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Internode cell wall composition, lignin, and monosaccharide contents. A-D, Representative pictures of the lignin deposition (phloroglucinol-HCl staining, red coloration) in the first internodes of fully senesced WT and cul4 TI laxa TI mutants. A and B, Whole stem and (C and D), stem magnification focusing on fascicular and interfascicular bundles of B. distachyon from WT (A and C) and cul4 TI laxa TI (B and D). In cul4 TI laxa TI , the lignin staining is less intense compared to WT, especially at the interfascicular regions. Scale bars: 100 lm. E, Quantification of acetyl bromide soluble lignin content. F, Quantification of b-O-4 linked H (circles), G (triangles), and S (crosses) lignin units by thioacidolysis. G, Main cell wall sugar contents in dried B. distachyon stem. ara, arabinoxylan; xyl, xylose and glc, glucose. For each line, pulverized stem tissue from 6 to 16 individuals from three independent replicates was analyzed. E-G, Asterisks indicate significant differences relative to WT (*P 50.05; ***P 5 0.001; ****P 5 0.0001; Mann-Whitney U-test), n = 3 and error bars represent standard deviation. %DM stands for percentage of dry mass. H, Relative transcript abundance of the lignin biosynthesis-related genes BdCAD1 and BdCOMT6, and of the cell wall-related genes BdCESA4, BdCESA7, BdCESA8, and BdSWAM1 in WT (white bars) and cul4 TI laxa TI (purple bars). Gene expression analysis was performed by RT-qPCR on stem samples. Gene expression was normalized against the constitutively expressed BdUBQ4 and BdUBC18 reference genes. Results represent means ± SEM (standard error of mean) of three biological repeats and three technical replicates. Primers used for RT-qPCR analysis are given in Supplemental Table S2. For the box plot, the median (middle horizontal line), upper and lower quartiles (boxes), as well as minimum and maximum values (whiskers) are indicated. Individual values are indicated by dots. Asterisks indicate significant differences relative to WT (*P 5 0.05; **P 5 0.01; one-way annalysis of variance (ANOVA)).
Source publication
In cultivated grasses, tillering, leaf and inflorescence architecture, as well as abscission ability, are major agronomical traits. In barley (Hordeum vulgare), maize (Zea mays), rice (Oryza sativa), and brachypodium (Brachypodium distachyon), NOOT-BOP-COCH-LIKE (NBCL) genes are essential regulators of vegetative and reproductive development. Grass...
Contexts in source publication
Context 1
... their tillering modifications, both the cul4 TI laxa TI and cul4 TI laxa CR double mutants were dwarf while no significant difference in plant height was recorded between cul4 TI , laxa TI , and laxa CR relative to WT (Figure 1). In addition, while cul4 TI showed no significant difference relative to WT, laxa TI , and laxa CR had shorter roots compared to WT (Supplemental Figure S3). As the LAXA gene is apparently not expressed in roots ( Winter et al., 2007;Sibout et al., 2017;Magne et al., 2020; http://bar.utoronto.ca/efp_brachy ...
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... it suggests that in B. distachyon, LAXA could play an indirect and positive effect on root development, maybe through resource allocation between the roots and the aerial parts. In addition, as expected for a dwarf line, the cul4 TI laxa TI root length was also reduced by 40% compared to the WT (Supplemental Figure S3). ...
Context 3
... observed that the interfascicular fibers of cul4 TI laxa TI double mutant were yellow or white relative to the red ones of the WT plants. In contrast, the vascular bundles of cul4 TI laxa TI were stained in red suggesting that vascular bundle tissues were not or less impacted than interfascicular fibers ( Figure 3, A-D). These results indicate that, in cul4 TI laxa TI , a very low amount of lignin is deposited onto the interfascicular fiber cell wall. ...
Context 4
... results indicate that, in cul4 TI laxa TI , a very low amount of lignin is deposited onto the interfascicular fiber cell wall. Subsequent quantification of the acetyl bromide soluble lignin content in cul4 TI laxa TI supported our findings and showed a significant decrease of 32% compared to the WT ( Figure 3E). Not unexpectedly, thioacidolysis analyses revealed a significant reduction of uncondensed guaiacyl (G), syringyl (S), and p-hydroxy-phenyl (H) lignin units content in cul4 TI laxa TI compared to the WT ( Figure 3F). ...
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... quantification of the acetyl bromide soluble lignin content in cul4 TI laxa TI supported our findings and showed a significant decrease of 32% compared to the WT ( Figure 3E). Not unexpectedly, thioacidolysis analyses revealed a significant reduction of uncondensed guaiacyl (G), syringyl (S), and p-hydroxy-phenyl (H) lignin units content in cul4 TI laxa TI compared to the WT ( Figure 3F). This striking change in lignin content led us to investigate the content of other cell wall compounds in the double mutant lines. ...
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... striking change in lignin content led us to investigate the content of other cell wall compounds in the double mutant lines. Similarly, the sugar composition, including glucose (mainly extracted from cellulose) was significantly affected in the double mutants with a reduction of the xylose and glucose contents ( Figure 3G). In addition, our study suggested that the content in arabinoxylans, the main hemicellulose in B. distachyon stems, was also impacted but in a positive manner. ...
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... BdSECONDARY WALL-ASSOCIATED MYB1 (BdSWAM1) acts as a positive regulator of cell wall biosynthesis ( Handakumbura et al., 2018). Our results showed that the expression level of BdCAD1, BdCOMT6, BdCESA4, BdCESA7, and BdCESA8 was significantly downregulated in cul4 TI laxa TI compared to WT, while the expression of BdSWAM1 was significantly upregulated ( Figure 3H; Supplemental Figure S5). ...
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MADS-box transcription factors are crucial regulators of inflorescence and flower development in plants. Therefore, the recent interest in this family has received much attention in plant breeding programs due to their impact on plant development and inflorescence architecture. The aim of this study was to investigate the role of HvMADS-box genes i...
Citations
... BTB-ankyrin proteins from both subclades can also function as E3 ubiquitin ligase adaptors involved in regulating protein abundance (Fu et al., 2012;Zhang et al., 2017;Chahtane et al., 2018;He et al., 2020). In monocots and dicots, BOPs contribute to a surprisingly large number of developmental processes, including lignin deposition as a defense (Zhang et al., 2019) and during secondary growth (Khan et al., 2012b;Woerlen et al., 2017;Shen et al., 2021;Liu et al., 2022). ...
... Phenotypic defects in bop1 bop2 double mutants are concentrated at lateral organ boundaries resulting in elongated leafy petioles, five-petalled flowers with a subtending bract, and defects in abscission (Hepworth et al., 2005;Norberg et al., 2005;McKim et al., 2008). In monocots and dicots, BOP1 and BOP2 paralogs participate in many of the same developmental processes (Wu et al., 2012;Tavakol et al., 2015;Couzigou et al., 2016;Jost et al., 2016;Xu et al., 2016;Toriba et al., 2019;Magne et al., 2020;Liu et al., 2022). ...
... The BOP family of co-transcriptional regulators are conserved in dicots and monocots and contribute to numerous developmental processes. Studies in dicots such as tobacco (Wu et al., 2012), legumes (Frankowski et al., 2015;Couzigou et al., 2016;Magne et al., 2018), tomato (Xu et al., 2016;Izhaki et al., 2018) and monocots such as rice (Toriba et al., 2020), barley (Tavakol et al., 2015;Jost et al., 2016) and Brachypodium (Magne et al., 2020;Liu et al., 2022) reveal highly conserved functions, comparable to Arabidopsis, that influence organ patterning, inflorescence architecture, and abscission (Khan et al., 2014;Hepworth and Pautot, 2015). Taxon-specific roles in the maintenance of symbiotic nodule identity (Couzigou et al., 2012;Magne et al., 2018) and rhizome development (Toriba et al., 2020) have also been identified. ...
BLADE-ON-PETIOLE (BOP) genes are essential regulators of vegetative and reproductive development in land plants. First characterized in Arabidopsis thaliana (Arabidopsis), members of this clade function as transcriptional co-activators by recruiting TGACG-motif binding (TGA) basic leucine zipper (bZIP) transcription factors. Highly expressed at organ boundaries, these genes are also expressed in vascular tissue and contribute to lignin biosynthesis during secondary growth. How these genes function in trees, which undergo extensive secondary growth to produce wood, remains unclear. Here, we investigate the functional conservation of BOP orthologs in Populus trichocarpa (poplar), a widely-used model for tree development. Within the poplar genome, we identified two BOP-like genes, PtrBPL1 and PtrBPL2, with abundant transcripts in stems. To assess their functions, we used heterologous assays in Arabidopsis plants. The promoters of PtrBPL1 and PtrBPL2, fused with a β-glucuronidase (GUS) reporter gene showed activity at organ boundaries and in secondary xylem and phloem. When introduced into Arabidopsis plants, PtrBPL1 and PtrBPL2 complemented leaf and flower patterning defects in bop1 bop2 mutants. Notably, Arabidopsis plants overexpressing PtrBPL1 and PtrBPL2 showed defects in stem elongation and the lignification of secondary tissues in the hypocotyl and stem. Finally, PtrBPL1 and PtrBPL2 formed complexes with TGA bZIP proteins in yeast. Collectively, our findings suggest that PtrBPL1 and PtrBPL2 are orthologs of Arabidopsis BOP1 and BOP2, potentially contributing to secondary growth regulation in poplar trees. This work provides a foundation for functional studies in trees.
... Other hormones, including GA, cytokinin (CK), ethylene (ET), directly or indirectly affect BR synergetic regulation of LA Cao et al., 2022b). Therefore, there is still much room to explore the LG1 regulatory network related to the development of other ligular regions in plants (Cao et al., 2022a;Liu et al., 2022). As a member of the SBP family, ...
Plant architecture is a culmination of the features necessary for capturing light energy and adapting to the environment. An ideal architecture can promote an increase in planting density, light penetration to the lower canopy, airflow as well as heat distribution to achieve an increase in crop yield. A number of plant architecture-related genes have been identified by map cloning, quantitative trait locus (QTL) and genome-wide association study (GWAS) analysis. LIGULELESS1 (LG1) belongs to the squamosa promoter-binding protein (SBP) family of transcription factors (TFs) that are key regulators for plant growth and development, especially leaf angle (LA) and flower development. The DRL1/2-LG1-RAVL pathway is involved in brassinosteroid (BR) signaling to regulate the LA in maize, which has facilitated the regulation of plant architecture. Therefore, exploring the gene regulatory functions of LG1, especially its relationship with LA genes, can help achieve the precise regulation of plant phenotypes adapted to varied environments, thereby increasing the yield. This review comprehensively summarizes the advances in LG1 research, including its effect on LA and flower development. Finally, we discuss the current challenges and future research goals associate with LG1.
... In comparison to previous reports, the protocol presented herein cuts one to two months to the creation of Bd transgenics [6,32] or CRISPR/Cas9-induced mutants [18,22,27], with no observable reduction in transformation efficiency. In routine procedures, we typically use ca. 100 immature primary embryos in a single Agrobacterium co-cultivation experiment for each construct of interest (five initial CIM plates, each with 20 izEmb), whether for straightforward transgenesis or simple CRISPR/Cas9 gene editing. ...
Background
Even for easy-to-transform species or genotypes, the creation of transgenic or edited plant lines remains a significant bottleneck. Thus, any technical advance that accelerates the regeneration and transformation process is welcome. So far, methods to produce Brachypodium distachyon (Bd) transgenics span at least 14 weeks from the start of tissue culture to the recovery of regenerated plantlets.
Results
We have previously shown that embryogenic somatic tissues grow in the scutellum of immature zygotic Bd embryos within 3 days of in vitro induction with exogenous auxin and that the development of secondary embryos can be initiated immediately thereafter. Here, we further demonstrate that such pluripotent reactive tissues can be genetically transformed with Agrobacterium tumefaciens right after the onset of somatic embryogenesis. In brief, immature zygotic embryos are induced for callogenesis for one week, co-cultured with Agrobacterium for three days, then incubated on callogenesis selective medium for three weeks, and finally transferred on selective regeneration medium for up to three weeks to obtain plantlets ready for rooting. This 7-to-8-week procedure requires only three subcultures. Its validation includes the molecular and phenotype characterization of Bd lines carrying transgenic cassettes and novel CRISPR/Cas9-generated mutations in two independent loci coding for nitrate reductase enzymes (BdNR1 and BdNR2).
Conclusions
With a short callogenesis stage and streamlined in vitro regeneration following co-cultivation with Agrobacterium, transgenic and edited T0 Bd plantlets can be produced in about 8 weeks, a gain of one to two months compared to previously published methods, with no reduction in transformation efficiency and at lower costs.
Medicago truncatula NODULE ROOT1 (MtNOOT1) and Pisum sativum COCHLEATA1 (PsCOCH1) are orthologous genes belonging to the NOOT-BOP-COCH-LIKE (NBCL) gene family which encodes key transcriptional co-regulators of plant development. In Mtnoot1 and Pscoch1 mutants, the development of stipule, flower and symbiotic nodules is altered. MtNOOT2 and PsCOCH2 represent the single paralogs of MtNOOT1 and PsCOCH1, respectively. In M. truncatula, MtNOOT1 and MtNOOT2 are both required for the establishment and maintenance of the symbiotic nodule identity.
In contrast to the NBCL1 genes, the role of NBCL2 genes in legume above-ground development is not known. To better understand the roles of NBCL genes in legumes, we used M. truncatula and P. sativum nbcl mutants from the literature, isolated a knockout mutant for the PsCOCH2 locus and generated Pscoch1coch2 double mutants in P. sativum. These new mutant lines enabled to compare the roles of MtNOOT2 and PsCOCH2 in both M. truncatula and P. sativum legume development.
Our work shows that the single Mtnoot2 and Pscoch2 mutants develop wild-type stipules, flowers and symbiotic nodules. However, the number of flowers was increased and the pods and seeds were smaller in comparison to wild type. Furthermore, in comparison to the corresponding nbcl1 single mutants, both the M. truncatula and P. sativum nbcl double mutants show a drastic alteration in stipule, inflorescence, flower and nodule development. Remarkably, in both M. truncatula and P. sativum nbcl double mutants, stipules are transformed into a range of aberrant leaf-like structures.
