem and xylem cells derive (De Rybel et al., 2016;Jouannet et al., 2015). Many of these procambium-expressed NLR genes were further induced after fungal infection (Fig. 2B). ...
Plant infection by microbial pathogens is a dynamic process. Here, we investigated the heterogeneity of plant responses in the context of pathogen location. A single-cell atlas of Arabidopsis thaliana leaves challenged by the fungus Colletotrichum higginsianum revealed cell type-specific gene expression that highlights an enrichment of intracellular immune receptors in vasculature cells. Using trajectory inference, we assigned cells that directly interacted with the invasive hyphae. Further analysis of cells at these infection sites revealed transcriptional plasticity based on cell type. A reprogramming of abscisic acid signalling was specifically activated in guard cells. Consistently, a contact-dependent stomatal closure was observed, possibly representing a defense response that anticipates pathogen invasive growth. We defined cell type-specific deployments of genes activating indole glucosinolate biosynthesis at the infection sites, and determined their contribution to resistance. This research highlights the spatial dynamics of plant response during infection and reveals cell type-specific processes and gene functions.
... Additionally, SNP si2:6562955 was associated with MSW and near ERF1 (ethylene-responsive transcription factor) ( Fig. 1B and Supplementary Data 3). ERF1 is implicated in cambium proliferation 29 , which might influence main stem width. Interestingly, the candidate gene PRR37 near the peak SNP si2:49328133 of the MSPL (Fig. 1D), suppressed heading and showed shorter panicle length than its mutant in rice 30 , which might directly regulate the panicle length of foxtail millet. ...
Genetic and environmental factors collectively determine plant growth and yield. In the past 20 years, genome-wide association studies (GWAS) have been conducted on crops to decipher genetic loci that contribute to growth and yield, however, plant genotype appears to be insufficient to explain the trait variations. Here, we unravel the associations between genotypic, phenotypic, and rhizoplane microbiota variables of 827 foxtail millet cultivars by an integrated GWAS, microbiome-wide association studies (MWAS) and microbiome genome-wide association studies (mGWAS) method. We identify 257 rhizoplane microbial biomarkers associated with six key agronomic traits and validated the microbial-mediated growth effects on foxtail millet using marker strains isolated from the field. The rhizoplane microbiota composition is mainly driven by variations in plant genes related to immunity, metabolites, hormone signaling and nutrient uptake. Among these, the host immune gene FLS2 and transcription factor bHLH35 are widely associated with the microbial taxa of the rhizoplane. We further uncover a plant genotype-microbiota interaction network that contributes to phenotype plasticity. The microbial-mediated growth effects on foxtail millet are dependent on the host genotype, suggesting that precision microbiome management could be used to engineer high-yielding cultivars in agriculture systems. Plant genotype alone appears to be insufficient to explain trait variations. This study integrates GWAS, MWAS and mGWAS in 827 foxtail millet cultivars, revealing that root-associated microbiota affect plant phenotypes in a host genotype-dependent manner.
... The ground meristem also increased in thickness after radicle protrusion. The procambium provides the basis for the differential modulation of long-distance transport capacities and plant body stability (Jouannet et al. 2015). ...
Previous studies on the embryo structure of legumes species had resulted in the division of the Fabaceae family into two great subfamilies based on embryo axis curvature. Research on seed morphology and anatomy adds to the knowledge of taxonomy, evolution and ecology. This study determined the seed developmental anatomy, pod and seed morphology as well as germination characteristics of the observed variants (T1-small prickles; T2-medium prickles; T3-long prickles) of Pterocarpus indicus Willd. f. echinatus locally known as prickly narra in the Mount Makiling Forest Reserve (MMFR). Based on the anatomy of the root (radicle) and shoot apex, the formation of the leaf primordium in T2 seeds after radicle protrusion was more progressive. It was observed that the germination rate and the percentage were the highest in T2, where the apical dome was well-developed. The germination, pod and seed morphological characters as well as seed anatomical characters were proven to be systematically informative by showing significant differences among the variants.
... Furthermore, understanding root development at earlier stages is pivotal since it has a huge impact on root function. The root developmental processes in Beta vulgaris are mainly focused on the regulation of cambium activity, which produces precursors for the formation of xylem and phloem elements (Jouannet et al. 2015). However, there is not enough evidence showing the effects of priming/biopriming on the root xylem vessels, precisely, on the stimulation of cambium division and differentiation. ...
This study assesses the effects of Azotobacter biopriming on the early development of sugar beet. Azotobacter chroococcum F8/2 was screened for plant growth promoting characteristics and biopriming effects were estimated through germination parameters and the structural changes of the root tissues. A. chroococcum F8/2 was characterized as a contributor to nitrogen, iron, and potassium availability, as well as a producer of auxin and 1-aminocyclopropane-1-carboxilic acid deaminase. Applied biopriming had reduced mean germination time by 34.44% and increased vigor I by 90.99% compared to control. Volatile blend comprised 47.67% ethanol, 32.01% 2-methyl-propanol, 17.32% 3-methyl-1-butanol, and a trace of 2,3-butanedione. Root micromorphological analysis of bioprimed sugar beet revealed a considerable increase in primary, secondary xylem area, and vessels size. Obtained results determine A. chroococcum F8/2 as a successful biopriming agent, and active participant in nutrient availability and hormonal status modulation affecting root vascular tissue.
... Development of the phloem, (pro)cambium, and xylem is a dynamic process that integrates diverse extrinsic and intrinsic factors. Molecular and genetic components, including peptide ligands, receptors, transcription factors, and phytohormones, have been identified and characterized that regulate vascular development (reviewed in Lucas et al. 2013;Miyashima et al. 2013;Furuta et al. 2014a;Ohashi-Ito and Fukuda 2014;Jouannet et al. 2015;Nieminen et al. 2015;Etchells et al. 2016;Cho et al. 2017;Wang 2020). In particular, interactions of the peptide ligand TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FAC-TOR (TDIF) and its cognate receptor PHLOEM INTER-CALATED WITH XYLEM/TDIF RECEPTOR (PXY/ TDR) have been well described (Hirakawa et al. 2008, Kangseok Ko and Eun Kyung Yoon have contributed equally to this work. ...
In plants, vascular development is a dynamic process that integrates extrinsic and intrinsic factors. In Arabidopsis (Arabidopsis thaliana), SHORTROOT (SHR) has been known to play key roles in regulating cell division and differentiation in root vascular development. However, the role of SHR in the shoot vasculature remains unknown. Here, we employed various experimental approaches to unravel SHR’s role in shoot vascular development. In hypocotyls and inflorescence stems, shr exhibited reduced cell numbers in the (pro)cambium. As a result, the daughter cells that would be incorporated into the phloem and xylem would become a shortage of supply, thereby causing precocious differentiation. By expression profiling, we identified a putative SHR-mediated transcriptional regulatory network (TRN), which includes a subset of the ETHYLENE RESPONSE FACTOR (ERF) family. Among these, we found that SHR directly regulated expression of ERF018, whose expression domains were largely overlapped with those of SHR. Additionally, we found that ERF018 overexpressors (ERF018-OXs) showed a reduction in cambium cell division in hypocotyls. Interestingly, the level of SHR transcripts was elevated in ERF018-OX hypocotyls. Taken together, our results provide insights into the previously uncharacterized role of SHR, implying that maintaining homeostasis of key regulators’ levels plays an important role in shoot vascular development.
... This finding suggests that there are similar but distinct roles Extensive studies of Arabidopsis mutants have demonstrated that BR signaling coupled with auxin polar transport is required for cambium initiation in stems [12]. As part of this pathway, MP/ARF5 and its upstream positive regulators, PINs, play major roles in translating auxin accumulation into the establishment of procambium identity [34]. mp/arf5 is activated in preprocambial strands and exhibits severe defects in procambium formation [35]. ...
Wood formation involves sequential developmental events requiring the coordination of multiple hormones. Brassinosteroids (BRs) play a key role in wood development, but little is known about the cellular and molecular processes that underlie wood formation in tree species. Here, we generated transgenic poplar lines with edited PdBRI1 genes, which are orthologs of Arabidopsis vascular-enriched BR receptors, and showed how inhibition of BR signaling influences wood development at the mRNA and/or proteome level. Six Populus PdBRI1 genes formed three gene pairs, each of which was highly expressed in basal stems. Simultaneous mutation of PdBRI1–1, −2, −3 and − 6, which are orthologs of the Arabidopsis vascular-enriched BR receptors BRI1, BRL1 and BRL3, resulted in severe growth defects. In particular, the stems of these mutant lines displayed a discontinuous cambial ring and patterning defects in derived secondary vascular tissues. Abnormal cambial formation within the cortical parenchyma was also observed in the stems of pdbri1–1;2;3;6. Transgenic poplar plants expressing edited versions of PdBRI1–1 or PdBRI1–1;2;6 exhibited phenotypic alterations in stem development at 4.5 months of growth, indicating that there is functional redundancy among these PdBRI1 genes. Integrated analysis of the transcriptome and proteome of pdbri1–1;2;3;6 stems revealed differential expression of a number of genes/proteins associated with wood development and hormones. Concordant (16%) and discordant (84%) regulation of mRNA and protein expression, including wood-associated mRNA/protein expression, was found in pdbri1–1;2;3;6 stems. This study found a dual role of BRs in procambial cell division and xylem differentiation and provides insights into the multiple layers of gene regulation that contribute to wood formation in Populus.
... Fig. 3. presents the main phases of embryogenesis where it was observed the formation of procambium (pc), surrounding by the meristem (gm) in the different locations of callus tissue (Fig. 3). A mutually inhibitory action between auxin and cytokinin signaling is an important aspect of procambium formation [29]. Procambial cells have propensity to form root meristems or vascular tissues in response to added auxin [30]. ...
A growing demand for the ecologically pure products brings us for searching novel biotechnological approaches for plant cultivation. One of these approaches is the in vitro cultivation and further acclimatization of valuable plant species. The object of our investigation was Ajuga reptance L. ornamental plant which possesses high metabolic activity. In vitro cultivation was carried out applying Murashige-Skoog nutrient medium and its modifications. Acclimatization of in vitro plants was implemented according Hazarika. In the presence of twice higher concentration of cytokinins over auxins and 0.2 mg/ml gibberellins callus culture was formed from the leaf explants. Callus tissue was formed in the presence of 0.2 mg/ml kinetin and 2 mg/ml indole-3-acetic acid which has denser structure than the first one. The shoot formation was observed on callus cultures growing on the same medium approximately after 5th passage. Callus culture growth was supported also by the adding of 2 mg/ml 2,4-dichlorophenoxyacetic acid. For the micropropagation, the already formed shoots were transferred to the nutrient medium which contains only 0.1 mg/ml 1-Naphthaleneacetic acid as a phytohormone. A. reptans culture has high regenerative ability and the micro-propagation index was 104 – 105. In vitro regenerated plants were successfully acclimatized to the soil conditions during two-week period
(6) (PDF) Isolated culture of A. reptance L., its' morphological and growth features. Available from: https://www.researchgate.net/publication/357678111_Isolated_culture_of_A_reptance_L_its'_morphological_and_growth_features [accessed Jan 11 2022].
... Sub-clusters 2 and 6 showed high expression levels of the epidermal cell markers PDF1, HDG2, or FIDDLEHEAD (FDH) (1) (Fig. 2B). In contrast, the vascular meristem markers LIKE AUXIN RESISTANT 2 (LAX2) (23) and HOMEOBOX GENE 8 (HB8) (24) were highly expressed in subcluster 4 (Fig. 2B). The transcript profile of these marker genes indicates that the PC population contains the cells that differentiate into the epidermis, mesophyll, and vascular cells. ...
Despite the enormous potential of novel approaches to explore gene expression at a single-cell level, we lack a high-resolution and cell type-specific gene expression map of the shoot apex in woody perennials. We use single-nuclei RNA sequencing to determine the cell type-specific transcriptome of the Populus vegetative shoot apex. We identified highly heterogeneous cell populations clustered into seven broad groups represented by 18 transcriptionally distinct cell clusters. Next, we established the developmental trajectories of epidermal cells, leaf mesophyll, and vascular tissue. Motivated by the high similarities between Populus and Arabidopsis cell population in the vegetative apex, we created and applied a pipeline for interspecific single-cell expression data integration. We contrasted the developmental trajectories of primary phloem and xylem formation in both species, establishing the first comparison of primary vascular development between a model annual herbaceous and a woody perennial plant species. Our results offer a valuable resource for investigating the basic principles underlying cell division and differentiation conserved between herbaceous and perennial species, which also allows the evaluation of the divergencies at single-cell resolution.
... Fig. 3. presents the main phases of embryogenesis where it was observed the formation of procambium (pc), surrounding by the meristem (gm) in the different locations of callus tissue (Fig. 3). A mutually inhibitory action between auxin and cytokinin signaling is an important aspect of procambium formation [29]. Procambial cells have propensity to form root meristems or vascular tissues in response to added auxin [30]. ...
A growing demand for the ecologically pure products brings us for searching novel biotechnological approaches for plant cultivation. One of these approaches is the in vitro cultivation and further acclimatization of valuable plant species. The object of our investigation was Ajugareptance L. ornamental plant which possesses high metabolic activity. In vitro cultivation was carried out applying Murashige-Skoog nutrient medium and its modifications. Acclimatization of in vitro plants was implemented according Hazarika. In the presence of twice higher concentration of cytokinins over auxins and 0.2 mg/ml gibberellins callus culture was formed from the leaf explants. Callus tissue was formed in the presence of 0.2 mg/ml kinetin and 2 mg/ml indole-3-acetic acid which has denser structure than the first one. The shoot formation was observed on callus cultures growing on the same medium approximately after 5 th passage. Callus culture growth was supported also by the adding of 2 mg/ml 2,4-dichlorophenoxyacetic acid. For the micropropagation, the already formed shoots were transferred to the nutrient medium which contains only 0.1 mg/ml 1-Naphthaleneacetic acid as a phytohormone. A. reptans culture has high regenerative ability and the micro-propagation index was 10 ⁴ – 10 ⁵ . In vitro regenerated plants were successfully acclimatized to the soil conditions during two-week period.
... Plant secondary growth takes place in the vascular cambium (VC) cells, as they divide anticlinally to add new cambium initial cells, and periclinally to produce xylem or phloem mother cells (Spicer and Groover, 2010). The alternation of cambium cell division and differentiation is tightly regulated by internal and environmental cues, such as light, cold, drought, various nutrients, and many phytohormones, including auxin, cytokinin, ethylene, brassinosteroids (BRs), gibberellins (GAs), and strigolactones (SLs) (Jouannet et al., 2015;Nieminen et al., 2015;Bhalerao and Fischer, 2017). ...
During secondary growth, meristematic cells in the cambium can either proliferate to maintain the stem cell population or differentiate into xylem or phloem. The balance between these two developmental trajectories is tightly regulated by many environmental and endogenous cues. Strigolactones (SLs), a class of plant hormones, were previously reported to regulate secondary growth by promoting cambium activity. However, the underlying molecular mechanisms of SL action in plant secondary growth are not well understood. We performed histological, genetic, and biochemical analyses using genetic materials in Arabidopsis (Arabidopsis thaliana) with altered activity of the transcription factors BRI1-EMS-SUPPRESSOR1 (BES1) or WUSCHEL-related HOMEOBOX4 (WOX4) or lacking MORE AXILLARY SHOOT2 (MAX2), a key positive component in the SL signaling pathway. We found that BES1, a downstream regulator in the SL signaling pathway that promotes shoot branching and xylem differentiation, also inhibits WOX4 expression, a key regulator of cambium cell division in the intercellular TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FACTOR (TDIF)-TDIF RECEPTOR (TDR) signaling pathway. The antagonistic roles of BES1 and WOX4 in the regulation of cambium activity may integrate intercellular TDIF signals to efficiently and bidirectionally modulate cambium cell proliferation and differentiation. As both BES1 and WOX4 are widely involved in various endogenous signals and responses to environmental stimuli, these findings may provide insight into the dynamic regulation of cambium development.
... MiRNAs are key players in controlling the establishment, patterning, and function of meristems. For example, miR165/6-mediated post-transcriptional regulation of homeodomain-leucine zipper transcription factors orchestrate shoot and root development (Muraro et al. 2014;Couzigou and Combier 2016), vascular differentiation, and patterning (Byrne 2006;Carlsbecker et al. 2010;Miyashima et al. 2011;Zhu and Helliwell 2011;Jouannet et al. 2015;Nieminen et al. 2015;Ramachandran et al. 2016). The differentiation of vascular tissue is closely associated with wood formation and properties (Côté et al. 2010). ...
Plant growth and development is usually characterized by chronological age over the plants’ lifetimes. Age-related changes actually originate with meristems because they control if, where, when, and how new tissues are formed along the axis of the shoot. The “time-keeping” of plant meristem development is a complex process. To uncover the post-transcriptional regulation underlying the chronological development of Larix kaempferi (Japanese larch) meristems, we investigated the miRNA-mediated regulatory network in the defoliated, uppermost main stems of 1-, 2-, 5-, 10-, 25-, and 50-year-old L. kaempferi using RNA-seq methods. We identified 29 high-confidence miRNAs, three of which were defined, age-related miRNAs whose expression changed depending on L. kaempferi age, and 17 showed coordinated expression patterns with three age-related miRNAs based on hierarchical correlations. All hierarchically coordinated miRNAs and their targets constituted a miRNA-mediated regulatory network. The developmental timing pathway lka-miR-1-5p-156-SBP/SPL (Squamosa Promoter Binding Protein-Like), the lignin biosynthesis pathway lka-miR-7,13-5p-397-LAC (Laccase), and an unknown pathway lka-miR-3-5p-CMSS1 (Cms1 Ribosomal Small Subunit Homolog) were age-driven, and information from auxin and light could be integrated by the lka-miR-9-5p-390-TAS/ARF (Trans-Acting siRNA3/Auxin Response Factor) and lka-miR-8-5p-IRL4 (Plant Intracellular Ras-Group-Related LRR Protein 4) pathways, respectively. Age-driven regulatory network will lead the way to understand which and how genes mutually cross-regulate their activity orchestrating development of meristems of L. kaempferi with age. We also discussed and contributed to miRNA annotation and nomenclature.
... Unlike in the RAM where primary phloem strands derive from precursor cells that also give rise to procambium strands (Mähönen et al. 2000, Lucas et al. 2013, secondary phloem tissues in the stem base derive from cambial cells. Cambial cells maintain a radial, secondary meristem and produce secondary vasculature (Lucas et al. 2013, Jouannet et al. 2015, Shi et al. 2019). Due to different cellular origins, it is intriguing to investigate whether the molecular mechanisms involving SMXL3/4/5 genes and promoting primary and secondary phloem development share similar features. ...
During post-embryonic development, plants rely on the integrity of phloem within their root systems. The phloem is part of the vasculature and transports energy metabolites from leaves into mitotically active regions such as the root apical meristem (RAM). Loss of function of genes regulating phloem development can result in severe changes in root growth and plant body architecture. The redundantly active genes SUPPRESSOR OF MAX2 1-LIKE3 (SMXL3), SMXL4 and SMXL5 are central regulators of early phloem formation. However, molecular mechanisms underlying SMXL3/4/5 gene activities during early phloem development are mostly unknown. The functional relevance of SMXL3/4/5 protein domains including the EAR motif is also unclear. The aim of my dissertation was to characterise the mode of action of SMXL3/4/5 during early events of phloem development in detail using Arabidopsis thaliana roots as model organ to investigate spatiotemporal tissue formation. First, I investigated at which developmental steps SMXL3/4/5 genes are required to promote phloem development in the RAM, how they interact genetically with positive regulators (OPS, BRX) and how their function is affected by negative regulators (CLE26, CLE45). I found that SMXL4/5 function is required to initiate and promote the activities other of of genes regulating phloem development (OPS, BRX, BAM3, CVP2 and APL), and that SMXL4/5 protein functions are possibly required to attenuate CLE-mediated suppression of phloem differentiation. Furthermore, I examined whether the highly conserved EAR motif of SMXL5 is functionally relevant to promote early phloem development. Here, I tested whether protein accumulation was altered for EAR motif-mutated SMXL5 proteins (SMXL5mEAR) in planta, and if phloem formation could be restored in smxl4;smxl5 double mutants complemented with SMXL5mEAR proteins. My data suggest that SMXL5 protein function is independent from the EAR motif indicating that SMXL5 proteins do not act as canonical EAR repressors in the context of phloem development. Last, I aimed at identifying new genes that are functionally related to SMXL3/4/5 during early phloem development. Therefore, I performed an ethyl methanesulfonate (EMS)-based mutagenesis of smxl4;smxl5 double mutants to screen for genetic suppressors that alleviate the phloem defects characteristic for smxl4;smxl5 mutants. I found that mutagenesis of yet unknown suppressor genes in the smxl4;smxl5 background could indeed restore phloem development. Further analysis including genome mapping is required to identify candidate genes that result in the suppression of the smxl4;smxl5 mutant phenotype. In conclusion, I postulate that SMXL3, SMXL4 and SMXL5 genes are required to establish the post-embryonic phloem lineage and regulate the phloem-specific developmental program in the RAM. Together, a complex, tightly balanced network of molecular players depending on SMXL3/4/5 activities ensures the formation of phloem within the root system.
... were involved in transferring root-derived NO 3 − into phloem of mature leaves for redistribution to young leaves (Hsu and Tsay, 2013). Root single cell transcriptomes indicated these NPF1s were broadly expressed in stellar cells, with the highest expression in the procambium (Figure 2A), the meristematic tissue that give rise to xylem and phloem (Jouannet et al., 2015;De Rybel et al., 2016). This expression pattern suggests NPF1.1/1.2 may function in NO 3 − loading to both meristematic and mature phloem and xylem cells in developing roots under the normal growth conditions. ...
Nitrogen (N), phosphorus (P), and potassium (K) are three major macronutrients essential for plant life. These nutrients are acquired and transported by several large families of transporters expressed in plant roots. However, it remains largely unknown how these transporters are distributed in different cell-types that work together to transfer the nutrients from the soil to different layers of root cells and eventually reach vasculature for massive flow. Using the single cell transcriptomics data from Arabidopsis roots, we profiled the transcriptional patterns of putative nutrient transporters in different root cell-types. Such analyses identified a number of uncharacterized NPK transporters expressed in the root epidermis to mediate NPK uptake and distribution to the adjacent cells. Some transport genes showed cortex- and endodermis-specific expression to direct the nutrient flow toward the vasculature. For long-distance transport, a variety of transporters were shown to express and potentially function in the xylem and phloem. In the context of subcellular distribution of mineral nutrients, the NPK transporters at subcellular compartments were often found to show ubiquitous expression patterns, which suggests function in house-keeping processes. Overall, these single cell transcriptomic analyses provide working models of nutrient transport from the epidermis across the cortex to the vasculature, which can be further tested experimentally in the future.
... Root and shoot apical meristems namely RAM and SAM respectively are created during the process of plant embryo development. The four initial cells of the procambium are formed as early as the globular embryo stage (Figure 1.1A) (reviewed in (Jouannet et al. 2015)). However, the vascular cambium, also known as lateral meristem, appears at later stages of plant development due to the influence of the hormone-triggered cellular differentiation mechanisms (Figure 1.1B) (reviewed in (Schuetz et al. 2013)). ...
Focus on biomass production has increased due to the depletion of nonrenewable sources of energy. Trees are the most important source of biomass. Being
perennial by nature, they produce the majority of terrestrially available biomass.
Therefore, there is a vital need for a good understanding of tree development. Using
Arabidopsis, previous studies in our lab have identified the influence of the receptor
kinase PXY and ligand CLE signalling pathways in regulating vascular cell division.
It has been shown that manipulating PXY-CLE signalling pathway increases
vascular cell division in Arabidopsis. PXY-CLE functions towards increasing the
stem cell population in the procambium/cambium of the Arabidopsis. In the current
study, we have altered the expression of poplar homologues of PXY and CLE41 in
attempt to increase cambial cell division. The poplar homologues of PXY and CLE41
genes were cloned from hybrid aspen (Populus tremula x P. tremuloides), and will
be referred to as PttPXY and PttCLE41. Using tissue-specific over-expression of
these PttPXY and PttCLE41 genes we were able to significantly increase the rate of
tree growth and biomass above ground. Not only did the poplars exhibit a two-fold
increase in the rate of wood formation, but they were also taller and possessed larger
leaves compared to the untransformed controls. Together these results suggest that
engineering PXY-CLE41 signalling offers an opportunity to dramatically increase
commercial tree productivity.
... Vascular bundles are composed of xylem, phloem and meristematic tissues (vascular cambium) (Sachs, 1981). Differentiation from the cambium results in addition of xylem and phloem both inwards and outwards (Jouannet et al., 2015). The differentiation and growth of the vascular system are hormonally controlled with both environmental and genetic cues that regulate cambium activity and formation (Aloni, 1987;Scarpella and Meijer, 2004). ...
Vascular bundles in the grape pedicel and berry contain the conduits, phloem and xylem, for transport of water, sugar, nutrients and signals into and through the grape berry and play a critical role in berry growth and composition. Here, we assess the vascular anatomy within the proximal region of the berry. Guided using a 3D berry model generated by micro-CT, differential staining of transverse sections of berries and receptacles was followed by fluorescent microscopy. Morphometric and vascular characteristics were analyzed within the central proximal region (brush zone, a fibrous extension from the pedicel vascular system into the berry) of the seeded cultivars Shiraz and Sauvignon Blanc, as well as the stenospermocarpic cultivars Ruby Seedless and Flame Seedless. Observations revealed a change in vascular arrangement from the receptacle into the berry brush zone and differences in xylem element size as well as xylem and phloem area relationships. Xylem anatomical and derived hydraulic parameters, as well as total tissue area of xylem and phloem varied between cultivars and in receptacle and berry components. Variation in vascular growth between grape pedicels and berries was independent of seededness. Differences in receptacle xylem vessel size and distribution could contribute to cultivar-dependent xylem backflow constraint.
... Synthesis of IAA and its polar transport are the basis for growth, organogenesis and histogenesis in the development of shoot apical meristem and the entire plant body [30,31,146,147]. The integrated action of auxin synthesis, transport and metabolism defines the spatiotemporal pattern of auxin accumulation [31]. ...
Auxin alone or supplemented with cytokinins and strigolactones were long considered as the main player(s) in the control of apical dominance (AD) and correlative inhibition of the lateral bud outgrowth, the processes that shape the plant phenotype. However, past decade data indicate a more sophisticated pathways of AD regulation, with the involvement of mobile carbohydrates which perform both signal and trophic functions. Here we provide a critical comprehensive overview of the current status of the AD problem. This includes insight into intimate mechanisms regulating directed auxin transport in axillary buds with participation of phytohormones and sugars. Also roles of auxin, cytokinin and sugars in the dormancy or sustained growth of the lateral meristems were assigned. This review not only provides the latest data on implicated phytohormone crosstalk and its relationship with the signaling of sugars and abscisic acid, new AD players, but also focuses on the emerging biochemical mechanisms, at first positive feedback loops involving both sugars and hormones, that ensure the sustained bud growth. Data show that sugars act in concert with cytokinins but antagonistically to strigolactone signaling. A complex bud growth regulating network is demonstrated and unresolved issues regarding the hormone–carbohydrate regulation of AD are highlighted.
... Xylem bridges comprise lignified xylem vessels that span the entire haustorium and connect parasite to host vasculature. The (pro-)cambium orchestrates vascular development in plants (Jouannet et al., 2015). (Pro-)cambial-like domains within the haustorium cortex express cambial marker genes such as CLE41, GSK3, BES1, and WOX4 in C. japonica (Shimizu et al., 2017). ...
One-sentence summary
Recent advances provide insight into the molecular mechanisms underlying host-dependent seed germination and haustorium formation in parasitic plants.
... We will emphasize the "syntax" of such control, rather than its "semantics." This latter has been reviewed recently and comprehensively elsewhere (e.g., Jouannet et al. 2015;De Rybel et al. 2016;Etchells et al. 2016;Cho et al. 2017;Ramachandran et al. 2017;Ruonala et al. 2017;Anne and Hardtke 2018;Fischer et al. 2019;Fukuda and Ohashi-Ito 2019). ...
Throughout plant development, vascular cells continually form from within a population of seemingly equivalent cells. Vascular cells connect end to end to form continuous strands, and vascular strands connect at both or either end to form networks of exquisite complexity and mesmerizing beauty. Here we argue that experimental evidence gained over the past few decades implicates the plant hormone auxin-its production, transport, perception, and response-in all the steps that lead to the patterned formation of the plant vascular system, from the formation of vascular cells to their connection into vascular networks. We emphasize the organizing principles of the cell- and tissue-patterning process, rather than its molecular subtleties. In the picture that emerges, cells compete for an auxin-dependent, cell-polarizing signal; positive feedback between cell polarization and cell-to-cell movement of the polarizing signal leads to gradual selection of cell files; and selected cell files differentiate into vascular strands that drain the polarizing signal from the neighboring cells. Although the logic of the patterning process has become increasingly clear, the molecular details remain blurry; the future challenge will be to bring them into razor-sharp focus.
... These processes are largely dependent on phosphorylation status and nucleocytoplasmic distribution, indicating that canonical BR signaling confers important signaling scaffolds in these cross-talk-driven developmental processes [29]. The primary growth occurs during the initial stages of the plant life cycle, and later, secondary growth additionally provides mechanical strength and generates cells that conduct nutrients and water, supporting the enlarged plant body [30]. Specifically, plant secondary growth is initiated with the formation of the vascular cambium, which continuously provides specialized conducting cells bidirectionally, enabling plants to transport water and nutrients [31]. ...
Brassinosteroids (BRs) play crucial roles in the physiology and development of plants. In the model plant Arabidopsis, BR signaling is initiated at the level of membrane receptors, BRASSINOSTEROIDS INSENSITIVE 1 (BRI1) and BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1) complex, thus activating the transcription factors (TFs) BRASSINAZOLE RESISTANT 1/BRI1-EMS-SUPPRESSOR 1 (BZR1/BES1) to coordinate BR responsive genes. BRASSINOSTEROIDS INSENSITIVE 2 (BIN2), glycogen synthase kinase 3 (GSK3) like-kinase, negatively regulates BZR1/BES1 transcriptional activity through phosphorylation-dependent cytosolic retention and shuttling. However, it is still unknown whether this mechanism is conserved in Panax ginseng C. A. Mayer, a member of the Araliaceae family, which is a shade-tolerant perennial root crop. Despite its pharmacological and agricultural importance, the role of BR signaling in the development of P. ginseng and characterization of BR signaling components are still elusive. In this study, by utilizing the Arabidopsisbri1 mutant, we found that ectopic expression of the gain of function form of PgBZR1 (Pgbzr1-1D) restores BR deficiency. In detail, ectopic expression of Pgbzr1-1D rescues dwarfism, defects of floral organ development, and hypocotyl elongation of bri1-5, implying the functional conservation of PgBZR1 in P. ginseng. Interestingly, brassinolide (BL) and BRs biosynthesis inhibitor treatment in two-year-old P. ginseng storage root interferes with and promotes, respectively, secondary growth in terms of xylem formation. Altogether, our results provide new insight into the functional conservation and potential diversification of BR signaling and response in P. ginseng.
... Xylem is a highly specialized vascular tissue with the roles of transporting water and minerals and also providing mechanical support for upright growth. The formation of xylem is mainly dependent on the activity of (pro)cambium and the rate of differentiation from it (Jouannet et al., 2015;Kondo et al., 2015). Several hormonal signals have been shown to be involved in maintenance of cambium and the formation of secondary vascular tissues, such as auxin, cytokinin, and gibberellin (Ragni et al., 2011;Ruzicka et al., 2015;Schaller et al., 2015). ...
Brassinosteroid (BR) signaling has long been reported to have an effect on xylem development, but the detailed mechanism remains unclear, especially in tree species. In this study, we find PdC3H17, which was demonstrated to mediate xylem formation driven by auxin in our previous report, is also involved in BR-promoted xylem development. Y1H analysis, EMSA, and transcription activation assay confirmed that PdC3H17 was directly targeted by PdBES1, which is a key transcriptional regulator in BR signaling. Tissue specificity expression analysis and in situ assay revealed that PdC3H17 had an overlapping expression profile with PdBES1. Hormone treatment examinations verified that xylem phenotypes in PdC3H17 transgenic plants, which were readily apparent in normal condition, were attenuated by treatment with either brassinolide or the BR biosynthesis inhibitor propiconazole. The subsequent quantitative real-time polymerase chain reaction (qRT-PCR) analyses further revealed that BR converged with PdC3H17 to influence transcription of downstream xylem-related genes. Additionally, the enhancement of xylem differentiation by auxin in PdC3H17 overexpression plants was significantly attenuated compared with wild-type and dominant negative plants due to BR deficiency, which suggested that the BR- and auxin-responsive gene PdC3H17 acted as an mediation of these two hormones to facilitate xylem development. Taken together, our results demonstrate that BR signaling converges with auxin-mediated PdC3H17 to regulate xylem formation in Populus and thus provide insight into the regulation mechanism of BRs and the crosstalk with auxin signaling on xylem formation.
... Later in development, when primary stem growth is completed, the procambium and its neighbouring interfascicular parenchyma cells (between the vascular bundles) differentiate into the cambial meristem, eventually forming a continuous cylinder of stem cells in the primary plant stem. This then initiates the process of secondary growth, during which the cambial meristem continuously generates secondary xylem usually towards the inside of the stem, and secondary phloem towards the outside of the stem, resulting in radial stem expansion (Jouannet et al., 2015). The vascular cambium is an important meristem, as it produces vascular tissues that transport water and nutrients throughout the plant body. ...
Plant secondary growth, also referred to as wood formation, includes the production of secondary xylem, which is derived from meristematic cambium cells embedded in vascular tissues. Despite the importance of secondary xylem in plant growth and wood formation, the molecular mechanism of secondary growth is not yet well understood. Here we identified an important role for the Arabidopsis thaliana (Arabidopsis) AT-HOOK MOTIF CONTAINING NUCLEAR LOCALIZED 15 (AHL15) gene, encoding for a putative transcriptional regulator, in controlling vascular cambium activity and secondary xylem formation. Secondary xylem development was significantly reduced in inflorescence stems of the Arabidopsis ahl15 lossof function mutant, whereas AHL15 overexpression led to extensive secondary xylem formation. AHL15 expression under a vascular meristem-specific promoter also enhanced the amount of interfascicular secondary xylem. Moreover, AHL15 appeared to be required for the enhanced secondary xylem formation in the Arabidopsis double loss-of-function mutant of the SUPPRESSOR OF OVEREXPRESSION OF CO 1 (SOC1) and FRUITFULL (FUL) genes. A well-known central regulator of cambial activity is the plant hormone cytokinin. We showed that the expression of two cytokinin biosynthesis genes (ISOPENTENYL TRANSERASE (IPT) 3 and 7) is decreased in ahl15 loss-of-function mutant stems, whereas the secondary xylem deficiency in these mutant stems can be resorted by cambium-specific expression of the Agrobacterium tumefaciens IPT gene, indicating that AHL15 acts through the cytokinin pathway. These findings support a model whereby AHL15 acts as a central factor inducing vascular cambium activity downstream of SOC1 and FUL and upstream of IPT3 , IPT7 and LOG4, LOG5 governing the rate of secondary xylem formation in Arabidopsis inflorescence stems.
... Lateral roots, adventitious roots, and callus all initiate from pericycle cells, a type of vascular initial [5,6]. Similarly, vascular cambium and stomatal lineage stem cells have comparable but more limited differentiation capacity [7,8]. ...
In plants and animals, self-renewing stem cell populations play fundamental roles in many developmental contexts. Plants differ from most animals in their retained ability to initiate new cycles of growth and development, which relies on the establishment and activity of branch meristems. In seed plants, branching is achieved by stem-cell-containing axillary meristems, which are initiated from a leaf axil meristematic cell population originally detached from the shoot apical meristem. It remains unclear how the meristematic cell fate is maintained. Here, we show that ARABIDOPSIS THALIANA HOMEOBOX GENE1 (ATH1) maintains the meristem marker gene SHOOT MERISTEMLESS (STM) expression in the leaf axil to enable meristematic cell fate maintenance. Furthermore, ATH1 protein interacts with STM protein to form a STM self-activation loop. Genetic and biochemical data suggest that ATH1 anchors STM to activate STM as well as other axillary meristem regulatory genes. This auto-regulation allows the STM locus to remain epigenetically active. Taken together, our findings provide a striking example of a self-activation loop that maintains the flexibility required for stem cell niche re-establishment during organogenesis.
... Therefore, although functionally equivalent, primary and secondary vascular tissues originate from different cellular environments. This raises the question to what extend molecular circuits instrumental for forming primary and secondary vascular tissues are comparable (Jouannet et al., 2014). ...
As a prerequisite for constant growth, plants produce vascular tissues at different sites within their postembryonic body. Interestingly, the formation of vascular tissues during longitudinal and radial expansion of shoot and root axes differs fundamentally with respect to its anatomical configuration. This raises the question to which level regulatory mechanisms of vascular tissue formation are shared throughout plant development. Here, we show that, similar to primary phloem formation during longitudinal growth, the cambium‐based formation of secondary phloem depends on the function of SUPPRESSOR OF MAX2 1‐LIKE (SMXL) genes. In particular, local SMXL5 deficiency results in the absence of secondary phloem. Moreover, the additional disruption of SMXL4 activity increases tissue production in the cambium region without that secondary phloem is formed. Using promoter reporter lines, we observe that SMXL4 and SMXL5 activities are associated with different stages of secondary phloem formation in the Arabidopsis stem. Based on genome‐wide transcriptional profiling and expression analyses of phloem‐related markers we conclude that early steps of phloem formation are impaired in smxl4;smxl5 double mutants and that the additional cambium‐derived cells fail to establish phloem‐related features. Our results show that molecular mechanisms determining primary and secondary phloem formation share important properties but differ slightly with SMXL5 playing a more dominant role in the formation of secondary phloem.
... Interestingly, several studies have provided evidence that maintenance and activity in shoot apical and vascular meristems are controlled by similar but different regulation loops involving transcription factors and hormones, especially CLAVATA3/EMBRYO SURROUNDING REGION-RELATED peptides (pCLE) and their receptors of the leucine-rich repeat (LRR) receptor kinase family as well as auxin supply and concentration gradient (Etchells et al., 2016;Bhalerao and Fischer, 2017;Woerlen et al., 2017;Ragni and Greb, 2018). In this comparison, the concept of the CLV-WUS feedback loop in the SAM can be partially transferred to another loop acting in the cambium where CLE41 targets and activates PHLOEM INTERCALATED WITH XYLEM/TDIF RECEPTOR (PXY/TDR), an RLK expressed in the Arabidopsis procambium (Jouannet et al., 2015). The conservation of CLE41-PXY/TDR loops among euphyllophytes supports the evidence of their fundamental role (see Ragny and Grebb (2018) for a recent review). ...
Plant architecture results from developmental processes occurring throughout the plant life span. In the current article, the processes responsible for the formation of aerial organs and their organization at the whole plant scale, i.e. primary and secondary growth, branching, and flowering are considered, whereas the below‐ground plant organization is not included. This article focuses on the advances made on the comprehension of the physiologic and genetic base of plant architecture initially elucidated in plant models, and which are progressively being deciphered in a large range of species. The current knowledge allows comparisons of the processes among groups and to examine their conservation versus their major differences. However, numerous questions remain especially when regularities in plant structure are observed at coarse scales of plant organization such as annual shoots, axes, or branching systems. Moreover, the overlap of numerous gene functions over several processes leads to pleiotropic effects that make the genetic effects complex to interpret. The evidence of such overlaps opens evo‐devo perspectives.
... In other reports, gene expression of AtSERK1 was found in the procambium of the vascular bundles in roots, hypocotyls, and inflorescence stems of Arabidopsis [10,22]. As procambium is regarded as a primary meristematic tissue, where procambium cells are differentiated as xylem and phloem cells [23], and SERK1 expression becomes confined to the procambium, it is therefore proposed that SERK1 expression marks the procambium vascular cell population to differentiate [22]. ...
Background
Plant gene homologs that control cell differentiation can be used as biotechnological tools to study the in vitro cell proliferation competence of tissue culture-recalcitrant species such as peppers. It has been demonstrated that SERK1 homologs enhance embryogenic competence when overexpressed in transformed tissues; therefore, cloning of a pepper SERK1 homolog was performed to further evaluate its biotechnological potential.
Results
A Capsicum chinense SERK full-length cDNA (CchSERK1) was cloned and characterized at the molecular level. Its deduced amino acid sequence exhibits high identity with sequences annotated as SERK1 and predicted-SERK2 homologs in the genomes of the Capsicum annuum CM-334 and Zunla-1 varieties, respectively, and with SERK1 homologs from members of the Solanaceae family. Transcription of CchSERK1 in plant tissues, measured by quantitative RT-PCR, was higher in stems, flowers, and roots but lower in leaves and floral primordia. During seed development, CchSERK1 was transcribed in all zygotic stages, with higher expression at 14 days post anthesis. During somatic embryogenesis, CchSERK1 was transcribed at all differentiation stages, with a high increment in the heart stage and lower levels at the torpedo/cotyledonal stages.
Conclusion
DNA sequence alignments and gene expression patterns suggest that CchSERK1 is the C. chinense SERK1 homolog. Significant levels of CchSERK1 transcripts were found in tissues with cell differentiation activities such as vascular axes and during the development of zygotic and somatic embryos. These results suggest that CchSERK1 might have regulatory functions in cell differentiation and could be used as a biotechnological tool to study the recalcitrance of peppers to proliferate in vitro.
How to cite: Jiménez-Guillen D, Pérez-Pascual D, Souza-Perera R, et al. Cloning and molecular characterization of a putative Habanero pepper SERK1 cDNA expressed during somatic and zygotic embryogenesis. Electron J Biotechnol 2019;41. https://doi.org/10.1016/j.ejbt.2019.07.006.
... The vascular tissues are comprised of xylem and phloem with a layer of cambial cells in between. The vascular cambium is characterized by its multipotent stem cell identity and continued proliferation and differentiation into new xylem and phloem cells during vascular development [1]. ...
Histone acetylation and deacetylation play essential roles in eukaryotic gene regulation. HD2 (HD-tuins) proteins were previously identified as plant-specific histone deacetylases. In this study, we investigated the function of the HDT1 gene in the formation of stem vascular tissue in Arabidopsis thaliana. The height and thickness of the inflorescence stems in the hdt1 mutant was lower than that of wild-type plants. Paraffin sections showed that the cell number increased compared to the wild type, while transmission electron microscopy showed that the size of individual tracheary elements and fiber cells significantly decreased in the hdt1 mutant. In addition, the cell wall thickness of tracheary elements and fiber cells increased. We also found that the lignin content in the stem of the hdt1 mutants increased compared to that of the wild type. Transcriptomic data revealed that the expression levels of many biosynthetic genes related to secondary wall components, including cellulose, lignin biosynthesis, and hormone-related genes, were altered, which may lead to the altered phenotype in vascular tissue of the hdt1 mutant. These results suggested that HDT1 is involved in development of the vascular tissue of the stem by affecting cell proliferation and differentiation.
... Given the continuity of procambium and vascular cambium, it should not be a surprise that genes involved in regulation of the latter are related to those active in the former, and that similarities in regulation exist between shoot apical meristem and vascular cambium (Schrader et al., 2004;Groover et al., 2006;Du & Groover, 2010;Agusti et al., 2011;Gursanscky et al., 2016). A feed-forward loop that involves HD-ZIP III genes, auxin, PIN and the auxin response factor MP/ARF5 is essential for the formation of procambium strands at the shoot apical meristem in Arabidopsis (Jouannet et al., 2015;M€ uller et al., 2016). These same regulatory components, carrying similar roles in the functioning or establishment of the vascular cambium in Populus, suggest that a conserved HD-ZIP III-auxin-PIN-MP/ARF5 signaling pathway is shared between procambium formation and vascular cambium establishment (Zhu et al., 2018). ...
Secondary growth from a vascular cambium, present today only in seed plants and isoetalean lycophytes, has a 400‐million‐yr evolutionary history that involves considerably broader taxonomic diversity, most of it hidden in the fossil record. Approaching vascular cambial growth as a complex developmental process, we review data from living plants and fossils that reveal diverse modes of secondary growth. These are consistent with a modular nature of secondary growth, when considered as a tracheophyte‐wide structural feature. This modular perspective identifies putative constituent developmental modules of cambial growth, for which we review developmental anatomy and regulation. Based on these data, we propose a hypothesis that explains the sources of diversity of secondary growth, considered across the entire tracheophyte clade, and opens up new avenues for exploring the origin of secondary growth. In this hypothesis, various modes of secondary growth reflect a mosaic pattern of expression of different developmental–regulatory modules among different lineages. We outline an approach that queries three information systems (living seed plants, living seed‐free plants, and fossils) and integrates data on developmental regulation, anatomy, gene evolution and phylogeny to test the mosaic modularity hypothesis and its implications, and to inform efforts aimed at understanding the evolution of secondary growth.
... The earliest indication of vascular development is the formation of procambial cells at early globular stage during embryogenesis (Mansfield and Briarty 1991). Further differentiation of procambial cells is halted during embryo maturation and resumed following germination (Cano-Delgado, Lee et al. 2010, Jouannet, Brackmann et al. 2015. In the course of embryogenesis, initiation of vascular procambial cells is controlled by hormonal and transcriptional regulators (ten Hove, Lu et al. 2015). ...
Plant vascular system is important for plant growth and development. There are three different cell types in vascular tissues, xylems, phloems and procambial or cambial. In the presence of cambium, plants continuously generate new vascular tissues. During the differentiation process, xylem fibers develop lignified secondary cell walls which provide abundant resources for biofuel industry. However, the underlying mechanisms of vascular development and secondary cell wall formation are still elusive. Our objectives are to identify new transcription factors function in vascular initiation and biomass deposition in Arabidopsis. In this thesis, we identified two activation-tagging mutants, hva-d with high vascular activity and stp-2d showing secondary wall thickening in pith cells. A novel mutant, hva-d, showed more developed vascular bundles. Histochemical analysis indicated cambium and phloem activity in hva-d were also elevated. The phenotype results from elevated expression of HVA. HVA functions as a transcriptional repressor and interacts with TPL. The results demonstrated HVA was an important regulator involved in vascular development. Another dominant mutant, stp–2d showed secondary wall thickening in pith cells (STP). Activation of microRNA 165b (miR165b) is responsible for the STP phenotype. The expression of three class III HD–ZIP transcription factor genes, including AtHB15, was repressed in the stp–2d mutant. MicroRNA-resistant mtAtHB15 complemented stp-2d defects and it indicated that repression of AtHB15 accounted for the abnormal lignification in stp-2d mutant. These results illustrated a microRNA165b-AtHB15 mediated regulatory pathway functions upstream of the secondary cell wall master regulators.
... However, the ratio of each vascular cell type in bil1 mutants was similar to that in wildtype plants ( Supplementary Fig. 3a,b). Vascular cambial activity at the stem base (immediately above the uppermost rosette leaves) is very high during the establishment of the interfascicular cambium from which the secondary xylem and phloem cells are newly produced via periclinal divisions 2,23 . To further investigate the vascular cambial activity, the lateral extension of interfascicular cambiumderived (ICD) tissue was examined at the stem base 13 . ...
Vascular cambium proliferation in plants is crucial for the generation of vascular tissues and for mechanical strength. Phytohormones and mobile peptides are key regulators of vascular cambial activity during secondary growth; however, the signalling cross-talk underlying their coordinated action is largely unknown. Here, we reveal that BIN2-LIKE 1 (BIL1), a glycogen synthase kinase 3, integrates the PHLOEM INTERCALATED WITH XYLEM/tracheary element differentiation inhibitory factor (TDIF) RECEPTOR (PXY/TDR) module into MONOPTEROS/AUXIN RESPONSE FACTOR 5 (MP/ARF5) transcription factor action during secondary growth. BIL1-mediated phosphorylation of MP/ARF5 enhances its negative effect on vascular cambial activity, which upregulates the negative regulators of cytokinin signalling ARABIDOPSIS RESPONSE REGULATOR 7 (ARR7) and ARR15. PXY/TDR inhibits BIL1 activity, which attenuates the effect of MP/ARF5 on ARR7 and ARR15 expression, thus increasing vascular cambial activity. Together, these results suggest that BIL1 is a key mediator that links peptide signalling with auxin-cytokinin signalling for the maintenance of cambial activity.
... Here, we review recent insights on stem ontogenesis and its regulation. We focus primarily on the initiation of stem tissues and on internode elongation, as secondary growth of the stem has been recently reviewed [9,10]. ...
In spite of its central importance in evolution, plant architecture and crop improvement, stem development remains poorly understood relative to other plant organs. Here, we summarise current knowledge of stem ontogenesis and its regulation, including insights from new image analysis and biophysical approaches. The stem initiates in the rib zone (RZ) of the shoot apical meristem, under transcriptional control by DELLA and BLH proteins. Links have emerged between these regulators and cell proliferation, patterning and oriented growth in the RZ. During subsequent internode elongation, cell wall properties and mechanics have been analysed in detail, revealing pectin modification as a prominent control point. Recent work has also highlighted signalling to coordinate growth of stem tissues with different mechanical properties.
For multicellular organisms to develop, cells must grow, divide, and differentiate along preferential or exclusive orientations or directions. Moreover, those orientations-or axes-and directions-or polarities-must be coordinated between cells within and between tissues. Therefore, how axes and polarities are coordinated between cells is a key question in biology. In animals, such coordination mainly depends on cell migration and direct interaction between proteins protruding from the plasma membrane. Both cell movements and direct cell-cell interactions are prevented in plants by cell walls that surround plant cells and keep them apart and in place. Therefore, plants have evolved unique mechanisms to coordinate their cell axes and polarities. Here I will discuss evidence suggesting that understanding how leaf veins form may uncover those unique mechanisms. Indeed-unlike previously thought-the cell-to-cell, polar transport of the plant hormone auxin along developing veins cannot account for many features of vein patterning. Instead, those features can be accounted for by models of vein patterning that combine polar auxin transport with auxin diffusion through plasmodesmata along the axis of developing veins. Though it remains unclear whether such a combination of polar transport and axial diffusion of auxin can account for the formation of the variety of vein patterns found in plant leaves, evidence suggests that such a combined mechanism may control plant developmental processes beyond vein patterning.
Auxin has been shown to exhibit a striking concentration gradient distribution in radial sections of angiosperm and gymnosperm species, in which peak auxin levels are concentrated in dividing cambial cells, while the absolute auxin concentration sharply declines toward developing secondary phloem and xylem regions. The coincidence of auxin concentration gradient across shoot tissues and xylem cell developmental gradient has prompted that auxin could act as “a plant morphogen” to provide a positional signal for cambial cell development. However, the specific location of vascular cambium and the lack of mutants altering auxin distribution in shoots of woody species made further verification experiments difficult to explore. To address this issue, different concentrations of exogenous IAA were applied to decapitated Chinese fir (Cunninghamia lanceolata (Lamb.) Hook.) trunks in this study to induce the change in the auxin concentration gradient in radial stems, and its effects on cambial activities were examined on the physiological, cellular and molecular levels. Our findings manifested that exogenous IAA treatments resulted in vast changes in endogenous hormone concentrations (including IAA, ZR, GA3 and ABA), cambial cell developmental behaviors and transcriptional activities of genes related to polar auxin transport (PAT), auxin signaling, the biosynthesis and signal transduction of other plant hormones and the genetic control of cambial activity. Based on above findings, we postulated a model of auxin concentration gradient involved in the control of cambial activity and secondary growth in tree trunks. In this model, the contrasting expression of AUX1/LAX and PIN family carriers in distinct Chinese fir wood-forming tissues dynamically modulates PAT into the cambial zone adjacent to the secondary phloem side and secondary xylem tissues, resulting in a sharp and wide auxin spatial gradient distribution across shoots in different stages of secondary growth, respectively. This change in auxin concentration gradient distribution in radial sections in turn acts on cambial developmental behaviors by modulating the expression of auxin signaling genes and key transcription factors and the production of other plant hormones in distinct woody tissues. Findings in this study provide important insights for understanding the biological significance of auxin concentration gradient existing in the radial stems of woody species.
Differentiation of stem cells in the plant apex gives rise to aerial tissues and organs. Presently we lack a lineage map of the shoot apex cells in woody perennials – a critical gap considering their role in determining primary and secondary growth. Here we used single-nuclei RNA sequencing to determine cell type-specific transcriptomes of the Populus vegetative shoot apex. We identified highly heterogeneous cell populations clustered into seven broad groups represented by 18 transcriptionally distinct cell clusters. Next, we established the developmental trajectories of the epidermis, leaf mesophyll, and vascular tissue. Motivated by the high similarities between Populus and Arabidopsis cell population in the vegetative apex, we applied a pipeline for interspecific single-cell gene expression data integration. We contrasted the developmental trajectories of primary phloem and xylem formation in both species, establishing the first comparison of vascular development between a model annual herbaceous and a woody perennial plant species. Our results offer a valuable resource for investigating the principles underlying cell division and differentiation conserved between herbaceous and perennial species while also allowing us to examine species-specific differences at single-cell resolution.
The leaf functions through photosynthesis and determines the yield and quality of the grape (Vitis vinifera L.). Understanding the morphology, anatomy, and molecular regulation of leaf development can provide insight into the function of the leaf. In the present study, the microstructure of the vein and stomata was observed using an improved tissue clearing method, and differently expressed transcription factors (TFs) underlying leaf development were determined by transcriptome analysis in the ‘Pinot Noir’ grape. The process of stomata development from a single epidermal cell to a mature stomata cell was observed clearly under light microscopy by improving the method of tissue clearing. The onset of stomata formation in the leaf tip occurred prior to that in the central leaf, and developing stomata were observed in the center of the leaves at different development stages except for L5, when only mature stomata were present. The major vein was initiated before leaf unfolding (L1) and numbers of vessels increased underlying leaf growth. The shape of the vessel presented tapering. Freely ending veins were not observed until stages L4 and L5, and the vessels were different from other order veins in shape and ways of connection. Development-dependent differences between stomata and vein suggested differences in function during leaf development. A total of 445 TFs involved in leaf morphology and anatomy were identified. The members of the bHLH family (SCRM) and MYB family (MYB88) can contribute to stomata patterning during leaf development. Members of the TF families of ARF (ARF5), Dof (TMO6 and HCA2), HD-Zip III (HB14), and GRAS (SCR, SCL14, and SCL28) were potential regulators involved in vein development. The grape orthologs of WOX9, TCP4, and GRF1-9 can participate in leaf outgrowth and expansion. These results will be helpful for our further study of the molecular regulation of leaf development and the relation between leaf structure and function, which can be applied to improve grape production.
The phellem is a specialized boundary tissue providing the first line of defense against abiotic and biotic stresses in organs undergoing secondary growth. Phellem cells undergo several differentiation steps, which include cell wall suberization, cell expansion and programmed cell death. Yet, the molecular players acting particularly in phellem cell differentiation remain poorly described, particularly in the widely used model plant Arabidopsis thaliana. Using specific marker lines we followed the onset and progression of phellem differentiation in A. thaliana roots, and further targeted the translatome of new developed phellem cells using Translating Ribosome Affinity Purification followed by mRNA sequencing (TRAP‐SEQ). We showed that phellem suberization is initiated early after phellogen (cork cambium) division. The specific translational landscape was organized in three main domains related to energy production, synthesis and transport of cell wall components, and response to stimulus. Novel players in phellem differentiation, related to suberin monomer transport and assembly, as well as novel transcription regulators were identified. This strategy provided an unprecedented resolution of the translatome of developing phellem cells, giving a detailed and specific view on the molecular mechanisms acting on cell differentiation in periderm tissues of the model plant Arabidopsis.
To discuss vascular differentiation, there is a need to introduce their basic structures, development, and patterns. Accordingly, major important developmental patterns that will be needed for discussion in the following chapters are presented and discussed. In addition to basic, developmental, and comparative anatomy of plants’ vascular tissues, the chapter includes many original photographs that illustrate known and new phenomena with their up-to-date explanations.
Many insect species have acquired the ability to redirect plant development to form unique organs called galls, which provide these insects with unique, enhanced food and protection from enemies and the elements. Many galls resemble flowers or fruits, suggesting that elements of reproductive development may be involved. We tested this hypothesis using RNA sequencing to quantify the transcriptional responses of wild grapevine (Vitis riparia) leaves to a galling parasite, phylloxera (Daktulosphaira vitifoliae). If development of reproductive structures is part of gall formation, we expected to find significantly elevated expression of genes involved in flower and/or fruit development in developing galls as opposed to ungalled leaves. We found that reproductive gene ontology categories were significantly enriched in developing galls, and that expression of many candidate genes involved in floral development were significantly increased, particularly in later gall stages. The patterns of gene expression found in galls suggest that phylloxera exploits vascular cambium to provide meristematic tissue and redirects leaf development towards formation of carpels. The phylloxera leaf gall appears to be phenotypically and transcriptionally similar to the carpel, due to the parasite hijacking underlying genetic machinery in the host plant.
In roots, stem cells in the cambium region form vascular tissues needed for the long-distance transport of water and nutrients. How these stem cells are specified and regulated has now been illuminated.
While plants and animals largely diverge in several major aspects, some biological functions are highly conserved between these kingdoms. During organism development, the correct implementation of organs with unique shape, size and function is the result of coordinated cellular processes as cell proliferation and expansion. Translationally Controlled Tumor Protein (TCTP) is a highly conserved protein among all eukaryotes. TCTP mutation leads to embryo lethality, indicating that it is mandatory for organism development. Moreover, it has been shown that TCTP controls organ growth by regulating the G1/S transition and cell cycle progression both in plants and animals. In animals, the molecular pathways by which TCTP controls cell proliferation are well known. However, in plants, the mechanism implicating TCTP in the control of development and cell cycle is less understood. To better understand how TCTP controls cell proliferation and development in Arabidopsis thaliana, the putative TCTP interactors were identified. Among them, CSN4, a subunit of the COP9 signalosome complex (CSN) known to be involved in the control of CULLINS (CUL) neddylation status and CULLIN-RING ubiquitin ligases (CRLs) activity, was identified. Through their ubiquitination activity, CRL complexes are known to control the accumulation of mains cell cycle regulators as Cyclins or Kip Related Proteins. Thus, during my PhD, I studied the interaction between TCTP and CSN4, in order to evaluate if CSN complex could link TCTP to cell cycle control. I used genetic, cellular and biochemical approaches to demonstrate that TCTP and CSN4 interact in the cytoplasm. Phenotypic characterization of plants and cell cultures down- or overexpressing these genes demonstrated that TCTP and CSN4 interact genetically to control G1/S transition. In order to understand if the interaction between these two proteins could interfere with the CSN complex function, I characterized CUL1 neddylation status in transgenic lines misexpressing TCTP and CSN4. Loss of function of TCTP increases the non-neddylated CUL1 fraction, while overexpression of TCTP increases neddylated CUL1 form. These data show that TCTP interferes with the role of CSN complex in regulating CUL1 neddylation. Accordingly, our data suggest that TCTP controls cell cycle progression via controlling CSN deneddylation activity, and thus influencing CRL activity. In the last part of my PhD, I addressed if this role of TCTP is conserved in animals. I used biochemical approach to evaluate CUL1 neddylation in Drosophila melanogaster downregulated for dTCTP. My data show that Drosophila larvae knockdown for dTCTP also leads to an increase of non-neddylated CUL1 fraction. These data suggest that the mechanism by which TCTP/CSN4 regulate cell cycle, is likely conserved between Arabidopsis and Drosophila
Coordination of cell division and pattern formation is central to tissue and organ development, particularly in plants where
walls prevent cell migration. Auxin and cytokinin are both critical for division and patterning, but it is unknown how these
hormones converge upon tissue development. We identify a genetic network that reinforces an early embryonic bias in auxin
distribution to create a local, nonresponding cytokinin source within the root vascular tissue. Experimental and theoretical
evidence shows that these cells act as a tissue organizer by positioning the domain of oriented cell divisions. We further
demonstrate that the auxin-cytokinin interaction acts as a spatial incoherent feed-forward loop, which is essential to generate
distinct hormonal response zones, thus establishing a stable pattern within a growing vascular tissue.
The role of auxin as main regulator of vascular differentiation is well established, and a direct correlation between the rate of xylem differentiation and the amount of auxin reaching the (pro)cambial cells has been proposed. It has been suggested that thermospermine produced by ACAULIS5 (ACL5) and BUSHY AND DWARF2 (BUD2), is one of the factors downstream to auxin contributing to the regulation of this process in Arabidopsis. Here, we provide an in-depth characterization of the mechanism through which ACL5 modulates xylem differentiation. We show that an increased level of ACL5 slows down xylem differentiation by negatively affecting the expression of homeodomain-leucine zipper (HD-ZIP) III and key auxin signalling genes. This mechanism involves the positive regulation of thermospermine biosynthesis by the HD-ZIP III protein ARABIDOPSIS THALIANA HOMEOBOX8 tightly controlling the expression of ACL5 and BUD2. In addition, we show that the HD-ZIP III protein REVOLUTA contributes to the increased leaf vascularization and long hypocotyl phenotype of acl5 likely by a direct regulation of auxin signalling genes such as LIKE AUXIN RESISTANT2 (LAX2) and LAX3. We propose that proper formation and differentiation of xylem depends on a balance between positive and negative feedback loops operating through HD-ZIP III genes.
Development of cambium and its activity is important for our knowledge of the mechanism of secondary growth. Arabidopsis thaliana emerges as a good model plant for such a kind of study. Thus, this paper reports on cellular events taking place in the interfascicular regions of inflorescence stems of A. thaliana, leading to the development of interfascicular cambium from differentiated interfascicular parenchyma cells (IPC). These events are as follows: appearance of auxin accumulation, PIN1 gene expression, polar PIN1 protein localization in the basal plasma membrane and periclinal divisions. Distribution of auxin was observed to be higher in differentiating into cambium parenchyma cells compared to cells within the pith and cortex. Expression of PIN1 in IPC was always preceded by auxin accumulation. Basal localization of PIN1 was already established in the cells prior to their periclinal division. These cellular events initiated within parenchyma cells adjacent to the vascular bundles and successively extended from that point towards the middle region of the interfascicular area, located between neighboring vascular bundles. The final consequence of which was the closure of the cambial ring within the stem. Changes in the chemical composition of IPC walls were also detected and included changes of pectic epitopes, xyloglucans (XG) and extensins rich in hydroxyproline (HRGPs). In summary, results presented in this paper describe interfascicular cambium ontogenesis in terms of successive cellular events in the interfascicular regions of inflorescence stems of Arabidopsis.
Significance
The vascular tissues form a continuous network providing the long-distance transport of water and nutrients in all higher plants (tracheophytes). To incorporate separate organs into this network, it is essential that the position of different vascular cell types is tightly regulated. Several factors required for root vascular patterning (including hormones and gene products) have previously been identified in the model plant Arabidopsis . We have now established a mathematical model formulizing the interaction between these factors, allowing us to identify a minimal regulatory network capable of maintaining a stable vascular pattern in Arabidopsis roots. We envisage that this model will help future researchers understand how similar regulatory units can be applied to create alternative patterns in other species.
The phytohormone auxin is a key developmental signal in plants. To date, only auxin perception has been described to trigger the release of transcription factors termed AUXIN RESPONSE FACTORs (ARFs) from their AUXIN/INDOLE-3-ACETIC ACID (AUX/IAA) repressor proteins. Here, we show that phosphorylation of ARF7 and ARF19 via BRASSINOSTEROID-INSENSITIVE2 (BIN2) can also potentiates auxin signalling output during lateral root organogenesis. BIN2-mediated phosphorylation of ARF7 and ARF19 suppresses their interaction with AUX/IAAs, and subsequently enhances the transcriptional activity to their target genes LATERAL ORGAN BOUNDARIES-DOMAIN16 (LBD16) and LBD29. In this context, BIN2 is under the control of the TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FACTOR (TDIF) - TDIF RECEPTOR (TDR) module. TDIF-initiated TDR signalling directly acts on BIN2-mediated ARF phosphorylation, leading to the regulation of auxin signalling during lateral root development. In summary, this study delineates a TDIF-TDR-BIN2 signalling cascade controlling auxin perception-independent regulation of ARF and AUX/IAA interaction during lateral root development.
Plant vasculatures are complex tissues consisting of (pro)cambium, phloem, and xylem. The (pro)cambium serves as vascular
stem cells that produce all vascular cells. The Arabidopsis ERECTA (ER) receptor kinase is known to regulate the architecture of inflorescence stems. It was recently reported that the
er mutation enhances a vascular phenotype induced by a mutation of TDR/PXY, which plays a significant role in procambial proliferation, suggesting that ER participates in vascular development. However, detailed molecular mechanisms of the ER-dependent vascular regulation are largely unknown. Here, this work found that ER and its paralogue, ER-LIKE1, were redundantly involved in procambial development of inflorescence stems. Interestingly, their activity in the phloem
was sufficient for vascular regulation. Furthermore, two endodermis-derived peptide hormones, EPFL4 and EPFL6, were redundantly
involved in such regulation. It has been previously reported that EPFL4 and EPFL6 act as ligands of phloem-expressed ER for
stem elongation. Therefore, these findings indicate that cell–cell communication between the endodermis and the phloem plays
an important role in procambial development as well as stem elongation. Interestingly, similar EPFL–ER modules control two
distinct developmental events by slightly changing their components: the EPFL4/6–ER module for stem elongation and the EPFL4/6–ER/ERL1
module for vascular development.
Polyamines are small polycationic amines widespread in living organisms. Thermospermine, synthesized by thermospermine synthase ACAULIS5 (ACL5), was recently shown to be an endogenous plant polyamine. Thermospermine is critical for proper vascular development and xylem cell specification but it is not known how thermospermine homeostasis is controlled in the xylem. We present data in the Populus model system supporting presence of a negative feedback control of thermospermine levels in stem xylem tissues, the main site of thermospermine biosynthesis. While overexpression of the ACL5 homolog in Populus, POPACAULIS5, resulted in strong upregulation of ACL5 expression and thermospermine accumulation in leaves, the corresponding levels in the secondary xylem tissues of the stem were similar or lower than those in the wild-type. POPACAULIS5 overexpression had a negative effect on accumulation of indole-3-acetic acid (IAA), while exogenous auxin had a positive effect on POPACAULIS5 expression, thus promoting thermospermine accumulation. Further, overexpression of POPACAULIS5 negatively affected Class III homeodomain-leucine zipper (HD-Zip III) transcription factor PttHB8, a homolog of AtHB8, while upregulation of PttHB8 positively affected POPACAULIS5 expression. These results support that excessive accumulation of thermospermine is prevented by a negative feedback control of POPACAULIS5 transcript levels through suppression of IAA levels, and that PttHB8 is involved in the control of POPACAULIS5 expression. We propose that this negative feedback loop functions to maintain steady state levels of thermospermine, required for proper xylem development, and that it is dependent on the presence of high concentrations of endogenous IAA, such as those present in the secondary xylem tissues. This article is protected by copyright. All rights reserved.
In plants, the cambium and procambium are meristems from which vascular tissue is derived. In contrast to most plant cells, stem cells within these tissues are thin and extremely long. They are particularly unusual as they divide down their long axis in a highly ordered manner, parallel to the tangential axis of the stem. CLAVATA3-LIKE/ESR-RELATED 41 (CLE41) and PHLOEM INTERCALATED WITH XYLEM (PXY) are a multifunctional ligand-receptor pair that regulate vascular cell division, vascular organisation and xylem differentiation in vascular tissue. A transcription factor gene, WUSCHEL HOMEOBOX RELATED 4 (WOX4) has been shown to act downstream of PXY. Here we show that WOX4 acts redundantly with WOX14 in the regulation of vascular cell division, but that these genes have no function in regulating vascular organisation. Furthermore, we identify an interaction between PXY and the receptor kinase ERECTA (ER) that affects the organisation of the vascular tissue but not the rate of cell division, suggesting that cell division and vascular organisation are genetically separable. Our observations also support a model whereby tissue organisation and cell division are integrated via PXY and ER signalling, which together coordinate development of different cell types that are essential for normal stem formation.
The formation of leaf vein patterns has fascinated biologists for centuries. Transport of the plant signal auxin has long been implicated in vein patterning, but molecular details have remained unclear. Varied evidence suggests a central role for the plasma-membrane (PM)-localized PIN-FORMED1 (PIN1) intercellular auxin transporter of Arabidopsis thaliana in auxin-transport-dependent vein patterning. However, in contrast to the severe vein-pattern defects induced by auxin transport inhibitors, pin1 mutant leaves have only mild vein-pattern defects. These defects have been interpreted as evidence of redundancy between PIN1 and the other four PM-localized PIN proteins in vein patterning, redundancy that underlies many developmental processes. By contrast, we show here that vein patterning in the Arabidopsis leaf is controlled by two distinct and convergent auxin-transport pathways: intercellular auxin transport mediated by PM-localized PIN1 and intracellular auxin transport mediated by the evolutionarily older, endoplasmic-reticulum-localized PIN6, PIN8, and PIN5. PIN6 and PIN8 are expressed, as PIN1 and PIN5, at sites of vein formation. pin6 synthetically enhances pin1 vein-pattern defects, and pin8 quantitatively enhances pin1pin6 vein-pattern defects. Function of PIN6 is necessary, redundantly with that of PIN8, and sufficient to control auxin response levels, PIN1 expression, and vein network formation; and the vein pattern defects induced by ectopic PIN6 expression are mimicked by ectopic PIN8 expression. Finally, vein patterning functions of PIN6 and PIN8 are antagonized by PIN5 function. Our data define a new level of control of vein patterning, one with repercussions on other patterning processes in the plant, and suggest a mechanism to select cell files specialized for vascular function that predates evolution of PM-localized PIN proteins.
The plant vascular system, composed of xylem and phloem, evolved to connect plant organs and transport various molecules between them. During the post-embryonic growth, these conductive tissues constitutively form from cells that are derived from a lateral meristem, commonly called procambium and cambium. Procambium/cambium contains pluripotent stem cells and provides a microenvironment that maintains the stem cell population. Because vascular plants continue to form new tissues and organs throughout their life cycle, the formation and maintenance of stem cells are crucial for plant growth and development. In this decade, there has been considerable progress in understanding the molecular control of the organization and maintenance of stem cells in vascular plants. Noticeable advance has been made in elucidating the role of transcription factors and major plant hormones in stem cell maintenance and vascular tissue differentiation. These studies suggest the shared regulatory mechanisms among various types of plant stem cell pools. In this review, we focus on two aspects of stem cell function in the vascular cambium, cell proliferation and cell differentiation.
The procambium and cambium are meristematic tissues from which vascular tissue is derived. Vascular initials differentiate into phloem towards the outside of the stem and xylem towards the inside. A small peptide derived from CLV-3/ESR1-LIKE 41 (CLE41) is thought to promote cell divisions in vascular meristems by signalling through the PHLOEM INTERCALLATED WITH XYLEM (PXY) receptor kinase. pxy mutants, however, display only small reductions in vascular cell number, suggesting a mechanism exists that allows plants to compensate for the absence of PXY. Consistent with this idea, we identify a large number of genes specifically upregulated in pxy mutants, including several AP2/ERF transcription factors. These transcription factors are required for normal cell division in the cambium and procambium. These same transcription factors are also upregulated by ethylene and in ethylene-overproducing eto1 mutants. eto1 mutants also exhibit an increase in vascular cell division that is dependent upon the function of at least 2 of these ERF genes. Furthermore, blocking ethylene signalling using a variety of ethylene insensitive mutants such as ein2 enhances the cell division defect of pxy. Our results suggest that these factors define a novel pathway that acts in parallel to PXY/CLE41 to regulate cell division in developing vascular tissue. We propose a model whereby vascular cell division is regulated both by PXY signalling and ethylene/ERF signalling. Under normal circumstances, however, PXY signalling acts to repress the ethylene/ERF pathway.
Multicellular organisms achieve final body shape and size by coordinating cell proliferation, expansion, and differentiation. Loss of function in the Arabidopsis ERECTA (ER) receptor-kinase gene confers characteristic compact inflorescence architecture, but its underlying signaling pathways remain unknown. Here we report that the expression of ER in the phloem is sufficient to rescue compact er inflorescences. We further identified two Epidermal Patterning Factor-like (EPFL) secreted peptide genes, EPFL4 and EPFL6/CHALLAH (CHAL), as redundant, upstream components of ER-mediated inflorescence growth. The expression of EPFL4 or EPFL6 in the endodermis, a layer adjacent to phloem, is sufficient to rescue the er-like inflorescence of epfl4 epfl6 plants. EPFL4 and EPFL6 physically associate with ER in planta. Finally, transcriptome analysis of er and epfl4 epfl6 revealed a potential downstream component as well as a role for plant hormones in EPFL4/6- and ER-mediated inflorescence growth. Our results suggest that intercell layer communication between the endodermis and phloem mediated by peptide ligands and a receptor kinase coordinates proper inflorescence architecture in Arabidopsis.
In the shoot pole of Arabidopsis embryos, radial symmetry is broken by cotyledon specification. Subsequently, the radial pattern of the embryo axis is converted to bisymmetric. In a recent publication, we showed that distinct boundaries of hormonal signalling output specify the vascular pattern in the root meristem through a mutually inhibitory feedback loop between the hormones auxin and cytokinin. We observed that during embryogenesis, symmetry breakage in the root pole coincided with an influx of auxin from the cotyledons. In this manuscript, we provide genetic data to support the role of the cotyledons in initiating symmetry breaking in the embryonic root pole. Mutants with alterations in cotyledon number fail to establish bisymmetry in the embryo axis. These data further support the idea that input from the cotyledons may be required for the propagation of bisymmetry from the cotyledons to the embryonic root.
Developmental and physiological studies of roots are frequently limited to a post-germination stage. In Arabidopsis, a developmental change in the root meristem architecture during plant ontogenesis has not previously been studied and is addressed presently. Arabidopsis thaliana have closed root apical organization, in which all cell file lineages connect directly to one of three distinct initial tiers. The root meristem organization is dynamic and changes as the root ages from 1 to 4 wk post-germination. During the ontogeny of the root, the number of cells within the root apical meristem (RAM) increases and then decreases due to changes in the number of cortical layers and number of cell files within a central cylinder. The architecture of the initial tiers also changes as the root meristem ages. Included in the RAM's ontogeny is a pattern associated with the periclinal divisions that give rise to the middle cortex and endodermis; the three-dimensional arrangement of periclinally dividing derivative cells resembles one gyre of a helix. Four- or 5-wk-old roots exhibit a disorganized array of vacuolated initial cells that are a manifestation of the determinate nature of the meristem. Vascular cambium is formed via coordinated divisions of vascular parenchyma and pericycle cells. The phellogen is the last meristem to complete its development, and it is derived from pericycle cells that delineate the outer boundary of the root.
Cell-to-cell communication is crucial for the development of multicellular organisms, especially during the generation of new tissues and organs. Secondary growth--the lateral expansion of plant growth axes--is a highly dynamic process that depends on the activity of the cambium. The cambium is a stem cell-like tissue whose activity is responsible for wood production and, thus, for the establishment of extended shoot and root systems. Attempts to study cambium regulation at the molecular level have been hampered by the limitations of performing genetic analyses in trees and by the difficulty of accessing this tissue in model systems such as Arabidopsis thaliana. Here, we describe the roles of two receptor-like kinases, REDUCED IN LATERAL GROWTH1 (RUL1) and MORE LATERAL GROWTH1 (MOL1), as opposing regulators of cambium activity. Their identification was facilitated by a novel in vitro system in which cambium formation is induced in isolated Arabidopsis stem fragments. By combining this system with laser capture microdissection, we characterized transcriptome remodeling in a tissue- and stage-specific manner and identified series of genes induced during different phases of cambium formation. In summary, we provide a means for investigating cambium regulation in unprecedented depth and present two signaling components that control a process responsible for the accumulation of a large proportion of terrestrial biomass.
Understanding the fate and dynamics of cells during callus formation is essential to understanding totipotency and the mechanisms of somatic embryogenesis. Here, the fate of leaf explant cells during the development of embryogenic callus was investigated in the model legume Medicago truncatula.
Callus development was examined from cultured leaf explants of the highly regenerable genotype Jemalong 2HA (2HA) and from mesophyll protoplasts of 2HA and wild-type Jemalong. Callus development was studied by histology, manipulation of the culture system, detection of early production of reactive oxygen species and visualization of SERK1 (SOMATIC EMBRYO RECEPTOR KINASE1) gene expression.
Callus formation in leaf explants initiates at the cut surface and within veins of the explant. The ontogeny of callus development is dominated by the division and differentiation of cells derived from pluripotent procambial cells and from dedifferentiated mesophyll cells. Procambium-derived cells differentiated into vascular tissue and rarely formed somatic embryos, whereas dedifferentiated mesophyll cells were competent to form somatic embryos. Interestingly, explants incubated adaxial-side down had substantially less cell proliferation associated with veins yet produced similar numbers of somatic embryos to explants incubated abaxial-side down. Somatic embryos mostly formed on the explant surface originally in contact with the medium, while in protoplast microcalli, somatic embryos only fully developed once at the surface of the callus. Mesophyll protoplasts of 2HA formed embryogenic callus while Jemalong mesophyll protoplasts produced callus rich in vasculature.
The ontogeny of embryogenic callus in M. truncatula relates to explant orientation and is driven by the dynamics of pluripotent procambial cells, which proliferate and differentiate into vasculature. The ontogeny is also related to de-differentiated mesophyll cells that acquire totipotency and form the majority of embryos. This contrasts with other species where totipotent embryo-forming initials mostly originate from procambial cells.
A key question in developmental biology is how cells exchange positional information for proper patterning during organ development. In plant roots the radial tissue organization is highly conserved with a central vascular cylinder in which two water conducting cell types, protoxylem and metaxylem, are patterned centripetally. We show that this patterning occurs through crosstalk between the vascular cylinder and the surrounding endodermis mediated by cell-to-cell movement of a transcription factor in one direction and microRNAs in the other. SHORT ROOT, produced in the vascular cylinder, moves into the endodermis to activate SCARECROW. Together these transcription factors activate MIR165a and MIR166b. Endodermally produced microRNA165/6 then acts to degrade its target mRNAs encoding class III homeodomain-leucine zipper transcription factors in the endodermis and stele periphery. The resulting differential distribution of target mRNA in the vascular cylinder determines xylem cell types in a dosage-dependent manner.
Brassinosteroids (BRs) are growth-promoting steroid hormones in plants. Genetic studies in Arabidopsis illustrated the essential roles of BRs in a wide range of developmental processes and helped identify many genes involved in BR biosynthesis and signal transduction. Recently, proteomic studies identified missing links. Together, these approaches established the BR signal transduction cascade, which includes BR perception by the BRI1 receptor kinase at the cell surface, activation of BRI1/BAK1 kinase complex by transphosphorylation, subsequent phosphorylation of the BSK kinases, activation of the BSU1 phosphatase, dephosphorylation and inactivation of the BIN2 kinase, and accumulation of unphosphorylated BZR transcription factors in the nucleus. Mass spectrometric analyses are providing detailed information on the phosphorylation events involved in each step of signal relay. Thus, the BR signaling pathway provides a paradigm for understanding receptor kinase-mediated signal transduction as well as tools for the genetic improvement of the productivity of crop plants.
Class III HD-ZIP and KANADI gene family members have complementary expression patterns in the vasculature and their gain-of-function and loss-of-function mutants have complementary vascular phenotypes. This suggests that members of the two gene families are involved in the establishment of the spatial arrangement of phloem, cambium and xylem. In this study, we have investigated the role of these two gene families in vascular tissue differentiation, in particular their interactions with the plant hormone auxin. We have analyzed the vasculature of plants that have altered expression levels of Class III HD-ZIP and KANADI transcription factors in provascular cells. Removal of either KANADI or Class III HD-ZIP expression in procambium cells led to a wider distribution of auxin in internal tissues, to an excess of procambium cell recruitment and to increased cambium activity. Ectopic expression of KANADI1 in provascular cells inhibited procambium cell recruitment due to negative effects of KANADI1 on expression and polar localization of the auxin efflux-associated protein PIN-FORMED1. Ectopic expression of Class III HD-ZIP genes promoted xylem differentiation. We propose that Class III HD-ZIP and KANADI transcription factors control cambium activity: KANADI proteins by acting on auxin transport, and Class III HD-ZIP proteins by promoting axial cell elongation and xylem differentiation.
Controlling the orientation of cell division is fundamental to the development of complex body plans. This is particularly apparent in plants, where development is determined by differential growth that results solely from changes in cell expansion and orientation of the cell division plane. Despite the fundamental importance of cell division orientation to plant development, the mechanisms regulating this process remain almost completely unknown. During vascular development, the meristematic cambial cells divide down their long axis in a highly orientated manner to generate clear files of cells. The receptor kinase PXY has previously be shown to be essential for this orientation. Here, we demonstrate that the division plane is determined by the interactions of PXY and its peptide ligand, CLE41. PXY is expressed within dividing meristematic cells of the procambium, whereas CLE41 localises to the adjacent phloem cells. Altering the pattern of CLE41 expression leads to a loss of cell division orientation and a dramatic loss of ordered vascular tissue development. By contrast, increasing phloem-specific expression of CLE41 results in more cell divisions, but the orientation of cell division is retained, leading to both increased and well-ordered vascular development. We demonstrate that PXY signalling is down-regulated by CLE41. This feedback mechanism is crucial in integrating the different roles of PXY signalling in controlling xylem differentiation, regulating the rate of vascular cell division and determining the orientation of cell division. Parallels with animal systems indicate that localised signalling from adjacent cells is a general mechanism for defining the plane of cell division.
Plant shoot organs arise from initial cells that are recruited from meristematic tissues. Previous studies have shown that members of the WUSCHEL-related HOMEOBOX (WOX) gene family function to organize various initial cell populations during plant development. The function of the WOX4 gene is previously undescribed in any plant species. Comparative analyses of WOX4 transcription and function are presented in Arabidopsis (Arabidopsis thaliana), a simple-leafed plant with collateral vasculature, and in tomato (Solanum lycopersicum), a dissected-leafed species with bicollateral venation. WOX4 is transcribed in the developing vascular bundles of root and shoot lateral organs in both Arabidopsis and tomato. RNA interference-induced down-regulation of WOX4 in Arabidopsis generated small plants whose vascular bundles accumulated undifferentiated ground tissue and exhibited severe reductions in differentiated xylem and phloem. In situ hybridization analyses of Atwox4-RNA interference plants revealed delayed and reduced expression of both the phloem developmental marker ALTERED PHLOEM1 and HOMEOBOX GENE8, a marker of the vascular procambium. Overexpression of SlWOX4 correlated with overproliferation of xylem and phloem in transgenic tomato seedlings. The cumulative data suggest that the conserved WOX4 function is to promote differentiation and/or maintenance of the vascular procambium, the initial cells of the developing vasculature.
The principles underlying the formation of veins in the leaf have long intrigued developmental biologists. In Arabidopsis leaves, files of anatomically inconspicuous subepidermal cells that will elongate into vein-forming procambial cells selectively activate ATHB8 gene expression. The biological role of ATHB8 in vein formation and the molecular events that culminate in acquisition of the ATHB8 preprocambial cell state are unknown, but intertwined pathways of auxin transport and signal transduction have been implicated in defining paths of vascular strand differentiation. Here we show that ATHB8 is required to stabilize preprocambial cell specification against auxin transport perturbations, to restrict preprocambial cell state acquisition to narrow fields and to coordinate procambium formation within and between veins. We further show that ATHB8 expression at preprocambial stages is directly and positively controlled by the auxin-response transcription factor MONOPTEROS (MP) through an auxin-response element in the ATHB8 promoter. We finally show that the consequences of loss of ATHB8 function for vein formation are masked by MP activity. Our observations define, at the molecular level, patterning inputs of auxin signaling in vein formation.
The development and activity of the procambium and cambium, which ensure vascular tissue formation, is critical for overall plant architecture and growth. However, little is known about the molecular factors affecting the activity of vascular meristems and vascular tissue formation. Here, we show that the His kinase CYTOKININ-INDEPENDENT1 (CKI1) and the cytokinin receptors ARABIDOPSIS HISTIDINE KINASE2 (AHK2) and AHK3 are important regulators of vascular tissue development in Arabidopsis thaliana shoots. Genetic modifications of CKI1 activity in Arabidopsis cause dysfunction of the two-component signaling pathway and defects in procambial cell maintenance. CKI1 overexpression in protoplasts leads to cytokinin-independent activation of the two-component phosphorelay, and intracellular domains are responsible for the cytokinin-independent activity of CKI1. CKI1 expression is observed in vascular tissues of inflorescence stems, and CKI1 forms homodimers both in vitro and in planta. Loss-of-function ahk2 and ahk3 mutants and plants with reduced levels of endogenous cytokinins show defects in procambium proliferation and an absence of secondary growth. CKI1 overexpression partially rescues ahk2 ahk3 phenotypes in vascular tissue, while the negative mutation CKI1H405Q further accentuates mutant phenotypes. These results indicate that the cytokinin-independent activity of CKI1 and cytokinin-induced AHK2 and AHK3 are important for vascular bundle formation in Arabidopsis.
The roots and stems of dicotyledonous plants thicken by the cell proliferation in the cambium. Cambial proliferation changes in response to environmental factors; however, the molecular mechanisms that regulate cambial activity are largely unknown. The quadruple Arabidopsis thaliana mutant atipt1;3;5;7, in which 4 genes encoding cytokinin biosynthetic isopentenyltransferases are disrupted by T-DNA insertion, was unable to form cambium and showed reduced thickening of the root and stem. The atipt3 single mutant, which has moderately decreased levels of cytokinins, exhibited decreased root thickening without any other recognizable morphological changes. Addition of exogenously supplied cytokinins to atipt1;3;5;7 reactivated the cambium in a dose-dependent manner. When an atipt1;3;5;7 shoot scion was grafted onto WT root stock, both the root and shoot grew normally and trans-zeatin-type (tZ-type) cytokinins in the shoot were restored to WT levels, but isopentenyladenine-type cytokinins in the shoot remained unchanged. Conversely, when a WT shoot was grafted onto an atipt1;3;5;7 root, both the root and shoot grew normally and isopentenyladenine-type cytokinins in the root were restored to WT levels, but tZ-type cytokinins were only partially restored. Collectively, it can be concluded that cytokinins are important regulators of cambium development and that production of cytokinins in either the root or shoot is sufficient for normal development of both the root and shoot.
• cambium
• isopentenyladenine
• phytohormone
• zeatin
The vascular tissues of flowering plants form networks of interconnected cells throughout the plant body. The molecular mechanisms directing the routes of vascular strands and ensuring tissue continuity within the vascular system are not known, but are likely to depend on general cues directing plant cell orientation along the apical-basal axis. Mutations in the Arabidopsis gene MONOPTEROS (MP) interfere with the formation of vascular strands at all stages and also with the initiation of the body axis in the early embryo. Here we report the isolation of the MP gene by positional cloning. The predicted protein product contains functional nuclear localization sequences and a DNA binding domain highly similar to a domain shown to bind to control elements of auxin inducible promoters. During embryogenesis, as well as organ development, MP is initially expressed in broad domains that become gradually confined towards the vascular tissues. These observations suggest that the MP gene has an early function in the establishment of vascular and body patterns in embryonic and post-embryonic development.
Higher organisms possess mechanisms to maintain stem cells’ proliferative and pluripotent states in stem cell niches [1]. Plants possess two types of stem cell niches in the root and shoot apical meristems, where regulatory interactions exist between stem cells and organizing cells. Recent studies provided new insights into the molecular mechanism of stem cell maintenance [2, 3 and 4]. However, earlier and more essential developmental events such as the acquisition of stem cell proliferative activity are still unknown. In vascular tissues, procambial cells function as stem cells and differentiate into xylem, phloem, and procambium. Procambial cell proliferation starts at root apical meristem (RAM) postembryonically; therefore, procambial cell development in RAM is a good model for investigating the regulation of stem cell proliferation. LONESOME HIGHWAY (LHW) and TARGET OF MONOPTEROS5 (TMO5), as well as its homolog, TMO5-LIKE1 (T5L1), encode bHLH proteins that function as heterodimers (LHW-TMO5 and LHW-T5L1) in vascular tissue organization [ 5, 6 and 7]. LHW-T5L1 promotes vascular-cell-specific proliferation in RAM [ 7]. Here, we demonstrate that LHW-T5L1 promotes expression of key cytokinin production genes, including LONELY GUY3 (LOG3) and LOG4, in xylem precursor cells, resulting in elevated cytokinin levels in the surrounding cells. LHW-T5L1 can also promote expression of AHP6, which suppresses cytokinin signaling and then maintains xylem precursor cells at a nondividing state. Our results indicate that LHW-T5L1 establishes xylem precursor cells as a signal center that promotes procambial-cell-specific proliferation through cytokinin response.
Formative cell divisions are critical for multicellular patterning. In the early plant embryo, such divisions follow from orienting the division plane. A major unanswered question is how division plane orientation is genetically controlled, and in particular whether this relates to cell geometry. We have generated a complete 4D map of early Arabidopsis embryogenesis and used computational analysis to demonstrate that several divisions follow a rule that uses the smallest wall area going through the center of the cell. In other cases, however, cell division clearly deviates from this rule, which invariably leads to asymmetric cell division. By analyzing mutant embryos and through targeted genetic perturbation, we show that response to the hormone auxin triggers a deviation from the "shortest wall" rule. Our work demonstrates that a simple default rule couples division orientation to cell geometry in the embryo and that genetic regulation can create patterns by overriding the default rule.
During plant radial growth typically seen in trees, procambial and cambial cells act as meristematic cells in the vascular system to self-proliferate and differentiate into xylem cells. These two processes are regulated by a signalling pathway composed of a peptide ligand and its receptor; tracheary element differentiation inhibitory factor (TDIF) and TDIF RECEPTOR (TDR). Here we show that glycogen synthase kinase 3 proteins (GSK3s) are crucial downstream components of the TDIF signalling pathway suppressing xylem differentiation from procambial cells. TDR interacts with GSK3s at the plasma membrane and activates GSK3s in a TDIF-dependent fashion. Consistently, a specific inhibitor of plant GSK3s strongly induces xylem cell differentiation through BRI1-EMS SUPPRESSOR 1 (BES1), a well-known target transcription factor of GSK3s. Our findings provide insight into the regulation of cell fate determination in meristem maintenance.
Plants can grow complex and elaborate structures, in some species for thousands of years. Despite the diversity in form and shape, plants are built from a limited number of fundamental tissue types, and their arrangement is deeply conserved in the plant kingdom. A key question in biology is how these fundamental tissues, i.e. epidermal, ground and vascular tissue, are specified and organized in time and space. In the present paper, I discuss the use of the early Arabidopsis embryo as a model system to dissect the control of tissue formation and patterning, as well as the specification of the stem cells that sustain post-embryonic growth. I present recent insights into the molecules and mechanisms that control both the specification and the subsequent growth of the different cell types within the embryonic root. Finally, I discuss major unanswered questions and future challenges in using the embryo as a model to decipher the regulatory logic of plant development.
How biological systems generate reproducible patterns with high precision is a central question in science. The shoot apical meristem (SAM), a specialized tissue producing plant aerial organs, is a developmental system of choice to address this question. Organs are periodically initiated at the SAM at specific spatial positions and this spatiotemporal pattern defines phyllotaxis. Accumulation of the plant hormone auxin triggers organ initiation, whereas auxin depletion around organs generates inhibitory fields that are thought to be sufficient to maintain these patterns and their dynamics. Here we show that another type of hormone-based inhibitory fields, generated directly downstream of auxin by intercellular movement of the cytokinin signalling inhibitor ARABIDOPSIS HISTIDINE PHOSPHOTRANSFER PROTEIN 6 (AHP6), is involved in regulating phyllotactic patterns. We demonstrate that AHP6-based fields establish patterns of cytokinin signalling in the meristem that contribute to the robustness of phyllotaxis by imposing a temporal sequence on organ initiation. Our findings indicate that not one but two distinct hormone-based fields may be required for achieving temporal precision during formation of reiterative structures at the SAM, thus indicating an original mechanism for providing robustness to a dynamic developmental system.
Using adaptive control techniques, we investigate the multi-switching synchronization of chaotic systems with parameters unknown.
Based upon the Lyapunov stability theory, we design the controllers and updating laws of different switching, and it is extended
to investigate the synchronization problems with different combinations of slave states with master systems. We take the Lorenz
system and the Chen system as an example to analyze the multi-switching synchronization process of different structures of
chaotic systems. Finally, numerical simulations have shown the effectiveness of the method.
Plant vascular cells are joined end to end along uninterrupted lines to connect shoot organs with roots; vascular strands are thus polar, continuous and internally aligned. What controls the formation of vascular strands with these properties? The "auxin canalization hypothesis"-based on positive feedback between auxin flow through a cell and the cell's capacity for auxin transport-predicts the selection of continuous files of cells that transport auxin polarly, thus accounting for the polarity and continuity of vascular strands. By contrast, polar, continuous auxin transport-though required-is insufficient to promote internal alignment of vascular strands, implicating additional factors. The auxin canalization hypothesis was derived from the response of mature tissue to auxin application but is consistent with molecular and cellular events in embryo axis formation and shoot organ development. Objections to the hypothesis have been raised based on vascular organizations in callus tissue and shoot organs but seem unsupported by available evidence. Other objections call instead for further research; yet the inductive and orienting influence of auxin on continuous vascular differentiation remains unique.
The monopteros (mp) gene contributes to apical-basal pattern formation in the Arabidopsis embryo. mp mutant seedlings lack basal body structures such as hypocotyl, radicle and root meristem, and this pattern deletion has been traced back to alterations in the octant-stage embryo. Cells of the embryo proper and the uppermost cell of the suspensor fail to establish division patterns that would normally generate the basal body structures. The resulting absence of a morphological axis seems to be responsible for another phenotypic trait of mp seedlings, variable positioning of cotyledons. This rela - tionship is suggested by weak mp seedling phenotypes in which the presence of a short hypocotyl is correlated with normal arrangement of cotyledons. Root formation has been induced in mp seedlings grown in tissue culture. This result supports the notion that the mp gene is required for organising the basal body region, rather than for making the root, in the developing embryo. SUMMARY
Plants have a remarkable potential for sustained (indeterminate) postembryonic growth. Following their specification in the early embryo, tissue-specific precursor cells first establish tissues and later maintain them postembryonically. The mechanisms underlying these processes are largely unknown. Here we define local control of oriented, periclinal cell division as the mechanism underlying both the establishment and maintenance of vascular tissue. We identify an auxin-regulated basic helix-loop-helix (bHLH) transcription factor dimer as a critical regulator of vascular development. Due to a loss of periclinal divisions, vascular tissue gradually disappears in bHLH-deficient mutants; conversely, ectopic expression is sufficient for triggering periclinal divisions. We show that this dimer operates independently of tissue identity but is restricted to a small vascular domain by integrating overlapping transcription patterns of the interacting bHLH proteins. Our work reveals a common mechanism for tissue establishment and indeterminate vascular development and provides a conceptual framework for developmental control of local cell divisions.
Plant vascular tissues are essential for the existence of land plants. Many studies of transcriptional regulation and cell-cell communication have revealed the process underlying the development of vascular tissues from vascular initial cells. However, the initiation of vascular cell differentiation is still a mystery. Here, we report that LONESOME HIGHWAY (LHW), which encodes a bHLH transcription factor, is expressed in pericycle-vascular mother cells at the globular embryo stage and is required for proper asymmetric cell division to generate vascular initial cells. In addition, ectopic expression of LHW elicits an ectopic auxin response. Moreover, LHW is required for the correct expression patterns of components related to auxin flow, such as PIN-FORMED 1 (PIN1), MONOPTEROS (MP) and ATHB-8, and ATHB-8 partially rescues the vascular defects of lhw. These results suggest that LHW functions as a key regulator to initiate vascular cell differentiation in association with auxin regulation.
The vascular system in plants, which comprises xylem, phloem and vascular stem cells, originates from provascular cells and
forms a continuous network throughout the plant body. Although various aspects of vascular development have been extensively
studied, the early process of vascular development remains largely unknown. LONESOME HIGHWAY (LHW), which encodes an atypical basic helix–loop–helix (bHLH) transcription factor, plays an essential role in establishing vascular
cells. Here, we report the analysis of LHW homologs in relation to vascular development. Three LHW homologs, LONESOME HIGHWAY LIKE 1–3 (LHL1–LHL3), were preferentially expressed in the plant vasculature. Genetic analysis indicated that, although the LHL3 loss-of-function mutant showed no obvious phenotype, the lhw lhl3 double mutant displayed more severe phenotypic defects in the vasculature of the cotyledons and roots than the lhw single mutant. Only one xylem vessel was formed at the metaxylem position in lhw lhl3 roots, whereas the lhw root formed one protoxylem and one or two metaxylem vessels. Conversely, overexpression of LHL3 enhanced xylem development in the roots. Moreover, N-1-naphthylphthalamic acid caused ectopic LHL3 expression in accordance with induced auxin maximum. These results suggest that LHL3 plays a positive role in xylem differentiation
downstream of auxin.
Unlabelled:
The direct induction of adventitious buds and somatic embryos from explants is a morphogenetic process that is under the influence of exogenous plant growth regulators and its interactions with endogenous phytohormones. We performed an in vitro histological analysis in peach palm (Bactris gasipaes Kunth) shoot apexes and determined that the positioning of competent cells and their interaction with neighboring cells, under the influence of combinations of exogenously applied growth regulators (NAA/BAP and NAA/TDZ), allows the pre-procambial cells (PPCs) to act in different morphogenic pathways to establish niche competent cells. It is likely that there has been a habituation phenomenon during the regeneration and development of the microplants. This includes promoting the tillering of primary or secondary buds due to culturing in the absence of NAA/BAP or NAA/TDZ after a period in the presence of these growth regulators. Histological analyses determined that the adventitious roots were derived from the dedifferentiation of the parenchymal cells located in the basal region of the adventitious buds, with the establishment of rooting pole, due to an auxin gradient. Furthermore, histological and histochemical analyses allowed us to characterize how the PPCs provide niches for multipotent, pluripotent and totipotent stem-like cells for vascular differentiation, organogenesis and somatic embryogenesis in the peach palm. The histological and histochemical analyses also allowed us to detect the unicellular or multicellular origin of somatic embryogenesis. Therefore, our results indicate that the use of growth regulators in microplants can lead to habituation and to different morphogenic pathways leading to potential niche establishment, depending on the positioning of the competent cells and their interaction with neighboring cells.
Key message:
Our results indicate that the use of growth regulators in microplants can lead to habituation and to different morphogenic pathways leading to potential niche establishment, depending on the positioning of the competent cells and their interaction with neighboring cells.
Plant vascular tissues form systems of interconnected cell files throughout the plant body. Vascular tissues usually differentiate at predictable positions but the wide range of functional patterns generated in response to abnormal growth conditions or wounding reveals partially self-organizing patterning mechanisms. Signals ensuring aligned cell differentiation within vascular strands are crucial in self-organized vascular patterning, and the apical–basal flow of indole acetic acid has been suspected to act as an orienting signal in this process. Several recent advances appear to converge on a more precise definition of the role of auxin flow in vascular tissue patterning.
Plant growth is directed by the activity of stem cells within meristems. The first meristems are established during early embryogenesis, and this process involves the specification of both stem cells and their organizer cells. One of the earliest events in root meristem initiation is marked by re-specification of the uppermost suspensor cell as hypophysis, the precursor of the organizer. The transcription factor MONOPTEROS (MP) is a key regulator of hypophysis specification, and does so in part by promoting the transport of the plant hormone auxin and by activating the expression of TARGET OF MP (TMO) transcription factors, both of which are required for hypophysis specification. The mechanisms leading to the activation of these genes by MP in a chromatin context are not understood. Here, we show that the PHD-finger proteins OBERON (OBE) and TITANIA (TTA) are essential for MP-dependent embryonic root meristem initiation. TTA1 and TTA2 are functionally redundant and function in the same pathway as OBE1 and OBE2. These PHD-finger proteins interact with each other, and genetic analysis shows that OBE-TTA heterotypic protein complexes promote embryonic root meristem initiation. Furthermore, while MP expression is unaffected by mutations in OBE/TTA genes, expression of MP targets TMO5 and TMO7 is locally lost in obe1 obe2 embryos. PHD-finger proteins have been shown to act in initiation of transcription by interacting with nucleosomes. Indeed, we found that OBE1 binds to chromatin at the TMO7 locus, suggesting a role in its MP-dependent activation. Our data indicate that PHD-finger protein complexes are crucial for the activation of MP-dependent gene expression during embryonic root meristem initiation, and provide a starting point for studying the mechanisms of developmental gene activation within a chromatin context in plants.
The variability of shoot architecture in plants is striking and one of the most extreme examples of adaptive growth in higher organisms. Mediated by the differential activity of apical and lateral meristems, flexibility in stem growth essentially contributes to this variability. In spite of this importance, the regulation of major events in stem development is largely unexplored. Recently, however, novel approaches exploiting knowledge from root and leaf development are starting to shed light on molecular mechanisms that regulate this essential plant organ. In this review, we summarize our understanding of initial patterning events in stems, discuss prerequisites for the initiation of lateral stem growth and highlight the burning questions in this context.
Multipotent stem cell populations, the meristems, are fundamental for the indeterminate growth of plant bodies. One of these meristems, the cambium, is responsible for extended root and stem thickening. Strikingly, although the pivotal role of the plant hormone auxin in promoting cambium activity has been known for decades, the molecular basis of auxin responsiveness on the level of cambium cells has so far been elusive. Here, we reveal that auxin-dependent cambium stimulation requires the homeobox transcription factor WOX4. In Arabidopsis thaliana inflorescence stems, 1-N-naphthylphthalamic acid-induced auxin accumulation stimulates cambium activity in the wild type but not in wox4 mutants, although basal cambium activity is not abolished. This conclusion is confirmed by the analysis of cellular markers and genome-wide transcriptional profiling, which revealed only a small overlap between WOX4-dependent and cambium-specific genes. Furthermore, the receptor-like kinase PXY is required for a stable auxin-dependent increase in WOX4 mRNA abundance and the stimulation of cambium activity, suggesting a concerted role of PXY and WOX4 in auxin-dependent cambium stimulation. Thus, in spite of large anatomical differences, our findings uncover parallels between the regulation of lateral and apical plant meristems by demonstrating the requirement for a WOX family member for auxin-dependent regulation of lateral plant growth.
The plant hormone auxin triggers a wide range of developmental and growth responses throughout a plant's life. Most well-known auxin responses involve changes in gene expression that are mediated by a short pathway involving an auxin-receptor/ubiquitin-ligase, DNA-binding auxin response factor (ARF) transcription factors and their interacting auxin/indole-3-acetic acid (Aux/IAA) transcriptional inhibitors. Auxin promotes the degradation of Aux/IAA proteins through the auxin receptor and hence releases the inhibition of ARF transcription factors. Although this generic mechanism is now well understood, it is still unclear how developmental specificity is generated and how individual gene family members of response components contribute to local auxin responses. We have established a collection of transcriptional reporters for the ARF gene family and used these to generate a map of expression during embryogenesis and in the primary root meristem. Our results demonstrate that transcriptional regulation of ARF genes generates a complex pattern of overlapping activities. Genetic analysis shows that functions of co-expressed ARFs converge on the same biological processes, but can act either antagonistically or synergistically. Importantly, the existence of an 'ARF pre-pattern' could explain how cell-type-specific auxin responses are generated. Furthermore, this resource can now be used to probe the functions of ARF in other auxin-dependent processes.
Cytokinin phytohormones regulate a variety of developmental processes in the root such as meristem size, vascular pattern, and root architecture [1-3]. Long-distance transport of cytokinin is supported by the discovery of cytokinins in xylem and phloem sap [4] and by grafting experiments between wild-type and cytokinin biosynthesis mutants [5]. Acropetal transport of cytokinin (toward the shoot apex) has also been implicated in the control of shoot branching [6]. However, neither the mode of transport nor a developmental role has been shown for basipetal transport of cytokinin (toward the root apex). In this paper, we combine the use of a new technology that blocks symplastic connections in the phloem with a novel approach to visualize radiolabeled hormones in planta to examine the basipetal transport of cytokinin. We show that this occurs through symplastic connections in the phloem. The reduction of cytokinin levels in the phloem leads to a destabilization of the root vascular pattern in a manner similar to mutants affected in auxin transport or cytokinin signaling [7]. Together, our results demonstrate a role for long-distance basipetal transport of cytokinin in controlling polar auxin transport and maintaining the vascular pattern in the root meristem.
Whereas the majority of animals develop toward a predetermined body plan, plants show iterative growth and continually produce new organs and structures from actively dividing meristems. This raises an intriguing question: How are these newly developed organs patterned? In Arabidopsis embryos, radial symmetry is broken by the bisymmetric specification of the cotyledons in the apical domain. Subsequently, this bisymmetry is propagated to the root promeristem.
Here we present a mutually inhibitory feedback loop between auxin and cytokinin that sets distinct boundaries of hormonal output. Cytokinins promote the bisymmetric distribution of the PIN-FORMED (PIN) auxin efflux proteins, which channel auxin toward a central domain. High auxin promotes transcription of the cytokinin signaling inhibitor AHP6, which closes the interaction loop. This bisymmetric auxin response domain specifies the differentiation of protoxylem in a bisymmetric pattern. In embryonic roots, cytokinin is required to translate a bisymmetric auxin response in the cotyledons to a bisymmetric vascular pattern in the root promeristem.
Our results present an interactive feedback loop between hormonal signaling and transport by which small biases in hormonal input are propagated into distinct signaling domains to specify the vascular pattern in the root meristem. It is an intriguing possibility that such a mechanism could transform radial patterns and allow continuous vascular connections between other newly emerging organs.
The indeterminate nature of plant growth and development depends on the stem cell system found in meristems. The Arabidopsis thaliana vascular meristem includes procambium and cambium. In these tissues, cell-cell signaling, mediated by a ligand-receptor pair made of the TDIF (for tracheary element differentiation inhibitory factor) peptide and the TDR/PXY (for TDIF RECEPTOR/ PHLOEM INTERCALATED WITH XYLEM) membrane protein kinase, promotes proliferation of procambial cells and suppresses their xylem differentiation. Here, we report that a WUSCHEL-related HOMEOBOX gene, WOX4, is a key target of the TDIF signaling pathway. WOX4 is expressed preferentially in the procambium and cambium, and its expression level was upregulated upon application of TDIF in a TDR-dependent manner. Genetic analyses showed that WOX4 is required for promoting the proliferation of procambial/cambial stem cells but not for repressing their commitment to xylem differentiation in response to the TDIF signal. Thus, at least two intracellular signaling pathways that diverge after TDIF recognition by TDR might regulate independently the behavior of vascular stem cells. Detailed observations in loss-of-function mutants revealed that TDIF-TDR-WOX4 signaling plays a crucial role in the maintenance of the vascular meristem organization during secondary growth.
Acquisition of cell identity in plants relies strongly on positional information, hence cell-cell communication and inductive signalling are instrumental for developmental patterning. During Arabidopsis embryogenesis, an extra-embryonic cell is specified to become the founder cell of the primary root meristem, hypophysis, in response to signals from adjacent embryonic cells. The auxin-dependent transcription factor MONOPTEROS (MP) drives hypophysis specification by promoting transport of the hormone auxin from the embryo to the hypophysis precursor. However, auxin accumulation is not sufficient for hypophysis specification, indicating that additional MP-dependent signals are required. Here we describe the microarray-based isolation of MP target genes that mediate signalling from embryo to hypophysis. Of three direct transcriptional target genes, TARGET OF MP 5 (TMO5) and TMO7 encode basic helix-loop-helix (bHLH) transcription factors that are expressed in the hypophysis-adjacent embryo cells, and are required and partially sufficient for MP-dependent root initiation. Importantly, the small TMO7 transcription factor moves from its site of synthesis in the embryo to the hypophysis precursor, thus representing a novel MP-dependent intercellular signal in embryonic root specification.
Land plants evolved a long-distance transport system of water and nutrients composed of the xylem and phloem, both of which are generated from the procambium- and cambium-comprising vascular stem cells. However, little is known about the molecular mechanism of cell communication governing xylem–phloem patterning. Here, we show that a dodecapeptide (HEVHypSGHypNPISN; Hyp, 4-hydroxyproline), TDIF (tracheary element differentiation inhibitory factor), is secreted from the phloem and suppresses the differentiation of vascular stem cells into xylem cells through a leucine-rich repeat receptor-like kinase (LRR-RLK). TDIF binds in vitro specifically to the LRR-RLK, designated TDR (putative TDIF receptor), whose expression is restricted to procambial cells. However, the combined analysis of TDIF with a specific antibody and the expression profiles of the promoters of two genes encoding TDIF revealed that TDIF is synthesized mainly in, and secreted from, the phloem and its neighboring cells. The observation that TDIF is capable of promoting proliferation of procambial cells while suppressing xylem differentiation suggests that this small peptide functions as a phloem-derived, non-cell-autonomous signal that controls stem cell fate in the procambium. Our results indicate that we have discovered a cell communication system governing phloem–xylem cross-talk.
• CLV3/ESR-related (CLE)
• leucine-rich repeat receptor-like kinase
• phloem
• procambium
• xylem