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NFL, the tobacco homolog of FLORICAULA and LEAFY, is transcriptionally expressed in both vegetative and floral meristems
Abstract
The homologous genes FLORICAULA (FLO) of Antirrhinum and LEAFY (LFY) of Arabidopsis regulate the formation of determinate floral meristems. Transcripts of these single-copy genes are confined to floral meristems and some floral organs as well as to the leaflike bracts that subtend Antirrhinum flowers. Based on these observations, we hypothesized that the transcription of genes homologous to FLO and LFY in tobacco, a determinate plant in which the primary shoot apex is consumed in the production of a terminal flower, would serve as a molecular marker for floral commitment. Surprisingly, transcripts of the tobacco homologs NFL1 and NFL2 (Nicotiana FLO/LFY) were found not only in floral meristems, but also in indeterminate vegetative meristems. This implies that the transcriptional expression of the FLO/LFY homologous genes in the apical meristem is not sufficient for the initiation of floral meristem development. In addition, the transcript patterns of the NFL genes identified a previously undescribed subset of cells within the shoot apical meristem that may indicate unique functional compartmentalization. This suggests that, unlike FLO and LFY, which specify determinacy only during floral development, the NFL genes act to specify determinacy in the progenitor cells for both flowers and leaves.
... But only LFY in gymnosperms is orthologous with LFY in angiosperms. The LFY and NEEDLY paralogs in gymnosperms are specifically expressed in both vegetative and reproductive meristems 18,19 . The LFY paralog provides more activity to rescue Arabidopsis lfy mutant phenotypes than NEEDLY 1,18 . ...
... The LFY and NEEDLY paralogs in gymnosperms are specifically expressed in both vegetative and reproductive meristems 18,19 . The LFY paralog provides more activity to rescue Arabidopsis lfy mutant phenotypes than NEEDLY 1,18 . ...
... LFY is strongly expressed in floral primordia, and faintly expressed in cauline leaf primordia 4 . Moreover, constitutive overexpression of LFY homologs results in precocious flowering in most angiosperms and gymnosperms 15,17,18 . However, most of the IsLFY1 and IsLFY2 transgenic lines did not show precocious flowering relative to the wild-type plants, suggesting that the two paralogs were not functional orthologs with LFY 4, 17, 18 . ...
LEAFY (LFY) is a plant-specific transcription factor, which is found in algae and all land plants. LFY homologs exert ancestral roles in regulating cell division and obtain novel functions to control floral identity. Isoetes L. is an ancient genus of heterosporous lycophytes. However, characters about LFY homologs in lycophytes remain poorly investigated. In this study, two LFY homologs, ILFY1 and ILFY2, were cloned from five Isoetes species, including I. hypsophila, I. yunguiensis, I. sinensis, I. orientalis, and I. taiwanensis. The full length of ILFY1 was 1449–1456 bp with an open reading frame (ORF) of 927–936 bp. The full length of ILFY2 was 1768 bp with ORF of 726 bp. Phylogenetic tree revealed that ILFY1 and ILFY2 were separated into two clades, and I. hypsophila were separated with the others. Expression analysis demonstrated that IsLFY1 and IsLFY2 for I. sinensis did not show functional diversity. The two transcripts were similarly accumulated in both vegetative and reproductive tissues and highly expressed in juvenile tissues. In addition, the IsLFY1 and IsLFY2 transgenic Arabidopsis similarly did not promote precocious flowering, and they were inactive to rescue lfy mutants. The results facilitate general understandings about the characteristics of LFY in Isoetes and evolutionary process.
... Conversely, ectopic LFY expression prevents TFL1 expression in the shoot apex (Weigel and Nilsson, 1995;Ratcliffe et al., 1999). In tobacco, the expression pattern of NFL (for Nicotiana FLO LFY), the likely FLO/LFY homolog, is different from that of FLO in Antirrhinum and LFY in Arabidopsis ( Kelly et al., 1995). NFL is expressed in the shoot apical region throughout development but not in the apical meristem itself; NFL is expressed in a ring of cells outside the central dome in floral meristems and leaf primordia. ...
... To assess whether NFL and CET2 and CET4 were coexpressed in axillary meristems or were restricted to separate domains, as occurs in Antirrhinum and Arabidopsis, we compared the expression patterns of NFL with those of CET2 and CET4 by using in situ hybridization (Figure 4). During the vegetative phase, young axillary meristems near the apex of wild-type tobacco plants strongly expressed CET2 and CET4 ( Figures 4A, 4B, and 4D), whereas NFL RNA was not detected in these axillary meristems but was restricted to the flanks of the main shoot meristem (Figures 4C and 4E; Kelly et al., 1995). In older axillary meristems, after some leaf primordia had been generated, NFL was detected in a pattern similar to that in the apical meristem ( Figure 4G), and CET2 and CET4 were expressed in the central region below the apical dome of these meristems ( Figure 4F). ...
... Floral meristem identity genes appear not to be expressed in the shoot apical meristem of indeterminate species but are restricted to primordia or meristems arising from the periphery. In contrast, many determinate species, such as tobacco, tomato, Impatiens balsimina, and petunia, have FLO-and LFY-like genes that are expressed strongly in the shoot apical meristem from very early in development ( Kelly et al., 1995;Pouteau et al., 1997;Pnueli et al., 1998;Souer et al., 1998). Recent data showed that LFY and AP1 repress the expression of TFL1 in Arabidopsis ( Ratcliffe et al., 1999). ...
Plant species exhibit two primary forms of flowering architecture, namely, indeterminate and determinate. Antirrhinum is an indeterminate species in which shoots grow indefinitely and only generate flowers from their periphery. Tobacco is a determinate species in which shoot meristems terminate by converting to a flower. We show that tobacco is responsive to the CENTRORADIALIS (CEN) gene, which is required for indeterminate growth of the shoot meristem in Antirrhinum. Tobacco plants overexpressing CEN have an extended vegetative phase, delaying the switch to flowering. Therefore, CEN defines a conserved system controlling shoot meristem identity and plant architecture in diverse species. To understand the underlying basis for differences between determinate and indeterminate architectures, we isolated CEN-like genes from tobacco (CET genes). In tobacco, the CET genes most similar to CEN are not expressed in the main shoot meristem; their expression is restricted to vegetative axillary meristems. As vegetative meristems develop into flowering shoots, CET genes are downregulated as floral meristem identity genes are upregulated. Our results suggest a general model for tobacco, Antirrhinum, and Arabidopsis, whereby the complementary expression patterns of CEN-like genes and floral meristem identity genes underlie different plant architectures.
... The LFY gene has been cloned and its function has been studied from a few species viz. Antirrhinum [16], tobacco [17], Eucalyptus [18], Pinus [19], rice [20], tomato [21], poplar [14], violet cress [22], apple [23], grapevine [24], papaya [25], rubber [26] and cedar [27]. ...
... thaliana, A. majus, pea, tomato, petunia and poplar) [6,14,16,21,31,32]. In apple genome here are two homologous copies of LFY genes (AFL1 t and AFL2) [23] and so also in tobacco (NFL1 and NFL2) [17]. Eucalyptus has three homologous genes in the genome, but two of them were found not to be expressed and had stop codons in their coding region [18]. ...
... This is similar to the expression pattern observed in A. thaliana and A. majus [6,16]. oleracea, ELF gene from Eucalyptus, AFL1 gene from apple and NFL genes from tobacco) have not been reported to express in leaf and stem [17,18,23,34]. ...
A LEAFY cDNA was cloned from the short-day (SD) plant Brassica juncea cv Varuna. (BjLFY) consists of 1261 bp encoding for a protein of 420 amino acids and an estimated isoelectric point of 6.8. The deduced amino acid showed 99% and 86% identity to Arabi-dopsis thaliana and Brassica oleracea LFY cDNA respectively. The LFY transcript was detected throughout the vegetative and reproductive phase but an increase in transcript level was observed during transition. The earlier induction of BjLFY was observed in early flowering variety of B. juncea as compared to late flowering varieties further proving the critical role LEAFY plays in floral transition.
... The UNI and FA genes are homologs of the floral meristem identity genes FLORICAULA ( FLO ;Coen et al., 1990) and LEAFY ( LFY ;Weigel et al., 1992) from Antirrhinum and Arabidopsis, respectively (Hofer et al., 1997;MolineroRosales et al., 1999). Other potential homologs of FLO / LFY have been identified in monocotyledonous (Colombo et al., 1998;Kyozuka et al., 1998) and dicotyledonous (Anthony et al., 1993;Rottman et al., 1993;Kelly et al., 1995;Pouteau et al., 1997;Souer et al., 1998;Molinero-Rosales et al., 1999) angiosperm species, in basal angiosperms and gnetales (Frohlich and Meyerowitz, 1997), and in the gymnosperm pine (Mouradov et al., 1998). Of the dicots studied to date, FLO/LFY transcripts were detected in the leaf primordia of tobacco, Arabidopsis, pea, Impatiens, tomato, and petunia (Kelly et al., 1995;Blázquez et al., 1997;Hofer et al., 1997;Pouteau et al., 1997;Pnueli et al., 1998;Souer et al., 1998;Molinero-Rosales et al., 1999), but mutant leaf phenotypes have been described only for those species with compound leaves: pea and tomato.Hofer et al. (1997)proposed a common function for UNI in regulating indeterminacy during both leaf and flower development. ...
... Other potential homologs of FLO / LFY have been identified in monocotyledonous (Colombo et al., 1998;Kyozuka et al., 1998) and dicotyledonous (Anthony et al., 1993;Rottman et al., 1993;Kelly et al., 1995;Pouteau et al., 1997;Souer et al., 1998;Molinero-Rosales et al., 1999) angiosperm species, in basal angiosperms and gnetales (Frohlich and Meyerowitz, 1997), and in the gymnosperm pine (Mouradov et al., 1998). Of the dicots studied to date, FLO/LFY transcripts were detected in the leaf primordia of tobacco, Arabidopsis, pea, Impatiens, tomato, and petunia (Kelly et al., 1995;Blázquez et al., 1997;Hofer et al., 1997;Pouteau et al., 1997;Pnueli et al., 1998;Souer et al., 1998;Molinero-Rosales et al., 1999), but mutant leaf phenotypes have been described only for those species with compound leaves: pea and tomato.Hofer et al. (1997)proposed a common function for UNI in regulating indeterminacy during both leaf and flower development. An opposite role in specifying lateral organ determinacy was suggested for the tobacco homolog NICOTIANA FLO/LFY ( NFL ;Kelly et al., 1995), although no loss-of-function tobacco mutant has been identified to support this role in leaf development. ...
... Of the dicots studied to date, FLO/LFY transcripts were detected in the leaf primordia of tobacco, Arabidopsis, pea, Impatiens, tomato, and petunia (Kelly et al., 1995;Blázquez et al., 1997;Hofer et al., 1997;Pouteau et al., 1997;Pnueli et al., 1998;Souer et al., 1998;Molinero-Rosales et al., 1999), but mutant leaf phenotypes have been described only for those species with compound leaves: pea and tomato.Hofer et al. (1997)proposed a common function for UNI in regulating indeterminacy during both leaf and flower development. An opposite role in specifying lateral organ determinacy was suggested for the tobacco homolog NICOTIANA FLO/LFY ( NFL ;Kelly et al., 1995), although no loss-of-function tobacco mutant has been identified to support this role in leaf development. There are no reported morphological variations in the leaves of lfy or the corresponding petunia mutant, aberrant leaf and flower ( alf) , and neither LFY nor ALF is thought to play a positive role in leaf development (Weigel et al., 1992;Souer et al., 1998). ...
The compound leaf primordium of pea represents a marginal blastozone that initiates organ primordia, in an acropetal manner, from its growing distal region. The UNIFOLIATA (UNI) gene is important in marginal blastozone maintenance because loss or reduction of its function results in uni mutant leaves of reduced complexity. In this study, we show that UNI is expressed in the leaf blastozone over the period in which organ primordia are initiated and is downregulated at the time of leaf primordium determination. Prolonged UNI expression was associated with increased blastozone activity in the complex leaves of afila (af), cochleata (coch), and afila tendril-less (af tl) mutant plants. Our analysis suggests that UNI expression is negatively regulated by COCH in stipule primordia, by AF in proximal leaflet primordia, and by AF and TL in distal and terminal tendril primordia. We propose that the control of UNI expression by AF, TL, and COCH is important in the regulation of blastozone activity and pattern formation in the compound leaf primordium of the pea.
... The LFY gene has been cloned and its function has been studied from a few species viz. Antirrhinum [16], tobacco [17], Eucalyptus [18], Pinus [19], rice [20], tomato [21], poplar [14], violet cress [22], apple [23], grapevine [24], papaya [25], rubber [26] and cedar [27]. ...
... thaliana, A. majus, pea, tomato, petunia and poplar) [6,14,16,21,31,32]. In apple genome here are two homologous copies of LFY genes (AFL1 t and AFL2) [23] and so also in tobacco (NFL1 and NFL2) [17]. Eucalyptus has three homologous genes in the genome, but two of them were found not to be expressed and had stop codons in their coding region [18]. ...
... This is similar to the expression pattern observed in A. thaliana and A. majus [6,16]. oleracea, ELF gene from Eucalyptus, AFL1 gene from apple and NFL genes from tobacco) have not been reported to express in leaf and stem [17,18,23,34]. ...
... The LFY gene has been cloned and its function has been studied from a few species viz. Antirrhinum [16], tobacco [17], Eucalyptus [18], Pinus [19], rice [20], tomato [21], poplar [14], violet cress [22], apple [23], grapevine [24], papaya [25], rubber [26] and cedar [27]. ...
... thaliana, A. majus, pea, tomato, petunia and poplar) [6,14,16,21,31,32]. In apple genome here are two homologous copies of LFY genes (AFL1 t and AFL2) [23] and so also in tobacco (NFL1 and NFL2) [17]. Eucalyptus has three homologous genes in the genome, but two of them were found not to be expressed and had stop codons in their coding region [18]. ...
... This is similar to the expression pattern observed in A. thaliana and A. majus [6,16]. oleracea, ELF gene from Eucalyptus, AFL1 gene from apple and NFL genes from tobacco) have not been reported to express in leaf and stem [17,18,23,34]. ...
A LEAFY cDNA was cloned from the short-day (SD) plant Brassica juncea cv Varuna. (BjLFY) consists of 1261 bp encoding for a protein of 420 amino acids and an estimated isoelectric point of 6.8. The deduced amino acid showed 99% and 86% identity to Arabi-dopsis thaliana and Brassica oleracea LFY cDNA respectively. The LFY transcript was detected throughout the vegetative and reproductive phase but an increase in transcript level was observed during transition. The earlier induction of BjLFY was observed in early flowering variety of B. juncea as compared to late flowering varieties further proving the critical role LEAFY plays in floral transition.
... However, the expression patterns of LFY/FLO homologs vary greatly. In tobacco, NFL is expressed in the vegetative and floral meristems, while there is no apparent upregulation during floral induction even in Maryland Mammoth, which is an absolute short-day variety (Kelly et al., 1995). By contrast, FLO expression in Antirrhinum is confined exclusively to the inflorescence and floral meristem (Bradley et al., 1996). ...
... The genomic DNA blot hybridization analysis using a CnFL probe in high stringency conditions detected a single band, which suggests that a single copy of the CnFL gene is present in the C. nankingense genome (Fig. 3). This is similar to most LFY homolog genes, although some species have two or more copies in their genomes (Archambault and Bruneau, 2004; Wada et al., 2002; Wang et al., 2008; Kelly et al., 1995). ...
... However, the upregulation of CnFL in induced C. nankingense contrasts with the expression of LFY/FLO homologs in other species, such as NFL in Nicotiana tabacum cv. Maryland Mammoth and CrFL in Chenopodium rubrum, which are both obligate photoperiodic short-day plants, although there was apparently no difference in the NFL and CrFL levels in the vegetative parts compared with the floral shoot apical meristems (Kelly et al., 1995; Veit et al., 2004 ...
... There, no statistically significant differences were detected between strong RNAi suppression events and non-suppressed transgenic trees in height, diameter at breast height, or volume index (Klocko et al., 2016a). We were particularly concerned about the effects on vegetative performance in LFY knockouts because of the reports of significant LFY expression in vegetative meristems in Eucalyptus (Dornelas et al., 2004); however, vegetative expression has also been reported in Arabidopsis, Impatiens, pea, petunia, tobacco, and tomato (Blazquez et al., 1997;Bradley et al., 1997;Hempel et al., 1997;Hofer et al., 1997;Kelly et al., 1995;Molinero-Rosales et al., 1999;Pouteau et al., 1997;Souer et al., 1998;Weigel et al., 1992). Despite expression in vegetative meristems, effects on leaf morphology have only been reported in unifoliata mutants (pea) (Blazquez et al., 1997;Bradley et al., 1997;Dornelas et al., 2004;Hempel et al., 1997;Hofer et al., 1997;Kelly et al., 1995;Molinero-Rosales et al., 1999;Pouteau et al., 1997;Souer et al., 1998;Weigel et al., 1992). ...
... We were particularly concerned about the effects on vegetative performance in LFY knockouts because of the reports of significant LFY expression in vegetative meristems in Eucalyptus (Dornelas et al., 2004); however, vegetative expression has also been reported in Arabidopsis, Impatiens, pea, petunia, tobacco, and tomato (Blazquez et al., 1997;Bradley et al., 1997;Hempel et al., 1997;Hofer et al., 1997;Kelly et al., 1995;Molinero-Rosales et al., 1999;Pouteau et al., 1997;Souer et al., 1998;Weigel et al., 1992). Despite expression in vegetative meristems, effects on leaf morphology have only been reported in unifoliata mutants (pea) (Blazquez et al., 1997;Bradley et al., 1997;Dornelas et al., 2004;Hempel et al., 1997;Hofer et al., 1997;Kelly et al., 1995;Molinero-Rosales et al., 1999;Pouteau et al., 1997;Souer et al., 1998;Weigel et al., 1992). ...
Eucalyptus is among the most widely planted taxa of forest trees worldwide. However, its spread as an exotic or genetically engineered form can create ecological and social problems. To mitigate gene flow via pollen and seeds, we mutated the Eucalyptus ortholog of LEAFY (LFY) by transforming a Eucalyptus grandis x urophylla hybrid and two Flowering Locus T (FT) overexpressing (and flowering) lines with CRISPR Cas9 targeting its LFY ortholog, ELFY. We achieved high rates of elfy biallelic knock‐outs, often approaching 100% of transgene insertion events. Frameshift mutations and deletions removing conserved amino acids caused strong floral alterations, including indeterminacy in floral development and an absence of male and female gametes. These mutants were otherwise visibly normal and did not differ statistically from transgenic controls in juvenile vegetative growth rate or leaf morphology in greenhouse trials. Genes upstream or near to ELFY in the floral development pathway were overexpressed, whereas floral organ identity genes downstream of ELFY were severely depressed. We conclude that disruption of ELFY function appears to be a useful tool for sexual containment, without causing statistically significant or large adverse effects on juvenile vegetative growth or leaf morphology.
... Results showed that derived LFY homologues of PmLFY1 contained a typical intron/exon distribution and a highly conserved region at the N-and C-terminal. At the N-terminal, the plant also has an alanine-rich region and a leucine zipper structure same as most woody plants have, while in the middle region there is a lysine and arginine-rich alkaline region and a region rich in glutamic acid and aspartic ammonia (Carmona et al. 2002;Dornelas et al. 2004;Rottmann et al. 2000;Wada et al. 2002), while proline-rich region at the N-terminal of most of the herbaceous plants (Coen et al. 1990;Kelly et al. 1995;Weigel et al. 1992). It suggests that this region may be a regulatory region for transcriptional activation. ...
... However, there are many homologous genes in multicopy form, such as three LFY homologous gene copies in Eucalyptus (Dornelas et al. 2004), but only one of them was shown to be functional, and the other two had no function. In diploid tobacco, LFY is present as a single copy, and two LFY homologous genes in tetraploid tobacco which have a homology of 96% (Ahearn et al. 2001;Kelly et al. 1995). There are also two LFY homologues genes, NEEDLY and PRFLL, in monterey pine (Mellerowicz et al. 1998;Mouradov et al. 1998). ...
The flowering transition is a qualitative change in the life cycle of higher plants and an important turning point in the transition from vegetative to reproductive growth. Despite the significant importance, the molecular regulation mechanism of the flowering transition of mei (Prunus mume) remains poorly defined. In this study, RT-PCR was used to clone a LFY (LEAFY)-like gene in P. mume ‘Changrui Lve’ and functional characterisation of the gene was carried out. The gene showed high homology to the homologous genes of other woody plants, suggesting that PmLFY-like gene exhibits relatively high conservation as flowering-related genes. Expression of PmLFY1 gene in different organs by real-time quantitative PCR showed that this gene was highly expressed in the late stage rather than early stages of flower bud differentiation in P. mume, thereby exhibiting an active regulatory role in flower organ differentiation. Overexpression of LFY1 in Arabidopsis exhibited phenomenal phenotypic changes in transgenic plants, wherein PmLFY-like gene demonstrated the ability to advance the flowering and promoted the transition of inflorescence meristem to floral meristem. Thus, the PmLFY-like gene was not only involved in regulating the duration of flowering and transition to flowering but also played a role during all stages of inflorescence and flower development. These results can be used to conduct a comprehensive and systematic study of the gene functions which can lay the foundation for the molecular research of flowering in mei.
... In the meristem and inflorescence tissues, the OpLFY2 and OpL-FY2v transcript accumulation was about 12%-14% and 16%-44% of OpLFY1, respectively, in the calibrator tissue. OpLFY transcripts were not observed in the root and leaf tissues, as it has been found for LFY homologs in Arabidopsis, Antirrhinum and tobacco [40]. A high abundance of OpLFY transcripts in male and female inflorescence implies participation of OpLFY in the oil palm floral development pathway. ...
... The qPCR results showed expression of OPLYF transcripts in the shoot apical meristem (SAM). The OpLFY transcripts are predicted to be localised at the peripheral zone of the SAM, where the cells divide and differentiate into lateral organs, similar to the expression of NFL1, a LFY ortholog, in tobacco [40]. Further investigation on its spatial localisation through in situ hybridisation will enable the identification of zone-specific expression of OpLFY at the SAM. ...
LEAFY (LFY) is an important regulator of plant's reproductive system, regulating the transition from vegetative growth to flowering, and also acts as an upstream regulator of floral homeotic MADS-box genes in Arabidopsis. Owing to its importance in flower development, oil palm LFY transcripts were isolated and the expression pattern was characterised to elucidate the role of LFY in oil palm. In addition, the gene expression pattern was analysed to determine a possible correlation of LFY to a type of floral abnormality in clonal palms termed mantling. In this study, three LFY transcripts were isolated from oil palm inflorescence tissue. The isolated cDNAs, OpLFY1, OpLFY2 and its splice variant, OpLFY2v were predicted to encode proteins each consisting of 430, 451 and 181 amino acids, respectively. OpLFY1 and OpLFY2 share 90% and 94% amino acid sequence identity to two different LFYs from Phoenix dactylifera, respectively. OpLFY1 was localised in all floral organ primordia. Quantitative polymerase chain reaction (qPCR) analysis revealed that OpLFY genes are expressed in shoot apical meristem and inflorescence tissues with the highest expression level. The expression of the OpLFY1 gene was significantly lower in stage 1 of inflorescence development in mantled palms compared with that of normal inflorescences, suggesting possible association of the gene with this somaclonal variation.
... In aspen and citrus trees, constitutive expression of the LFY gene can overcome the trees' long juvenile growth phase and quickly shifts the trees to the reproductive phase (Weigel and Nilsson 1995;Peña et al. 2001). LFY orthologs have also been identified in pine, Gnetum, maize, grape, and tobacco, and molecular and morphological analyses indicate that these LFY orthologs function in the transition from vegetative growth to reproductive growth in all species studied (Kelly, Bonnlander, and Meeks-Wagner 1995;Mouradov et al. 1998;Shindo et al. 2001;Carmona, Cubas, and Martinez-Zapater 2002;Bomblies et al. 2003;Dornelas and Rodriguez, 2005). ...
... Duplication of LFY is previously thought to be rare in angiosperms (Cronk 2001). However, more and more plants have been reported to possess more than one copy of LFY (e.g., tobacco, eucalypt, apple, corn, and some Lamialean plants), and these duplicate copies are usually considered results of whole genome duplication (polyploidization) events (Kelly et al. 1995;Southerton et al. 1998;Wada et al. 2002;Bomblies et al. 2003;Aagaard et al. 2005). Viola pubescens is a diploid plant with n ¼ 6 chromosomes, in a Northern Hemisphere sublineage of the genus characterized by a base chromosome number of X ¼ 6. ...
Many plants, including most species in Viola, produce both open, outcrossing chasmogamous and closed, self-pollinating cleistogamous flowers. The chasmogamous/cleistogamous mixed breeding system is considered an evolutionarily successful reproductive strategy, but the underlying molecular basis remains largely unknown. The LEAFY (LFY) gene in Arabidopsis is responsible for the initiation of floral meristems and the regulation of flower development; therefore, we sought to identify LFY orthologs in Viola pubescens Aiton with the goal of understanding possible differences in the regulatory genetics of flower development and the molecular genetics underlying chasmogamous/cleistogamous mixed breeding systems. A genomic library of V. pubescens was constructed to identify LFY orthologs, and reverse transcription polymerase chain reaction was employed to study gene expression in both flower types. In addition, overexpression studies of the orthologs were carried out in Arabidopsis to explore their role in the breeding system of Viola. Two LFY orthologs, VpLFY1 and VpLFY2, were isolated from a genomic library of V. pubescens. Both genes were expressed in both flower types, especially in young floral buds. However, over-expression of VpLFY2, but not VpLFY1, caused precocious flowering in Arabidopsis, indicating a potentially unique role of each ortholog. The results suggest that VpLFY1 and VpLFY2 function together in flower development of V. pubescens. The different effects caused by VpLFY1 and VpLFY2 in Arabidopsis indicate that the two orthologs may have unique, gene-specific properties that might contribute to the flower type in violets.
... In aspen and citrus trees, constitutive expression of the LFY gene can overcome the trees' long juvenile growth phase and quickly shifts the trees to the reproductive phase (Weigel and Nilsson 1995;Peña et al. 2001). LFY orthologs have also been identified in pine, Gnetum, maize, grape, and tobacco, and molecular and morphological analyses indicate that these LFY orthologs function in the transition from vegetative growth to reproductive growth in all species studied (Kelly, Bonnlander, and Meeks-Wagner 1995;Mouradov et al. 1998;Shindo et al. 2001;Carmona, Cubas, and Martinez-Zapater 2002;Bomblies et al. 2003;Dornelas and Rodriguez, 2005). ...
... Duplication of LFY is previously thought to be rare in angiosperms (Cronk 2001). However, more and more plants have been reported to possess more than one copy of LFY (e.g., tobacco, eucalypt, apple, corn, and some Lamialean plants), and these duplicate copies are usually considered results of whole genome duplication (polyploidization) events (Kelly et al. 1995;Southerton et al. 1998;Wada et al. 2002;Bomblies et al. 2003;Aagaard et al. 2005). Viola pubescens is a diploid plant with n ¼ 6 chromosomes, in a Northern Hemisphere sublineage of the genus characterized by a base chromosome number of X ¼ 6. ...
... Both LFY and AP1 genes have been identifi ed in members of diverse plant families, and in some cases, have been shown to be functionally equivalent, indicating that there is some conservation of fl oral regulatory signaling pathways among plant families (Kelly et al., 1995;Mena et al., 1995). Consistent with this, overexpression of either AtLFY or AtAP1 is suffi cient to promote precocious fl owering in distantly related species, including citrus (Pena et al., 2001;Rottmann et al., 2000;Weigel and Nilsson, 1995). ...
... Consistent with other studies, CsLFY expression was restricted to fl oral organs and whole stems, which included early developing fl oral infl orescences. However, some LFY homologues have been detected in early developing vegetative leaf primordia using in situ hybridization (Kelly et al., 1995;Mellerowicz et al., 1998;Mouradov et al., 1998;Walton et al., 2001). The CsAP1 transcript was detected in all four fl oral organ whorls. ...
Homologues of the floral meristem identity genes LEAFY (LFY) and APETALA1 (AP1) were isolated from the hybrid perennial tree crop 'Washington' navel orange (Citrus sinensis) and designated CsLFY and CsAP1, respectively. Citrus has an extended juvenile period unlike herbaceous plants and responds to different floral stimuli than herbaceous plants or deciduous tree species. Despite these differences, the deduced amino acid sequences of CsLFY and CsAP1 genes are at least 65% identical with their Arabidopsis thaliana L. Heynh counterparts and share even greater sequence similarity to LFY and AP1 from the deciduous woody perennials, Populus balsamifera Bradshaw and Populus tremuloides Michaux, respectively. Like A. thaliana LFY (AtLFY) and AP1 (AtAP1), CsLFY and CsAP1 expression was restricted almost exclusively to reproductive tissues, but observed expression of CsAP1 in the fourth whorl carpel tissue of mature flowers was distinct from other plant AP1 genes. Transgenic A. thaliana plants ectopically expressing CsLFY or CsAP1 showed early-flowering phenotypes similar to those described for overexpression of AtLFY and AtAP1. In addition, the 35S:CsLFY and 35S:CsAP1 transgenes partially complemented the lfy-10 and apl-3 mutants, respectively. The severity of the overexpression phenotypes correlated with the accumulation of CsLFY or CsAP1 transcripts. LFY is a single-copy gene in flowering plants but consistent with its hybrid origin, the genome of C. sinensis 'Washington' has two easily distinguishable CsLFY and CsAP1 alleles derived from it's parental genotypes, C. maxima L. Osbeck (pummelo) and C. reticulata Blanco (mandarin). Allelic polymorphism at both the CsLFY and CsAP1 loci was restricted to the 5′- and 3′-flanking regions.
... However, the expression patterns of LFY/FLO homologues vary widely. NFL in tobacco is expressed in both vegetative and floral meristems, with no apparent up-regulation on floral induction even in Maryland Mammoth, the absolute short day variety [18]. By contrast, FLO expression in Antirrhinum is confined exclusively to the inflorescence and floral meristems [19]. ...
... The up-regulation of DFL in induced C. lavandulifolium contrasts with the expression of LFY/FLO homologues in other species such as NFL in N. tabacum cv. Maryland Mammoth and CrFL in Chenopodium rubrum, two obligate photoperiodic SD plant, where there is apparently no difference in levels of NFL and CrFL in vegetative compared with floral SAMs [18,21]. In Antirrhinum, a facultative LD plant, FLO transcription is absolutely correlated with floral induction and is completely absent in vegetative meristems even when measured by RT-PCR [28]. ...
he expression of the floral regulators DFL, a LFY/FLO homologue from Chrysanthemum lavandulifolium was examined during short day light treatments. Quantitative real-time RT-PCR experiments showed that DFL was expressed in the vegetative apices and throughout the shoot apex following photoperiodic induction. After 5 days of induction, DFL expression was increased markly and expressed in highest levels after 20 days induction. Expression of DFL in the shoot apex at the time of floral determination indicating that DFL gene is involved in the first steps of the transition from vegetative to reproductive development.
... Expression of LFY homologues in the vegetative shoot apical meristem and leaf primordia has previously been reported in many angiosperms, such as Nicotiana tabacum (Kelly et al., 1995), Arabidopsis (Blázquez et al., 1997;Hempel et al., 1997), Impatiens sp. (Pouteau et al., 1997), Solanum lycopersicum (Molinero-Rosales et al., 1999), Ionopsidium acaule (Shu et al., 2000), Populus (Rottmann et al., 2000), Malus domestica (Wada et al., 2002), Carica papaya (Yu et al., 2005), Idahoa (Sliwinski et al., 2007), Silene (Allnutt et al., 2007), Oryza sativa (Rao et al., 2008) and Dendranthema (Ma et al., 2008), which suggests a role of LFY in leaf organogenesis. ...
... Li et al. (2013) demonstrated that LFY stimulates the formation of floral and leaf primordia through auxin-regulated pathways. Expression of LFY homologues in developing leaves has also been reported in many investigated angiosperm species (Kelly et al., 1995;Bradley et al., 1997;Blázquez et al., 1997;Hofer et al., 1997;Pouteau et al., 1997;Molinero-Rosales et al., 1999;Rottmann et al., 2000;Souer et al., 1998;Southerton et al., 1998;Walton et al., 2001;Carmona et al., 2002;Dong et al., 2005;Ma et al., 2008), including Cornus in this study (Fig. 3). In pea and tomato plants, lfy mutants showed changes in leaf morphology, confirming the function of LFY homologues in leaf development (Hofer et al., 1997;Molinero-Rosales et al., 1999). ...
Background and Aims LFY homologues encode transcription factors that regulate the transition from vegetative to reproductive growth in flowering
plants and have been shown to control inflorescence patterning in model species. This study investigated the expression patterns
of LFY homologues within the diverse inflorescence types (head-like, umbel-like and inflorescences with elongated internodes) in
closely related lineages in the dogwood genus (Cornus s.l.). The study sought to determine whether LFY homologues in Cornus species are expressed during floral and inflorescence development and if the pattern of expression is consistent with a function
in regulating floral development and inflorescence architectures in the genus.
... Nicotinia, Petunia hybrida, and Impatiens balsimina are likely examples of plants whose ability to respond to LFY activity is altered, as LFY homologues have been found to be expressed in both vegetatively and florally induced apices of these plants [138][139][140]. The genes PENNYWISE (PNY) and POUND-FOOLISH (PNF) have also been identified as being important in regulating the apex's ability to respond to floral inducing cues. ...
Plants are extremely sensitive to changes in their environment, particularly variations in photoperiod or day length. Photoperiodism refers to a plant's capacity to detect variations in day length and make use of this knowledge to control key developmental processes including flowering, growth, and dormancy. Through a process known as photoperiodism, plants can detect and react to variations in the number of daylight hours, or photoperiod. The physiological response of plants to the length of day or night is known as photoperiodism. The plant uses this physiological response to time-critical developmental events like flowering. In this essay, I will cover the current understanding of how plants respond to photoperiod and the molecular mechanisms underpinning this response. Three groups of plants' photoperiodic responses can be distinguished: short-day plants (SDPs), long-day plants (LDPs), and day-neutral plants (DNPs). Whereas LDPs bloom when the length of the day exceeds the crucial threshold, SDPs do so only when it is shorter than the critical threshold. Conversely, DNPs do not have a crucial day duration and can bloom at any day length. Many genes and biochemical processes control how a plant responds to the photoperiod. The creation and movement of the hormone florigen, which starts blooming in response to photoperiodic signals, is a crucial regulating mechanism. On the other hand, a class of photoreceptors known as phytochromes is involved in the biochemical mechanisms driving photoperiodic responses in plants. The perception of light's duration, quality, and amount is caused by phytochromes. The red-light-absorbing Pr form and the far-red-light-absorbing Pfr form are the two interconvertible states in which they can exist. The ratio of Pr to Pfr is altered by the duration of light exposure and is utilizes by plants to assess day length. Exposure to light in SDPs causes the expression of the CONSTANS (CO) gene, and the CO protein causes the expression of the FLOWERING LOCUS T (FT), a gene that encourages flowering. By exposing LDPs to light, a different gene called GI (GIGANTEA) is induced rather than CO, which is normally expressed. The FT gene's expression is encouraged by GI's interaction with the protein ZEITLUPE (ZTL), which also encourages flowering. In addition to these essential elements, several proteins and signalling pathways are also involved in photoperiodic responses in plants. For instance, to optimise the response to variations in day length, the photoperiodic pathway interacts with the circadian clock, which controls numerous physiological processes in plants. In some species, the hormone gibberellin (GA) also aids in the promotion of flowering. One essential adaptation that enables plants to synchronize their developmental processes with seasonal changes is their capacity to react to variations in day length. Phytochromes play a key role in how plants perceive the day in the complex network of proteins and signalling channels that make up the molecular mechanisms behind photoperiodic responses in plants. There is still much to learn about the diversity and complexity of the photoperiodic response across several plant groupings, even if much is known about it in particular species.
... On the other hand, a LFY homolog from a conifer, NEEDLY (NLY), appears to function similarly to its dicot counterparts. In contrast to RFL, NLY is expressed in primordia arising on the flanks of both the vegetative and reproductive shoot apical meristem (Mouradov et al., 1998), a pattern that is reminiscent of LFY in Arabidopsis and its ortholog NFL in tobacco (Kelly et al., 1995;Blázquez et al., 1997). While NLY RNA levels do not increase during the vegetative to reproductive transition, similar to what is reported for tobacco NFL, constitutive expression of NLY in transgenic Arabidopsis has the same effect as constitutive expression of LFY. ...
Onset of flowering, or heading date, is an important agronomic trait of cereal crops such as rice and early-heading varieties are required for certain regions in which rice is cultivated. Since the floral control gene LEAFY from Ara-bidopsis can dramatically accelerate flowering in dictoyledonous plants, the usefulness of LEAFY for manipulating heading date in rice has been tested. Constitutive expression of LEAFY from the cauliflower mosaic virus 35S promoter caused early flowering in transgenic rice, with a heading date that was 26-34 days earlier than that of wild-type plants. Early flowering was accompanied by a small yield penalty and some panicle abnormality. These observations suggest that floral regulatory genes from Arabidopsis are useful tools for heading date improvement in cereal crops.
... We used an ubiquitin gene (UBQ2) as an internal reference (Mohamed et al., 2010) and normalized the Ct values across plates, determining relative quantities using comparative Ct method (2 −ΔΔCt ) as previously described (Livak and Schmittgen, 2001). For in situ hybridization, immature inflorescences were collected from wild Populus trichocarpa trees near Corvallis, OR, United States, and fixed and embedded as previously described (Kelly et al., 1995). Transcripts were detected using antisense riboprobes from the 3′ ends (Supplementary Figure S2) of P. trichocarpa LAP1a (396 bp) and LAP1b (360 bp) cDNAs. ...
Both the evolution of tree taxa and whole-genome duplication (WGD) have occurred many times during angiosperm evolution. Transcription factors are preferentially retained following WGD suggesting that functional divergence of duplicates could contribute to traits distinctive to the tree growth habit. We used gain- and loss-of-function transgenics, photoperiod treatments, and circannual expression studies in adult trees to study the diversification of three Populus FLOWERING LOCUS D-LIKE (FDL) genes encoding bZIP transcription factors. Expression patterns and transgenic studies indicate that FDL2.2 promotes flowering and that FDL1 and FDL3 function in different vegetative phenophases. Study of dominant repressor FDL versions indicates that the FDL proteins are partially equivalent in their ability to alter shoot growth. Like its paralogs, FDL3 overexpression delays short day-induced growth cessation, but also induces distinct heterochronic shifts in shoot development—more rapid phytomer initiation and coordinated delay in both leaf expansion and the transition to secondary growth in long days, but not in short days. Our results indicate that both regulatory and protein coding sequence variation contributed to diversification of FDL paralogs that has led to a degree of specialization in multiple developmental processes important for trees and their local adaptation.
... For example, many herbaceous flowering plants show low levels of LFY expression in a range of vegetative tissues. NFL, the LFY homolog from Nicotiana tabacum (tobacco), is expressed in vegetative meristems [6]. Similarly, FA, the Lycopersicon esculentum (tomato) homolog is expressed in vegetative meristems, leaf primordia, and leaves [7]. ...
The central floral development gene LEAFY (LFY), whose mutation leads to striking changes in flowering and often sterility, is commonly expressed in non-floral structures; however, its role in vegetative development is poorly understood. Sterility associated with suppression of LFY expression is an attractive means for mitigating gene flow by both seeds and pollen in vegetatively propagated forest trees, but the consequences of its suppression for tree form and wood production are unclear. To study the vegetative effects of RNAi suppression of LFY, we created a randomized, multiple-year field study with 30–40 trees (ramets) in each of two sterile gene insertion events, three transgenic control events, and a wild-type control population. We found that floral knock-down phenotypes were stable across years and propagation cycles, but that several leaf morphology and productivity traits were statistically and often substantially different in sterile vs. normal flowering RNAi-LFY trees. Though trees with suppressed LEAFY expression looked visibly normal, they appear to have reduced growth and altered leaf traits. LFY appears to have a significant role in vegetative meristem development, and evaluation of vegetative impacts from LFY suppression would be prudent prior to large-scale use for genetic containment.
... For example, the wild P. regulatory gene of the floral meristem. To date, LEAFY homologous genes, as an example LFY, FLO, NFL, and RFL, have been isolated from many angiosperms (Coen et al., 1990;Kelly et al., 1995;Weigel et al., 1995；Kyozuka et al., 1998. The LFY gene is the earliest expression of floral meristem-specific gene, and AP1/CAL and other genes are their downstream target genes (William et al., 2004). ...
Flowering is an important stage in the life cycle of plants and also a turning point from vegetative growth to reproductive growth. This process is affected by many exogenous and endogenous factors. Some examples of the latter are endogenous hormones, plant growth status, nutrient composition, and flowering regulatory genes. Many gymnosperms have a long juvenile period. Previous studies attempted to shorten this period using traditional asexual propagation methods, but significant results have not been achieved. In recent years, molecular biology is used to study the flowering regulatory gene to obtain transgenic plants with early flowering trait. Thus, the production of gymnosperms is hastened, and economic efficiency is improved. Studies have shown that the flowering genes of plants act synergistically to form a complex network. In this paper, we reviewed the recent development in the study of the regulation of the flowering genes of gymnosperms, that is, from the floral meristem-specific gene, floral organ-specific gene, genes that inhibit plant flowering, and microRNA regulation of flowering. We provide a reference for the in-depth study on the genetic improvement of the flowering gene.
... The dataset was pruned and then supplemented with further sequences (Supplementary file 1) to enable trees to be inferred that would (i) provide a more balanced distribution across the major plant groups and (ii) infer fern relationships. Only a subset of available angiosperm sequences was retained (keeping both monocot and dicot representatives) but protein sequences from other angiosperm species where function has been defined through loss-offunction analyses were added from NCBI -Antirrhinum majus FLO AAA62574.1 , Pisum sativum UNI AAC49782.1 (Hofer et al., 1997), Cucumis sativus CsLFY XP_004138016.1 (Zhao et al., 2018), Medicago truncatula SGL1 AY928184 (Wang et al., 2008), Petunia hybrida ALF AAC49912.1 (Souer et al., 1998), Nicotiana tabacum NFL1 AAC48985.1 and NFL2 AAC48986.1 (Kelly, 1995), Eschscholzia californica EcFLO AAO49794.1 (Busch and Gleissberg, 2003), Gerbera hybrida cv. 'Terraregina' GhLFY ANS10152.1 (Zhao et al., 2016), Lotus japonicus LjLFY AAX13294.1 (Dong et al., 2005) and Populus trichocarpa PTLF AAB51533.1 (Rottmann et al., 2000). ...
During land plant evolution, determinate spore-bearing axes (retained in extant bryophytes such as mosses) were progressively transformed into indeterminate branching shoots with specialized reproductive axes that form flowers. The LEAFY transcription factor, which is required for the first zygotic cell division in mosses and primarily for floral meristem identity in flowering plants, may have facilitated developmental innovations during these transitions. Mapping the LEAFY evolutionary trajectory has been challenging, however, because there is no functional overlap between mosses and flowering plants, and no functional data from intervening lineages. Here, we report a transgenic analysis in the fern Ceratopteris richardii that reveals a role for LEAFY in maintaining cell divisions in the apical stem cells of both haploid and diploid phases of the lifecycle. These results support an evolutionary trajectory in which an ancestral LEAFY module that promotes cell proliferation was progressively co-opted, adapted and specialized as novel shoot developmental contexts emerged.
... Copy number was determined by Digital Droplet PCR (Glowacka et al., 2016) using primer and probes specific for the HPH selectable marker and N. tabacum NFL2 reference gene (Kelly et al., 1995) (Accession #AH006599). BamHI and HindIII digested genomic DNA was added to 2x ddPCR mastermix at concentrations between 20 and 120 ng DNA per 20 µL PCR reaction. ...
Synthesis and accumulation of plant oils in the entire vegetative biomass offers the potential to deliver yields surpassing those of oilseed crops. However, current levels still fall well short of those typically found in oilseeds. Here we show how transcriptome and biochemical analyses pointed to a futile cycle in a previously established Nicotiana tabacum line, accumulating up to 15 % (dry weight) of the storage lipid triacylglycerol in leaf tissue. To overcome this metabolic bottleneck, we either silenced the SDP1 lipase or overexpressed the Arabidopsis thaliana LEC2 transcription factor in this transgenic background. Both strategies independently resulted in the accumulation of 30-33 % triacylglycerol in leaf tissues. Our results demonstrate that the combined optimization of de novo fatty acid biosynthesis, storage lipid assembly and lipid turnover in leaf tissue results in a major overhaul of the plant central carbon allocation and lipid metabolism. The resulting further step changes in oil accumulation in the entire plant biomass offers the possibility of delivering yields that outperform current oilseed crops.
... LFY homologues are highly conserved, but their expression patterns differ among species. Unlike FLO, which is expressed only in floral meristems in Antirrhinum (Coen et al., 1990), LFY homologues are expressed in both vegetative and floral meristems in Arabidopsis, tobacco, tomato, and pea (Weigel et al., 1992;Kelly et al., 1995;Blazquez et al., 1997;Hofer et al., 1997). ...
Recurrent flowering is an important characteristic in modern roses. Previous studies indicate that RoTFL1 (RTFL1c, RoKSN) plays an important role in this trait, but homologues of its downstream target gene LEAFY (LFY) have not been identified in roses. In this study, we isolated LFY homologues from three Rosa species (here collectively designated Rosa LEAFY [RoLFY]) with recurrent or non-recurrent flowering habits. The RoLFY genes were isolated from Rosa chinensis, R. multiflora, and R. rugosa using a combination of degenerate and gene-specific primers by thermal asymmetric interlaced-PCR and normal PCR. The full-length cDNA was 2142 bp. The coding sequence was 1242 bp and encoded 413 amino acids. The sequence identity among the three species was 96.1%, compared with 57.2% with Arabidopsis LFY. A phylogenetic analysis clustered the RoLFY proteins in one group, which was most similar to LFY homologues of strawberry (FaLFY) within the Rosaceae. The deduced tertiary structure of the RoLFY proteins was almost identical and the domains were consistent with those of FaLFY-2 from strawberry. The RoLFY protein was localized in the nucleus consistent with other transcription factors. Over-expression of RoLFY promoted reproductive growth and earlier flowering in transgenic Arabidopsis. Marked differences in expression levels were detected during early flower bud development, but not at subsequent stages, among the non-recurrent flowering R. multiflora and R. rugosa, and the recurrent flowering R. chinensis. These results suggest that once floral initiation is complete, recurrent-flowering roses can flower continuously without repeated accumulation of RoLFY mRNA.
... It was expressed in floral meristems and in developing floral organs. The pattern was similar to the expression of most of the FLO/LFY homologue genes in dicotyledon, such as tobacco, petunia, tomato, grapevine, Papaveraceae, Impatiens and Eschscholzia californica (Kelly et al., 1995;Pouteau et al., 1997;Souer et al., 1998;Southerton et al., 1998;Molinero-Rosales et al., 1999;Carmona et al., 2002;Andrea and Gleissberg, 2003). However, the expression patterns of monocot FLO/LFY homologues are even more divergent from dicotyledonous counterparts. ...
Ten polymorphic microsatellite loci were isolated and characterized from an enriched genomic library of Paphiopedilum concolor (Batem.) Pfitzer. The number of alleles per microsatellite locus ranged from three to 11 with an average of 6.4 in a sample of 30 individuals from three populations. The observed and expected heterozygosity ranged from 0.200 to 0.800 and from 0.544 to 0.827, respectively. These microsatellites can be used as tools to investigate the genetic structure of P. concolor populations and relationship patterns with closely related taxa.
... LEAFY is a homeotic gene that regulates floral meristem induction during the early stages of reproductive ontogeny (Schultz & Haughn, 1991;Weigel, 1995;Bl azquez, 1997). It is considered to have only a single copy in diploid angiosperms (Frohlich & Parker, 2000;Himi et al., 2001), although the copy number can vary in polyploids (Kelly et al., 1995;Frohlich & Parker, 2000;Wada et al., 2002). ...
Low copy nuclear genes show great potential to provide phylogenetic information, but their use has been hampered by several inherent adverse factors. Polymerase chain reaction (PCR)-mediated recombination ranks among these factors and occurs when high levels of similar paralogs for a low-copy nuclear gene coexist within a single PCR amplification reaction. In this study, the LEAFY gene was cloned and sequenced for 63 Cinnamomum species and two copies of the second intron were found within species of the diploid sect. Cinnamomum. Although these two copies are very similar, they can be distinguished easily due to a specific ca. 47-bp segment that is missing in the "short sequence" copy. The "long sequence" copy performed well in phylogenetic analysis of Cinnamomum and is largely consistent with phylogenetic relationships based on internal transcribed spacer region sequences. In contrast, the "short sequence" copy was problematic for phylogenetic reconstruction and PCR-mediated recombination was detected in 20 of the 38 Cinnamomum species with two LEAFY copies. Thirty-one recombinants were discriminated and the breakpoints suggested by the programs RDP3 and GARD were distributed randomly along the recombination sequences. This study shows that duplication in low-copy nuclear genes and problems associated with PCR-mediated recombination need to be given more attention in phylogenetic studies.
... However, for most, there is no information on their parental origin or transcription activity. Both progenitor homoeologs of the floral meristem identity gene FLORICAULA/ LEAFY have been isolated in tobacco, and both are transcriptionally active in the shoot apex (Kelly et al. 1995). Three floral organ identity genes, B-class DEFICIENS and GLOBOSA and C-class AGAMOUS, have been identified and functionally characterized in tobacco (Hansen et al. 1993;Kempin et al. 1993;Davies et al. 1996); however, only a single cDNA sequence has been reported for each gene. ...
In plants, genome duplication followed by genome diversification and selection is recognized as a major evolutionary process. Rapid epigenetic and genetic changes that affect the transcription of parental genes are frequently observed after polyploidization. The pattern of alternative splicing is also frequently altered, yet the related molecular processes remain largely unresolved. Here, we study the inheritance and expression of parental variants of three floral organ identity genes in allotetraploid tobacco. DEFICIENS and GLOBOSA are B-class genes, and AGAMOUS is a C-class gene. Parental variants of these genes were found to be maintained in the tobacco genome, and the respective mRNAs were present in flower buds in comparable amounts. However, among five tobacco cultivars, we identified two in which the majority of paternal GLOBOSA pre-mRNA transcripts undergo exon 3 skipping, producing an mRNA with a premature termination codon. At the DNA level, we identified a G-A transition at the very last position of exon 3 in both cultivars. Although alternative splicing resulted in a dramatic decrease in full-length paternal GLOBOSA mRNA, no phenotypic effect was observed. Our finding likely serves as an example of the initiation of homoeolog diversification in a relatively young polyploid genome.
... Variations in the expression patterns of ALF/LFY homologs are as important for morphological divergence as those of DOT/UFO homologs, but, again, not all the variation relates necessarily to developmental changes. Many species, with few exceptions (Coen et al., 1990), express their LFY homologs in vegetative tissues with different spatio-temporal patterns, where they have no apparent (architectural) role that is obvious from mutant phenotypes (Weigel et al., 1992;Kelly et al., 1995;Souer et al., 1998;Molinero-Rosales et al., 1999), except for a clade of legumes where LFY is involved in the development of compound leaves (Champagne et al., 2007). Several Brassicaceae with (rosette-flowering) indeterminate racemose inflorescences express their LFY homologs in the lateral (floral) meristems, and in the apical IM, which nevertheless remains indeterminate (Shu et al., 2000;Sliwinski et al., 2007). ...
Higher plant species diverged extensively with regard to the moment (flowering time) and the position (inflorescence architecture) where flowers are formed. This seems largely caused by variation in the expression patterns of conserved genes that specify floral meristem identity (FMI), rather than changes in the encoded proteins. Here we report a functional comparison of the promoters of homologous FMI genes from Arabidopsis, petunia, tomato and Antirrhinum. Analysis of promoter-reporter constructs in petunia and Arabidopsis and complementation experiments showed that the divergent expression of LEAFY (LFY) and the petunia homolog ABERRANT LEAF AND FLOWER (ALF) results from alterations in the upstream regulatory network rather than cis-regulatory changes. The divergent expression of UNUSUAL FLORAL ORGANS (UFO) from Arabidopsis and the petunia homolog DOUBLE TOP (DOT), on the other hand, is caused by the loss or gain of cis-regulatory promoter elements, which respond to trans-acting factors that are expressed in similar patterns in both species. Introduction of pUFO:UFO causes no obvious defects in Arabidopsis, but in petunia it causes the precocious and ectopic formation of flowers. This provides an example of how a change in a cis-regulatory region can account for a change in the plant body plan.
© 2015. Published by The Company of Biologists Ltd.
... In situ hybridizations were performed as described by Kelly et al. (1995) with the following modifications. Eightto 10-m sections were probed with either antisense PTD cRNA transcribed using the T7 promoter in the pBlueScript vector or with sense (control) PTD cRNA transcribed from the T3 promoter. ...
We isolated PTD, a member of theDEFICIENS (DEF) family of MADS box transcription factors, from the dioecious tree, black cottonwood (Populus trichocarpa). In females, in situ hybridization experiments showed that PTD mRNA was first detectable in cells on the flanks of the inflorescence meristem, before differentiation of individual flowers was visually detectable. In males, the onset of PTD expression was delayed until after individual flower differentiation had begun and floral meristems were developing. Although PTD was initially expressed throughout the inner whorl meristem in female and male flowers, its spatial expression pattern became sex-specific as reproductive primordia began to form. PTD expression was maintained in stamen primordia, but excluded from carpel primordia, as well as vegetative tissues. Although PTD is phylogenetically most closely related to the largely uncharacterized TM6subfamily of theDEF/APETELA3(AP3)/TM6group, its spatio-temporal expression patterns are more similar to that of DEF and AP3 than to other members of the TM6 subfamily.
... zfl mRNA is expressed throughout early floral meristems and subsequently relegated to developing organ primordia (Fig. 2C-H). This pattern is similar to the floral expression patterns of FLO/LFYlike genes reported in several dicot species (Coen et al., 1990;Hofer et al., 1997;Kelly et al., 1995;Molinero-Rosales et al., 1999;Souer et al., 1998;Weigel et al., 1992). The conserved expression pattern and similar mutant effects on floral development, suggest that dicot FLO/LFY and zfl play a conserved role in floral development. ...
The homologous transcription factors FLORICAULA of Antirrhinum and LEAFY of Arabidopsis share conserved roles in flower meristem identity and floral patterning. While roles for FLORICAULA/LEAFY homologs in flower development have been demonstrated in numerous dicots, little is known about the function of these meristem identity genes in the more distantly related flowering plants, the monocots. We used reverse genetics to investigate the role of two duplicate FLORICAULA/LEAFY homologs in maize (Zea mays L. ssp. mays) – a monocot species with dramatically different flower and inflorescence morphology from that of dicot species. Transposon insertions into the maize genes, zfl1 and zfl2, led to a disruption of floral organ identity and patterning, as well as to defects in inflorescence architecture and in the vegetative to reproductive phase transition. Our results demonstrate that these genes share conserved roles with their dicot counterparts in flower and inflorescence patterning. The phenotype of zfl1; zfl2 double mutants suggests that these maize FLORICAULA/LEAFY homologs act as upstream regulators of the ABC floral organ identity genes, and this along with previously published work, indicates that the transcriptional network regulating flower development is at least partially conserved between monocots and dicots. Our data also suggest that the zfl genes may play a novel role in controlling quantitative aspects of inflorescence phyllotaxy in maize, consistent with their candidacy for quantitative trait loci that control differences in inflorescence structure between maize and its progenitor, teosinte.
... The polyploidy of 'Carnival' and its parental species, however, warrants further investigation. Many studies suggest that multiple copies of LFY may be detrimental for plant development due to the importance of LFY in floral architecture and reproductive success (Kelly et al., 1995;Baum et al., 2005), however, no adverse effects due to the expression of multiple ProLFY copies are evident in 'Carnival'. From heterologous expression studies in A. thaliana one of these paralogous ProLFY genes, ProLFY-WL4 isolated from 'wavy leaf' meristems, was found to be functionally equivalent to AtLFY as it could restore the abnormal floral morphology (Figs. 5 and 6) and fertility of lfy 2 mutant plants (Fig. 7) in the T2 population. ...
a r t i c l e i n f o Proteas are woody perennial plants that develop large, complex inflorescences which are popular as cut flowers. Four paralogous genes with sequence similarity to the meristem identity gene LEAFY (LFY) of Arabidopsis thaliana were identified in the Protea cultivar 'Carnival' (Protea compacta x Protea neriifolia). The deduced amino acid sequences of Protea LFY (ProLFY) paralogues displayed highly conserved N-and C-terminal regions comparable to that of other LFY proteins. Paralogous sequences shared 97–99% similarity at nucleotide and deduced amino acid levels. To determine if these ProLFY paralogues are involved in floral transition and development, three paralogues were heterologously expressed in A. thaliana Col-0 wild-type and lfy-2 mutant plants. These three paralogues were chosen as they had the greatest sequence dissimilarities which may suggest a different function. Expression of the ProLFY-WL4 paralogue rescued the low fertility of the lfy-2 mutant by restoring seed yield to wild-type levels, and resulted in the conversion of 'leaf-like' inflorescences to flowers with a wild-type appearance. ProLFY-WL4 expression levels during 'Carnival' meristem development confirmed a role for this paralogue in inflorescence development. The other two paralogues used in heterologous A. thaliana expression studies, ProLFY-L5 and ProLFY-D1, did not rescue fertility or the abnormal floral phenotype of lfy-2. However, transgenic plants developed either smaller leaves when expressing 35S::ProLFY-L5, or larger leaves when expressing 35S::ProLFY-D1, compared to the non-transgenic lfy-2 plants. Together these results suggest that ProLFY paralogues may be involved in both floral and vegetative development of Protea.
... Expression of loquat LFY gene (ejLFY) in Arabidopsis resulted in early-flowering too (Liu et al., 2008). So far, LFY family genes have been isolated in many plant species, including snapdragon, rice, wheat, pea and so on (Coen et al., 1990;Dornelas and Rodriguez, 2006;Hofer et al., 1997;Kelly et al., 1995;Kyozuka et al., 1998;Shitsukawa et al., 2006). The identified LFY family genes are highly conserved in both their structures and functions (Ma et al., 2005). ...
LEAFY-like genes are crucial regulators of flowering in angiosperms. A partial homologue of LEAFY, designated as AcLFY (Genebank accession no HQ433335), was isolated from pineapple (Ananas comosus L. cv. Comte de Paris) by PCR. The conserved cDNA fragment of AcLFY is 256 bp in length and contained an open reading frame of 248 bp, which encodes 82 amino acids protein. The amino acid alignment showed that AcLFY had a high identity with the related fragment of PhalLFY in Phalaenopsis hybrid cultivar (91%), LFY in Oncidium (90%) and OrcLFY in Orchis italica (88%). RT-PCR analysis showed that the AcLFY could be expressed constitutionally. Moreover, it was expressed in the flesh of young fruit and bract at low levels, and was highly expressed in the fruit, stem, petal and sepal.
... Embora estas observações reflitam uma possível conservação funcional entre os homólogos de FLO/LFY em espécies pouco relacionadas filogeneticamente, variações significativas a respeito dos padrões de expressão destes genes vêm sendo relatadas (Dornelas et al. 2004, Veit et al. 2004. Em Antirrhinum, FLO é expresso exclusivamente durante a fase reprodutiva nas brácteas e meristemas florais (Coen et al. 1990), enquanto que ortólogos deste gene são também expressos em baixas intensidades nos primórdios foliares de outras espécies, como em Arabidopsis (Blázquez et al. 1997), tabaco (Kelly et al. 1995), ervilha (Hofer et al. 1997), petúnia (Souer et al. 1998) e tomate (Rosales et al. 1999). Contrapondo o padrão de expressão aproximado dos ortólogos de FLO/LFY em dicotiledôneas, as espécies de monocotiledôneas reforçam a hipótese de divergência funcional deste gene. ...
Hyptis suaveolens (L.) Poit. (Lamiaceae) is a typical invasive plant that produces an essential oil with several biological properties. The largest essential oil content is found at the flowers. So, advances in the current knowledge about the H. suaveolens reproductive development are important when the essential oil productivity is sought. Furthermore, the comprehension of the mechanisms involved in the H. suaveolens floral induction would be useful to clarify the adaptive responses in invasive plants. We here reported some H. suaveolens reproductive development features. The plants were grown at two photoperiodic treatments (natural photoperiod from Alfenas, Minas Gerais, Brazil and natural photoperiod extended of 16 h). Samples of floral and vegetative aerial apices were taken at five different periods of time, for anatomical and scanning electron microscopic analysis, as well as in situ hybridization experiments, in order to detect the transcripts of a putative ortholog of Arabidopsis thaliana LEAFY gene. Hyptis suaveolens is a short day plant, with critical photoperiod nearby 13 h. The cell density has increased according to higher vegetative or floral meristem activities. The putative ortholog of LEAFY in H. suaveolens has shown an expression pattern similar to that reported in Antirrhinum and Arabidopsis, although it was also expressed in vegetative apices at the natural photoperiod plants. Nevertheless, under extended natural photoperiod (16 h) no gene expression was detected in vegetative meristem. These results suggest a basal expression of LEAFY in H. suaveolens vegetative meristem, which is strongly decreased by extended day length conditions, or is increased under photoperiods lower than 13 h, when suitable levels are reached for floral determination.
... Chez le Tabac (Kelly et a/., 1995) cellules qui quittent Ia zone axiale pour une destinee nouvelle. Cette position tres particuliere des transcrits NFL rappelle celle des zones d'harmonisation de croissance (Lance, 1957). ...
During the last decade progress has been made on the shoot apex morphogenesis. Mutants, whose deficiencies reveal developmental anomalies, were characterised specially on Arabidopsis thaliana, Antirrhinum majus et Zea mays. Several genes were isolated, cloned and their products (mRNA, protein) localised at the level of the shoot apical meristem (SAM). Transgenic expression of reporter genes was also used. These researches allow one to present very new hypotheses and a new insight on the control of apical morphogenesis. Essential results concerning the vegetative shoot apex structure and functioning are presented. However the various types of genes, whose expression has been localised in the SAM, are emphasised. It is now possible to understand how the identity and the fate of cells are acquired to ensure the SAM construction during the embryonic phase, its maintenance during the vegetative phase and its gradual change during the inflorescence development. Early genic events correlated with the preparation to foliar initiation are set out as well as the most recent scenario on the intervention of several types of genes during the vegetative functioning of the SAM.
... Consistent with this interpretation, mutants of UFO orthologs DOUBLE TOP (DOT) and ANANTHA (AN) in the related Solanaceae species petunia and tomato, respectively, are unable to transit into a flower and instead reiterate the inflorescence program. The LFY orthologs NFL1 (tobacco), ABERRANT LEAF AND FLOWER (ALF petunia) and FALSIFLORA (FA tomato) are expressed like LFY in the lateral meristems produced by the vegetative and inflorescence meristems, but the NFL1 transcript levels are higher than those of LFY during the vegetative stage [39,47,48]. The UFO orthologs DOT in petunia and AN in tomato are not expressed in the vegetative meristems, but are first activated in the IMs upon transition to flowering [15,49]. ...
Plants are unique in their ability to continuously produce new meristems and organ primordia. In Arabidopsis, the transcription factor LEAFY (LFY) functions as a master regulator of a gene network that is important for floral meristem and organ specification. UNUSUAL FLORAL ORGANS (UFO) is a co-activator of LEAFY and is required for proper activation of APETALA3 in the floral meristem during the specification of stamens and petals. The ufo mutants display defects in other parts of the flower and the inflorescence, suggestive of additional roles. Here we show that the normal determinacy of the developing Arabidopsis leaves is affected by the expression of a gain-of-function UFO fusion protein with the VP16 transcriptional activator domain. In these lines, the rosette and cauline leaf primordia exhibit reiterated serration, and upon flowering produce ectopic meristems that develop into flowers, bract leaves and inflorescences. These striking phenotypes reveal that developing leaves maintain the competency to initiate flower and inflorescence programs. Furthermore, the gain-of-function phenotypes are dependent on LFY and the SEPALLATA (SEP) MADS-box transcription factors, indicative of their functional interactions with UFO. The findings of this study also suggest that UFO promotes the establishment of the lateral meristems and primordia in the peripheral zone of the apical and floral meristems by enhancing the activity of LFY. These novel phenotypes along with the mutant phenotypes of UFO orthologs in other plant species suggest a broader function for UFO in plants.
The sections in this article are
Introduction
Shoot Development
Organogenesis of the Leaf
Organogenesis of the Root
Conclusions
Acknowledgements
Plant morphogenesis proceeds through a multitude of events which occur according to a spatial and temporal sequence, some of them at specific time and sites (cell / tissue / organ), some gradually or permanently intertwined, some constitutively, and some with switches on and off which are unpredictable by the investigator although “programmed” in the plant. Unlike the animals, a plant which from its birth to its death is confined to grow and to develop at the geographical site of its birth, has to adapt itself to predictable and unpredictable seasonal environmental conditions. Those adaptive mechanisms acquired through the evolution of the species will be switched on or off according to the perception of the signals by the plant. Unlike the animals, the plant signal perception and transduction mechanisms are not as specific. Therefore, using mutants, developmental processes have been more clearly elucidated in animals than in plants.
Many genes are known that, upon mutation, significantly change plant development and architecture in a coordinated fashion. Of particular interest are the floral meristem identity genes, which specify the floral fate of meristems that appear at the flanks of inflorescence meristems. Several genes of this type have been cloned during recent years. Important representatives are FLORICAULA (FLO) and LEAFY (LFY), two orthologous genes from Antirrhinum majus (snapdragon) and Arabidopsis thaliana (thale cress), respectively, which encode members of a novel family of transcription factors. The expression of these genes strongly influences the time to flowering and the number of flowers formed. Studying the origin and evolution of FLO-like meristem identity genes may thus provide new insights into the evolutionary origin of flower development. Moreover, the time to flowering is a critical parameter that significantly contributes to the agronomic value of crop and forest plants. FLO-like genes may thus also be used as suitable molecular tools to design such plants according to our desires. The past few years have seen important progress in our understanding of both aspects of FLO-like genes, their evolution, and their suitability as genetic tools for the agronomic improvement of crop and forest plants.
During the last decade progress has been made on the shoot apex morphogenesis. Mutants, whose deficiencies reveal developmental anomalies, were characterised specially on Arabidopsis thaliana, Antirrhinum majus et Zea mays. Several genes were isolated, cloned and their products (mRNA, protein) localised at the level of the shoot apical meristem (SAM). Transgenic expression of reporter genes was also used. These researches allow one to present very new hypotheses and a new insight on the control of apical morphogenesis. Essential results concerning the vegetative shoot apex structure and functioning are presented. However the various types of genes, whose expression has been localised in the SAM, are emphasised. It is now possible to understand how the identity and the fate of cells are acquired to ensure the SAM construction during the embryonic phase, its maintenance during the Vegetative phase and its gradual change during the inflorescence development. Early genic events correlated with the preparation to foliar initiation are set out as well as the most recent scenario on the intervention of several types of genes during the vegetative functioning of the SAM.
Many floral homeotic genes are expressed early in floral development and exhibit floral-specific expression, making them excellent candidates for use in engineering reproductively sterile plants. We have cloned and partially characterized Populus trichocarpa cDNAs and genes homologous to the Arabidopsis/Antirrhinum genes LEAFY (LFY)/ FLORICAULA (FLO), APETALA3 (AP3)/DEFICIENS (DEF), and AGAMOUS (AG)I PLENA (PLE). The P. trichocarpa LFY homolog, PtFL, shares 75% amino acid identity with LFY. PtD shows 81% amino acid similarity with DEF, and PtAG shows 85% homology withAG. PtFL is expressed in immature inflorescences on which floral primordia are developing. PtD is expressed strongly in stamen primordia from the onset of organogenesis, and is expressed at low levels in carpel primordia.
Terminal flower1 (TFL1) is a key gene in charge of flowering time in Arabidopsis thaliana. It belongs to a family of phosphatidyl ethanolamine binding protein domain (PEBP). The main issues addressed in this paper are current advances in TFL1 homologs isolated from the monocot plants include rice (Oryza sativa) and ryegrass (Lolium perenne). Taking advantage of previous studies of cloning, in this review we have evaluated function of these genes in regulating flowering time as well as the effect of altered expression of these genes. It will then discuss or address places or parts of these plants where these genes express. Additionally, structural and functional relationships between them are compared.
Flowering is important for the production of strawberry, but its molecular mechanism is not clear now. To investigate the molecular mechanism of flowering induction in octoploid strawberry, we isolated and analyzed three important flowering related genes (FaLFY, FaAP1 and FaTFL1) from cultivated strawberry 'Huaji'. Partial sequences of these three genes were isolated with PCR and their expression roles in different tissues and organs of strawberry were detected. The deduced amino acids of FaLFY showed the highest degree of similarity to its homologue in Rosa lucieae (93%), FaAP1 showed the highest degree of similarity to its homologue in Eriobotrya japonica (88%) and FaTFL1 showed the highest degree of similarity to its homologue in Fragaria vesca (99%). FaLFY expression mainly detected in the reproductive tissues, FaAP1 expression mainly detected in the reproductive tissues and partially in floral organs, and FaTLF1 expression mainly detected both in the reproductive tissues and young vegetative tissues. These results suggested that all these three genes may be involved in the flowering process of strawberry.
The FLORICAULA/LEAFY (FLO/LFY) homologues' genes are necessary for normal flower development and play a key role in diverse angiosperm species. In this paper, an orthologue of FLORICAULA/LEAFY, CmLFY (chestnut FLO/LFY), was isolated from the inflorescence of chestnut trees. Its expression was detected in various tissues. Furthermore, the flowering effectiveness of the gene was assessed with transgenic Arabidopsis. CmLFY protein showed a high degree of similarity to PEAFLO (78%), which is a homologue of FLO/LFY from pea. RT-PCR analysis showed that, CmLFY expressed at high levels in inflorescences, but not in young leaves, fruits or stems. The transgenic Arabidopsis with over-expressed CmLFY showed accelerated flowering, which supports that CmLFY encodes a functional orthologue of the FLORICAULA/LEAFY genes of angiosperms despite its sequence divergence. These results suggest that CmLFY is involved in inflorescence development in chestnut.
Trochodendron aralioides is the sole member of the family Trochodendraceae, which belongs to the basal eudicots, has vesselless wood, and lacks a distinct perianth. Our observations confirmed that there are numerous perianth-like structures and that the number of these structures differs between protandrous and protogynous flowers and among the positions within an inflorescence. The epidermal cells on many floral parts of T. aralioides are papillate or conical, similar to the ones of ordinary showy petals of other species. The data in this article support the hypothesis that the perianth of Trochodendron has been secondarily lost and suggest that some aspects of petal identity, e.g., papillate cells, have been retained that might be important for pollinator attraction. We have identified 11 homologues of floral organ identity genes—two A-class, three B-class, two C-class, and four E-class homologous genes—from T. aralioides. Phylogenetic analysis shows that all of the genes arose before a major duplication of MADS-box genes at the base of the core eudicots. Expression patterns for those floral organ identity gene homologues was determined by reverse transcriptase PCR, which showed variations that do not conform well to the current floral ABCDE model. In addition, all paralogous genes have distinct expression patterns, suggesting that they had undergone functional divergence.
For studies of the evolution of development in angiosperms, early-diverging eudicot taxa are of particular interest for comparisons with established core eudicot model plants, such as Arabidopsis. Here we provide a detailed description of shoot and floral development of the basal eudicot California poppy (Eschscholzia californica). Rosette formation in the vegetative phase is accompanied by increased leaf complexity and shoot apex size. The flowering phase is characterized by internode elongation, formation of dissected cauline leaves, terminal flowers, and basipetal inflorescence branching. For developing flowers and fruits, we have defined 14 stages according to important landmark events, from inflorescence primordium initiation through seed dispersal. Floral organ initiation, morphogenesis, increase in floral meristem size, and the surface structure of mature floral organs are recorded in detail. The duration of the later floral stages, as well as the path of pollen tube growth in the gynoecium, is documented. Comparison of California poppy floral development with that of Arabidopsis indicates considerable differences in terms of organ fusion, whorl proliferation, and variability of size and organ number between the two species. Transitions in meristem identity from germination to floral organogenesis were monitored using expression of the developmental control gene EcFLO, the Eschscholzia ortholog of FLORICAULA/LEAFY. We found that the pattern of expression of EcFLO in the flanks of the shoot apex is maintained from late embryogenesis to flower initiation, indicating a continuous role for this gene in meristem function. As flower organs develop, EcFLO expression becomes more restricted to petal and stamen primordia. Development of the gynoecium occurs without EcFLO expression, indicating that EcFLO may not be necessary for the activation of C-class genes.
A LEAFY/FLORICAULA (LFY/FLO) homolog PpLFL (P
runus
p
ersica
L
EAFY/
F
LORICAULA
L
ike) gene was isolated from axillary buds of peach (Prunus persica (L.) Batsch. cv. Bayuecui) during flower induction period. The open reading frame of PpLFL spanned 1,248 bp, encoding a putative protein of 415 amino acid residues, which was with high similarity (50.48 %–84.69 %) to other FLO/LFY inferred proteins from different species. The spatial expression patterns of PpLFL were detected in axillary buds during the periods of flower induction by using immunohistolocalisation. The results showed that PpLFL gene was mainly expressed during flower induction time, and also in leaf and petal promordia at the SAM. For further functional analysis, the PpLFL was constitutively expressed in the Arabidopsis lfy mutant background, and the results showed that overexpression of PpLFL under the control of CaMV 35S promoter can accelerate flowering and give rise to normal flower organs. Our results suggest that PpLFL might play an important role in flower induction, and could act as a functional flower meristem identity gene in peach.
Hickory (Carya cathayensis Sarg.) is an important economic nut tree in China. However, its long juvenile phase is a deterrent to expanding its cultivation and production to wider areas. To understand the genetic and molecular mechanisms that underlie the reproductive process in hickory trees, CcLFY, a homologue of FLORICAULA/LEAFY, was cloned and its expression patterns were studied during vegetative and reproductive development. The cDNA sequence of CcLFY consisted of 1,442 bp encoding a putative protein of 385 amino acids. The protein consisted of a Pro-rich region, a central acidic domain, a putative Leucine zipper, and a basic region formed by a core of Arg and Lys residues that are critical motifs for transcription factors. sThe amino acid sequence of the CcLFY protein shares 89.2% and 65.6% identities with LFY proteins of Castanea mollissima and Arabidopsis thaliana, respectively. CcLFY was highly expressed in flower buds and leaves, weakly expressed in stems, and was undetectable in roots. In situ hybridization revealed that CcLFY was highly expressed in both immature leaves and flower buds. Expression of CcLFY was initiated during floral transition in the spring, and subsequently, continued to increase, reaching peak levels at 16 days after full bloom (DAFB). Moreover, during this period, the major floral buds formed. Heterologous expression of CcLFY in transgenic tobacco plants induced precocious flowering of growing shoots, and flowers were of normal phenotypes. These results suggested that CcLFY might play a pivotal role in flower initiation and in the development of floral organs in C. cathayensis.
Plant MADS-box genes of the AG/PLE subfamily control the identity of sexual organs. An AG-homologous gene (PrAG1) was characterized in the Pinus radiata genome. PrAG1 transcript was detected only in the female and male strobili. To investigate its regulatory function during floral development, three binary vectors were constructed and transformed into tobacco plants for overexpression of the PrAG1 full-length protein (35S::MIKC), and truncated proteins with deletion of C domain (35S::MIK) or deletion of both C and K domains (35S::MI) under control of the CaMV 35S promoter. All transgenic tobacco lines with ectopic expression of 35S::MIKC and 35S::MIK showed no phenotypic effect on floral development. However, 23.8% of the 35S::MI transgenic lines displayed altered inflorescence architecture and variety of floral development changes, including complete male sterility, suggesting that PrAG1 may be the P. radiata
AG-homologous gene with C-function and that it may play a role in the determination of meristem identities in both inflorescence and flowering. Expression of truncated AG MI genes could be useful in reducing plant pollen and seed formation, as well as increasing inflorescence branching and flower production, providing a novel engineered sterility strategy for transgenic plants with potential commercial application in molecular breeding of horticultural flowering plants.
Flowers are reproductive shoots produced by determinate floral meristems. The role of FLORICAULA/LEAFY-like genes in the specification of flowers varies between lineages and has not been characterized in a basal eudicot species. Here we report the phenotypic effects of virus-induced silencing of EcFLO, a FLORICAULA/LEAFY homologue in the California poppy Eschscholzia californica. EcFLO silencing resulted in repeated sepal and petal whorl formation and internodes between calyxes. A subset of silenced flowers showed reduced petal and stamen numbers and mosaic identities of floral organs. We also provide expression data of other floral regulator genes and conclude that EcFLO contributes to certain aspects of flower development, reflecting both functional conservation and differentiation among basal eudicots, core eudicots, and monocot grasses. EcFLO is involved in stamen and petal initiation, floral organ identity delimitation, suppression of internodes, and limitation of the number of perianth whorls. Further, silencing of EcFLO allows insights into perigynous flower development in Eschscholzia. We demonstrate that receptacle cup formation, by which the perianth and the androecium are elevated above the gynoecium late in flower development, responds to the initiation of the synsepalous calyx.
The first step in flower development is the generation of a floral meristem by the inflorescence meristem. We have analyzed how this process is affected by mutant alleles of the Arabidopsis gene LEAFY. We show that LEAFY interacts with another floral control gene, APETALA1, to promote the transition from inflorescence to floral meristem. We have cloned the LEAFY gene, and, consistent with the mutant phenotype, we find that LEAFY RNA is expressed strongly in young flower primordia. LEAFY expression procedes expression of the homeotic genes AGAMOUS and APETALA3, which specify organ identify within the flower. Furthermore, we demonstrate that LEAFY is the Arabidopsis homolog of the FLORICAULA gene, which controls floral meristem identity in the distantly related species Antirrhinum majus.
The analysis of mutations affecting flower structure has led to the identification of some of the genes that direct flower development. Cloning of these genes has allowed the formulation of molecular models of how floral meristem and organ identity may be specified, and has shown that the distantly related flowering plants Arabidopsis thaliana and Antirrhinum majus use homologous mechanisms in floral pattern formation.
Deficiens (defA+) is a homeotic gene involved in the genetic control of Antirrhinum majus flower development. Mutation of this gene (defA-1) causes homeotic transformation of petals into sepals and of stamina into carpels in flowers displaying the 'globifera' phenotype, as shown by cross sections and scanning electronmicroscopy of developing flowers. A cDNA derived from the wild type defA+ gene has been cloned by differential screening of a subtracted 'flower specific' cDNA library. The identity of this cDNA with the defA+ gene product has been confirmed by utilizing the somatic and germinal instability of defA-1 mutants. According to Northern blot analyses the defA+ gene is expressed in flowers but not in leaves, and its expression is nearly constant during all stages of flower development. The 1.1 kb long mRNA has a 681 bp long open reading frame that can code for a putative protein of 227 amino acids (mol. wt 26.2 kd). At its N-terminus the DEF A protein reveals homology to a conserved domain of the regulatory proteins SRF (activating c-fos) in mammals and GRM/PRTF (regulating mating type) in yeast. We discuss the structure and the possible function of the DEF A protein in the control of floral organogenesis.
Mutations in the homeotic gene agamous of the plant Arabidopsis cause the transformation of the floral sex organs. Cloning and sequence analysis of agamous suggest that it encodes a protein with a high degree of sequence similarity to the DNA-binding region of transcription factors from yeast and humans and to the product of a homeotic gene from Antirrhinum. The agamous gene therefore probably encodes a transcription factor that regulates genes determining stamen and carpel development in wild-type flowers.
We have characterized the floral phenotypes produced by the recessive homeotic apetala 1-1 (ap1-1) mutation in Arabidopsis. Plants homozygous for this mutation display a homeotic conversion of sepsis into brachts and the concomitant formation of floral buds in the axil of each transformed sepal. In addition, these flowers lack petals. We show that the loss of petal phenotype is due to the failure of petal primordia to be initiated. We have also constructed double mutant combinations with ap1 and other mutations affecting floral development. Based on these results, we suggest that the AP1 and the apetala 2 (AP2) genes may encode similar functions that are required to define the pattern of where floral organs arise, as well as for determinate development of the floral meristem. We propose that the AP1 and AP2 gene products act in concert with the product of the agamous (AG) locus to establish a determinate floral meristem, whereas other homeotic gene products are required for cells to differentiate correctly according to their position. These results extend the proposed role of the homeotic genes in floral development and suggest new models for the establishment of floral pattern.
The shoot apex of higher plants contains undifferentiated meristematic cells that serve as the origin of post-embryonic organs. The transition from vegetative to reproductive growth results in the commitment of the apical meristem to floral organ formation. To identify the molecular signals that initiate floral development, we have pursued the isolation of genes that are transcriptionally active in the shoot apex of tobacco during the transition from vegetative to floral growth. The small size of the apex led us to utilize polymerase chain reaction shoot apices. This approach enabled the isolation of the apex-specific and floral apex-specific cDNA clones described in this paper. One clone, A3, detected an equivalent level of transcript in the shoot apex during all developmental stages observed. The second clone, FA2, detected a unique transcript that increased in abundance in the shoot apex during the transition to flowering and showed high levels of expression in developing petals, stamens, and pistils.
We describe the effects of four recessive homeotic mutations that specifically disrupt the development of flowers in Arabidopsis thaliana. Each of the recessive mutations affects the outcome of organ development, but not the location of organ primordia. Homeotic transformations observed are as follows. In agamous-1, stamens to petals; in apetala2-1, sepals to leaves and petals to staminoid petals; in apetala3-1, petals to sepals and stamens to carpels; in pistillata-1, petals to sepals. In addition, two of these mutations (ap2-1 and pi-1) result in loss of organs, and ag-1 causes the cells that would ordinarily form the gynoecium to differentiate as a flower. Two of the mutations are temperature-sensitive. Temperature shift experiments indicate that the wild-type AP2 gene product acts at the time of primordium initiation; the AP3 product is active later. It seems that the wild-type alleles of these four genes allow cells to determine their place in the developing flower and thus to differentiate appropriately. We propose that these genes may be involved in setting up or responding to concentric, overlapping fields within the flower primordium.
Mutations in the AGAMOUS (AG) gene of Arabidopsis thaliana result in the conversion of reproductive organs, stamens and carpels, into perianth organs, sepals and petals. We have isolated and characterized the putative AG gene from Nicotiana tabacum, NAG1, whose deduced protein product shares 73% identical amino acid residues with the Arabidopsis AG gene product. RNA tissue in situ hybridizations show that NAG1 RNA accumulates early in tobacco flower development in the region of the floral meristem that will later give rise to stamens and carpels. Ectopic expression of NAG1 in transgenic tobacco plants results in a conversion of sepals and petals into carpels and stamens, respectively, indicating that NAG1 is sufficient to convert perianth into reproductive floral organs.
There has been an explosion of information about flower development recently, largely because of genetic and molecular studies in Arabidopsis thaliana and Antirrhihum majus. A number of homeotic genes have been identified that regulate flower development, and models have been proposed for the specification of meristem and floral organ identities. Molecular cloning of many of these genes has allowed the testing of specific predictions of the models but also has led to modifications of a floral organ identity model. Furthermore, several of the floral genes contain a conserved region, the MADS box, which encodes a domain with striking sequence similarity with known transcription factors from human and yeast. Additional MADS box genes have been isolated from several plants; these genes are likely to play important regulatory roles during flower development. The genetic and molecular studies have uncovered many of the components of a complex network of regulatory proteins that directs flower development. Further characterization of these and other yet to be discovered components promises to contribute a great deal to our understanding of the mechanisms controlling flower development. Flowering plants, like other land plants, have vegetative organs such as roots, stems, and leaves, which absorb nutrients and water from the soil, transport them to other parts of the plant, and synthesize organic compounds using the sun's energy. In addition, flowering plants produce elaborate reproductive structures, the flowers, which, following fertilization, become fruits and bear seeds. From the seasoned gardener to the casual observer, from the naturalist to the florist shopper, people have always been fascinated by the enormous variety of flowers, ranging from 2 mm to > 10 cm in length, covering the whole visual spectrum with their colors, and differing in the arrangement of flowers and the symmetry within a flower. How do flowers develop? What genes regulate this complex process? In recent years, rapid advances have been made in addressing these questions, largely as a result of genetic and molecular studies in two distantly related flowering plants, Arabidopsis thaliana, a relative of cauliflower and cabbage, and the snapdragon, Antirrhinum majus.
Plant development is governed by intrinsic and environmen- tal factors that regulate the identity and activity of meristems, organized tissues of pluripotent "stem" cells, that together de- termine plant form and architecture. However, little is known about how these factors act at the molecular leve1 to affect meristem identity and function. Genetic studies in Arabidop- sis and other plant species such as snapdragon, petunia, and maize have revealed a hierarchy of regulatory genes that func- tion together to promote the formation of the floral meristem and to regulate floral organogenesis. Mutations in these genes result in dramatic defects in flower development that can af- fect both meristem identity and organ development. Table 1 lists the Arabidopsis genes that are known to control meristem and organ identity and their snapdragon equivalents. One class of regulatory genes, the homeotic organ identity genes, have provided important insights into the genetic and molecular mechanisms that govern floral organ identity and development (Bowman et al., 1991; Coen and Meyerowitz, 1991; Coen and Carpenter, 1993, this issue; van der Krol and Chua, 1993, this issue). In Arabidopsis, these genes include
Members of the lin-12/Notch gene family encode receptors for intercellular signals and are found throughout the animal kingdom. In many animals, the presence of at least two lin-12/Notch genes raises the issue of the significance of this duplication and divergence. In Caenorhabditis elegans, two lin-12/Notch genes, lin-12 and glp-1, encode proteins that are 50% identical, with different numbers of epidermal growth factor-like motifs in their extracellular domains. Many of the cell fate decisions mediated by lin-12 and glp-1 are distinct. Here, we express glp-1 protein under the control of lin-12 regulatory sequences in animals lacking endogenous lin-12 activity and find that glp-1 can substitute for lin-12 in mediating cell fate decisions. These results imply that the lin-12 and glp-1 proteins are biochemically interchangeable, sharing common ligand and effector proteins, and that the discrete lin-12 and glp-1 mutant phenotypes result from differential gene expression. In addition, these results suggest that the duplicate lin-12/Notch genes found in vertebrates may also be biochemically interchangeable.
Variation in plant shoot structure may be described as occurring through changes within a basic unit, the metamer. Using this terminology, the apical meristem of Arabidopsis produces three metameric types sequentially: type 1, rosette; type 2, coflorescence-bearing with bract; and type 3, flower-bearing without bract. We describe a mutant of Arabidopsis, Leafy, homozygous for a recessive allele of a nuclear gene LEAFY (LFY), that has an inflorescence composed only of type 2-like metamers. These data suggest that the LFY gene is required for the development of type 3 metamers and that the transition from type 2 to type 3 metamers is a developmental step distinct from that between vegetative and reproductive growth (type 1 to type 2 metamers). Results from double mutant analysis, showing that lfy-1 is epistatic to the floral organ homeotic gene ap2-6, are consistent with the hypothesis that a functional LFY gene is necessary for the expression of downstream genes controlling floral organ identity.
We studied the temporal and spatial regulation of three mRNA sequence sets that are present exclusively, or at elevated levels, in the tobacco anther. One mRNA set accumulates in the tapetum and decays as the tapetum degenerates later in anther development. The second mRNA set accumulates after the tapetal-specific mRNAs, is localized within the stomium and connective, and also decays as these cell types degenerate during anther maturation. The third mRNA sequence set persists throughout anther development and is localized within most anther tissues. A tapetal-specific gene, designated as TA29, was isolated from a tobacco genome library. Runoff transcription studies and experiments with chimeric [beta]-glucuronidase and diphtheria toxin A-chain genes showed that the TA29 gene is regulated primarily at the transcriptional level and that a 122-base pair 5[prime] region can program the tapetal-specific expression pattern. Destruction of the tapetum by the cytotoxic gene had no effect on the differentiation and/or function of surrounding sporophytic tissues but led to the production of male-sterile plants. Together, our studies show that several independent gene expression programs occur during anther development and that these programs correlate with the differentiated state of specific anther cell types.
Changes in gene expression during flower formation were studied in the long-day plant Sinapis alba. The day length dependence was exploited to synchronize flower formation in a large population of mustard plants. After an inductive light treatment, apices were harvested after different lengths of time, and changes in gene expression were analyzed. Two major groups of genes were identified whose expression was affected during flower formation. Transcripts of the first group (group I) were present at low concentration in the apex of noninduced plants. They began to accumulate strongly after the end of the inductive light period. They reached a maximum 2 days to 10 days after flower induction and then declined slowly. Transcripts of the second group of genes (group II) could be detected for the first time 10 days after flower induction. Within a very short time, these transcripts accumulated dramatically and reached a maximum 15 days after flower induction before beginning to decline. They dropped beyond the limit of detection before the flower reached maturity.
The cellular parameters of leaf development in tobacco (Nicotiana tabacum L.) have been characterized using clonal analysis, an approach that provides unequivocal evidence of cell lineage. Our results indicate that the tobacco leaf arises from a group of around 100 cells in the shoot apical meristem. Each of these cells contributes to a unique longitudinal section of the axis and transverse section of the lamina. This pattern of cell lincage indicates that primordial cells contribute more or less equally to the growth of the axis, in contrast to the more traditional view of leaf development in which the leaf is pictured as arising from a group of apical initials. Clones induced prior to the initiation of the lamina demonstrate that the subepidermal layer of the lamina arises from at least six files of cells. Submarginal cells usually divide with their spindles parallel to the margin, and therefore contribute relatively little to the transverse expansion of the lamina. During the expansion of the lamina the orientation and frequency of cell division are highly regulated, as is the duration of meristematic growth. Initially, cell division is polarized so as to produce lineages that are at an oblique angle to the midrib; later cell division is in alternating perpendicular planes. The distribution of clones generated by irradiation at various stages of development indicates that cell division ceases at the tip of the leaf when the leaf is about one tenth its final size, and then ceases in progressively more basal regions of the lamina. Variation in the mutation frequency within the lamina reflects variation in the frequency of mitosis. Prior to the mergence of the leaf the frequency of mutation is maximal near the tip of the leaf and extremely low at its base; after emergence, the frequency of mutation increases at the base of the leaf. In any given region of the lamina the frequency of mutation is highest in interveinal regions, and is relatively low near the margin. Thus, both the orientation and frequency of cell division at the leaf margin indicate that this region plays a minor role in the growth of the lamina.
Floral determination in the terminal bud of the short-day plant Nicotiana tabacum cv. Maryland Mammoth has been investigated. Plants grown continuously in short days flowered after producing 31.4±1.6 (SD) nodes while plants grown continuously in long days did not flower and produced 172.5±9.5 nodes after one year. At various ages, expressed as number of leaves that were at least 1.0 cm in length above the most basal 10-cm leaf, one of three treatments was performed on plants grown from seed in short days: 1) whole plants were shifted from short days to long days, 2) the terminal bud was removed and then rooted and grown in long days, and 3) the terminal bud was removed and then rooted and grown in short days. Whole plants flowered only when shifted from short days to long days at age 15 or later. Only rooted terminal buds from plants at age 15 or older produced plants that flowered when grown in long days. Only terminal buds from plants at age 15 or older that were rooted and grown in short days produced the same number of nodes as they would have produced in their original locations while buds from younger plants produced more nodes than they would have in their original locations. Thus, determination for floral development in the terminal bud, as assayed by rooting, is simultaneous with the commitment to flowering as assayed by shifting whole plants to non-inductive conditions.
There is a finite upper limit to the number of gene loci which an organism can afford to have. An organism having 3 × 106 or so gene loci would exterminate itself from an unbearable mutation load. Yet, the mammalian genome is large enough to accommodate that many gene loci. Natural selection is essentially conservative. A new gene with a hitherto non-existent function can not be created unless a temporary escape from the relentless pressure of natural selection is provided by gene duplication. However, the chance of a redundant copy of an old gene emerging as a new gene is considerably smaller than of that copy becoming degenerate. Perhaps for this reason, evolution from simpler life forms to mammals appears to have been accompanied not only by successive additions of new gene loci, but also by accumulation in the genome of a great deal of degenerate "nonsense" DNA base sequences. At least 90% of the mammalian genomic DNA appears to represent "nonsense" DNA base sequence of various kinds. The creation of additional regulatory systems contributed more to big leaps in evolution than did the creation of new structural genes. Yet, in order not to be burdened with an unbearable mutation load, the necessary increase in the number of regulatory systems had to be compensated by simplification of each regulatory system. It would not be surprising if each mammalian regulatory system is shown to have fewer components than the lac-operon system of Escherichia coli.
The terminal and axillary buds of the day-neutral plant, Nicotiana tabacum cv. Wisconsin 38, become determined for floral development during the growth of the plant. This state of determination can be demonstrated with a simple experiment: buds determined for floral development produce the same number of nodes in situ and if rooted. After several months of growth and the production of many leaves, the terminal bud became determined for floral development within a period of about 2 days. After the terminal bud became florally determined, it produced four nodes and a terminal flower. The buds located in the axils of leaves borne just below the floral branches became florally determined 5 to 9 days after the terminal bud became florally determined. Since florally-determined axillary buds were not clonally derived from a florally-determined terminal meristem, axillary buds and the terminal bud acquired the state of floral determination independently. These data indicate that a pervasive signal induced a state of floral determination in competent terminal and axillary buds.
We compared the single-copy DNA sequences of the tetraploid tobacco plant, Nicotiana tabacum, with those of its diploid progenitors N. sylvestris and N. tomentosiformis. We observed that 65% of N. sylvestris and N. tomentosiformis single-copy DNA fragments reacted with each other using moderately stringent hybridization conditions (60 C, 0.18 M Na+). An additional 10% sequence homology was detected when the hybridization temperature was reduced by 10 C. The thermal stability of interspecific single-copy DNA duplexes indicated that they were approximately 6% more mispaired than homologous single-copy DNA duplexes. In contrast, we observed almost no single-copy DNA divergence between N. tabacum and its diploid progenitors. Greater than 99% of N. sylvestris and N. tomentosiformis single-copy DNAs reacted with N. tabacum DNA using moderately stringent hybridization conditions. The thermal stability of these duplexes indicated that they contained no more sequence mismatch than homologous single-copy duplexes. Together, our results show that significant single-copy DNA sequence divergence has occurred between the diploid N. sylvestris and N. tomentosiformis genomes. However, by applying our experimental criteria these single-copy DNAs are indistinguishable from their counterparts in the hybrid N. tabacum nucleus.
A new method for determining nucleotide sequences in DNA is described. It is similar to the "plus and minus" method [Sanger, F. & Coulson, A. R. (1975) J. Mol. Biol. 94, 441-448] but makes use of the 2',3'-dideoxy and arabinonucleoside analogues of the normal deoxynucleoside triphosphates, which act as specific chain-terminating inhibitors of DNA polymerase. The technique has been applied to the DNA of bacteriophage varphiX174 and is more rapid and more accurate than either the plus or the minus method.
The comparison of human and chimpanzee macromolecules leads to several inferences: 1) Amino acid sequencing, immunological, and electrophoretic methods of protein comparison yield concordant estimates of genetic resemblance. These approaches all indicate that the average human polypeptide is more than 99 percent identical to its chimpanzee counterpart. 2) Nonrepeated DNA sequences differ more than amino acid sequences. A large proportion of the nucleotide differences between the two species may be ascribed to redundancies in the genetic code or to differences in non-transcribed regions. 3) The genetic distance between humans and chimpanzees, based on electrophoretic comparison of proteins encoded by 44 loci is very small, corresponding to the genetic distance between sibling species of fruit flies or mammals. Results obtained with other biochemical methods are consistent with this conclusion. However, the substantial anatomical and behavioral differences between humans and chimpanzees have led to their classification in separate families. This indicates that macromolecules and anatomical or behavioral features of organisms can evolve at independent rates. 4) A relatively small number of genetic changes in systems controlling the expression of genes may account for the major organismal differences between humans and chimpanzees. Some of these changes may result from the rearrangement of genes on chromosomes rather than from point mutations (53).
Mutations in the APETALA3 (AP3) gene of A. thaliana result in homeotic transformations of petals to sepals and stamens to carpels. We have cloned the AP3 gene from Arabidopsis based on its homology to the homeotic flower gene deficiens (DEFA) from the distantly related plant Antirrhinum majus. The sequence of four ap3 mutant alleles and genetic mapping analysis prove that the DEFA homolog is AP3. Like several other plant homeotic genes, the AP3 gene contains a MADS box and likely acts as a transcription factor. The region-specific spatial expression pattern of AP3 rules out certain types of sequential models of flower development and argues in favor of a spatial model based on positional information. Since DEFA and AP3 have very similar protein products, mutant phenotypes, and spatial expression patterns, it is likely that these genes are cognate homologs.
The first step in flower development is the transition of an inflorescence meristem into a floral meristem. Each floral meristem differentiates into a flower consisting of four organ types that occupy precisely defined positions within four concentric whorls. Genetic studies in Arabidopsis thaliana and Antirrhinum majus have identified early-acting genes that determine the identify of the floral meristem, and late-acting genes that determine floral organ identity. In Arabidopsis, at least two genes, APETALA1 and LEAFY, are required for the transition of an influorescence meristem into a floral meristem. We have cloned the APETALA1 gene and here we show that it encodes a putative transcription factor that contains a MADS-domain. APETALA1 RNA is uniformly expressed in young flower primordia, and later becomes localized to sepals and petals. Our results suggest that APETALA1 acts locally to specify the identity of the floral meristem, and to determine sepal and petal development.
Anomalous flowering of the Antirrhinum majus mutant squamosa (squa) is characterized by excessive formation of bracts and the production of relatively few and often malformed or incomplete flowers. To study the function of squamosa in the commitment of an inflorescence lateral meristem to floral development, the gene was cloned and its genomic structure, a well as that of four mutant alleles, was determined. SQUA is a member of a family of transcription factors which contain the MADS-box, a conserved DNA binding domain. In addition, we analysed the temporal and spatial expression pattern of the squa gene. Low transcriptional activity of squa is detectable in bracts and in the leaves immediately below the inflorescence. High squa transcript levels are seen in the inflorescence lateral meristems as soon as they are formed in the axils of bracts. Squa transcriptional activity persists through later stages of floral morphogenesis, with the exception of stamen differentiation. Although necessary for shaping a normal racemose inflorescence, the squa function is not absolutely essential for flower development. We discuss the function of the gene during flowering, its likely functional redundancy and its possible interaction with other genes participating in the genetic control of flower formation in Antirrhinum.
We characterized the distribution of AGAMOUS (AG) RNA during early flower development in Arabidopsis. Mutations in this homeotic gene cause the transformation of stamens to petals in floral whorl 3 and of carpels to another ag flower in floral whorl 4. We found that AG RNA is present in the stamen and carpel primordia but is undetectable in sepal and petal primordia throughout early wild-type flower development, consistent with the mutant phenotype. We also analyzed the distribution of AG RNA in apetela2 (ap2) mutant flowers. AP2 is a floral homeotic gene that is necessary for the normal development of sepals and petals in floral whorls 1 and 2. In ap2 mutant flowers, AG RNA is present in the organ primordia of all floral whorls. These observations show that the expression patterns of the Arabidopsis floral homeotic genes are in part established by regulatory interactions between these genes.
Plants carrying the floricaula (flo) mutation cannot make the transition from inflorescence to floral meristems and have indeterminate shoots in place of flowers. The flo-613 allele carries a Tam3 transposon insertion, which allowed the isolation of the flo locus. The flo gene encodes a putative protein (FLO) containing a proline-rich N-terminus and a highly acidic region. In situ hybridization shows that the flo gene is transiently expressed in the very early stages of flower development. The earliest expression seen is in bract primordia, followed by sepal, petal, and carpel primordia, but no expression is detected in stamen primordia. This pattern of expression has implications for how flo affects phyllotaxis, organ identity, and determinacy. We propose that flo interacts in a sequential manner with other homeotic genes affecting floral organ identity.
The above-ground portion of a plant develops from the shoot apical meristem. An abundant source of apical meristems was obtained from cauliflower heads. Meristematic cDNAs were identified by differential screening and used to isolate corresponding Arabidopsis thaliana genes. Transcriptional promoters from Arabidopsis clones were fused to the beta-glucuronidase (GUS) reporter gene and introduced into plants, and GUS expression was used to analyze temporal and spatial regulation of the promoters. One promoter (meri-5) directed GUS expression in the meristematic dome and not the surrounding leaf primordia. The meri-5 promoter also directed GUS expression at branching points in the shoot and root. A second meristematic gene was found to be a histone (H3) gene. The H3 promoter was isolated and fused to GUS. Expression of the H3-GUS fusion in transgenic tobacco showed preferential expression in the peripheral zone and a lack of noticeable staining in the central zone.
To isolate and study genes controlling floral development, we have carried out a large-scale transposon-mutagenesis experiment in Antirrhinum majus. Ten independent floral homeotic mutations were obtained that could be divided into three classes, depending on whether they affect (1) the identity of organs within the same whorl; (2) the identity and sometimes also the number of whorls; and (3) the fate of the axillary meristem that normally gives rise to the flower. The classes of floral phenotypes suggest a model for the genetic control of primordium fate in which class 2 genes are proposed to act in overlapping pairs of adjacent whorls so that their combinations at different positions along the radius of the flower can specify the fate and number of whorls. These could interact with class 1 genes, which vary in their action along the vertical axis of the flower to generate bilateral symmetry. Both of these classes may be ultimately regulated by class 3 genes required for flower initiation. The similarity between some of the homeotic phenotypes with those of other species suggests that the mechanisms controlling whorl identity and number have been highly conserved in plant evolution. Many of the mutations obtained show somatic and germinal instability characteristic of transposon insertions, allowing the cell-autonomy of floral homeotic genes to be tested for the first time. In addition, we show that the deficiens (def) gene (class 2) acts throughout organ development, but its action may be different at various developmental stages, accounting for the intermediate phenotypes conferred by certain def alleles. Expression of def early in development is not necessary for its later expression, indicating that other genes act throughout the development of specific organs to maintain def expression. Direct evidence that the mutations obtained were caused by transposons came from molecular analysis of leaf or flower pigmentation mutants, indicating that isolation of the homeotic genes should now be possible.
The beta subunit of the mitochondrial ATP synthase in Nicotiana plumbaginifolia is encoded by two nuclear genes, atp2-1 and atp2-2, which are both expressed. The complete nucleotide sequence of atp2-1 has been determined. It contains eight introns ranging from 88 to 1453 bp. The last intron contains a putative insertion element (Inp), of 812 bp bordered by 35-bp inverted repeats which share an 11-bp homology with Agrobacterium tumefaciens T-DNA borders. Sequences homologous to Inp are present in multiple copies in the N. plumbaginifolia and the N. tabacum genome but not in more distant species. The atp2-1 encoded polypeptide is highly homologous to beta subunits from other ATP synthases but it contains an extension at the N terminus which is probably involved in mitochondrial targeting. A sequence homology between exon 4 of atp2-1 and exon 1 of the human ras genes suggests a common ancestral origin for these exons.
To assess the significance of macromolecular sequence differences among species, we compared the serum albumins of 81 pairs of vertebrate species capable of producing viable hybrids. Micro-complement fixation experiments showed that the average difference between the albumins within such pairs was only 3 immunological distance units for placental mammals (31 pairs), but 36 units for frogs (50 pairs). Albumin immunological distance is strongly correlated with other measures of genetic distance, including those made with DNA annealing techniques. It therefore seems likely that mammalian species pairs capable of hybridization are far more similar at the macromolecular sequence level than is the case for most hybridizable frogs.
We think the most likely explanation for the marked molecular restriction on hybridization among mammals is that the ratio of regulatory evolution to protein evolution is higher for mammals than for frogs. Mammals may have experienced unusually rapid regulatory evolution; indeed, this could be the factor responsible for their unusually rapid anatomical evolution.
We have compared the relative rates of protein evolution and chromosomal evolution in frogs and mammals. The average rate of change in chromosome number has been about 20 times faster in mammals than in frogs. Whereas it takes only 3.5 million years, on the average, for a pair of mammal species to develop a difference in chromosome number, the corresponding period for frogs is 70 million years. In contrast, the rate of protein evolution in mammals has been roughly equal to that in frogs. The rapid rate of gene rearrangement in mammals parallels both their rapid anatomical evolution and their rapid evolutionary loss of the potential for interspecific hybridization. Thus, gene rearrangements may be more important than point mutations as sources for evolutionary changes in anatomy and way of life.
The University of Wisconsin Genetics Computer Group (UWGCG) has been organized to develop computational tools for the analysis
and publication of biological sequence data. A group of programs that will interact with each research-article has been developed
for the Digital Equipment Corporation VAX computer using the VMS operating system. The programs available and the conditions
for transfer are described.
It is generally accepted that the specific function of a gene depends on its coding sequence. The three paired-box and homeobox genes paired (prd), gooseberry (gsb) and gooseberry neuro (gsbn) have distinct developmental functions in Drosophila embryogenesis. During the syncytial blastoderm stage, the pair-rule gene prd activates segment-polarity genes, such as gsb, wingless (wg), and engrailed (en), in segmentally repeated stripes. After germ-band extension, gsb maintains the expression of wg, which in turn specifies the denticle pattern by repressing a default state of ubiquitous denticle formation in the ventral epidermis. In addition, gsb activates gsbn, which is expressed mainly in the central nervous system, suggesting that gsbn is involved in neural development. Here we show that, despite the functional difference and the considerably diverged coding sequence of these genes, their proteins have conserved the same function. The finding that the essential difference between genes may reside in their cis-regulatory regions exemplifies an important evolutionary mechanism of how function diversifies after gene duplication.
Rapid advances have been made in the understanding of the genetic basis of development and pattern formation in a variety
of model systems. By examining the extent to which these developmental systems are conserved or altered between different
organisms, insight can be gained into the evolutionary events that have generated the diversity of organisms around us. The
molecular and genetic basis of early pattern formation in Drosophila melanogaster has been particularly well studied, and
comparisons to other insects have revealed conservation of some aspects of development, as well as differences that may explain
variations in early patterning events.
Recessive mutations at the plena (ple) locus result in a homeotic conversion of sex organs to sterile perianth organs in flowers of Antirrhinum majus. A complementary phenotype, in which sex organs replace sterile organs, is conferred by semidominant ovulata mutations. The ple locus was identified and isolated using a homologous gene, agamous from Arabidopsis, as a probe. The expression of ple is normally restricted to the inner two whorls of the flower, where sex organs develop. However, in ovulata mutants, ple is expressed ectopically in the outer two whorls of the flower and in vegetative organs. These mutants correspond to gain-of-function alleles of ple, suggesting that ple is sufficient for promoting sex organ development within the context of the flower. The plena and ovulata phenotypes result from opposite orientations of the transposon Tam3 inserted in the large intron of ple.
In this paper, we describe the synthesis of a cDNA library from the vegetative shoot apical meristem and the analysis of clones selected from it. Using in situ hybridization, we characterized the patterns of expression of these genes in the tomato shoot apical meristem, as well as the patterns obtained from other sources. The results from the analysis of 15 cDNAs indicated the following six main patterns of gene expression in the shoot apical meristem: overall expression, zero expression, expression limited to the epidermis, expression excluded from the epidermis, punctate expression, and expression elevated in the flanks of the meristem. The patterns observed and the nature and number of the genes showing these patterns necessitate a reinterpretation of the models of meristem structure and function. In particular, the data suggest a compartmentation within the shoot apical meristem based on the spatial expression of particular subsets of genes. This paper also reports on the specific and precise criteria essential for the correct identification of meristem-specific gene expression. The data give new insight into the molecular organization of the shoot apical meristem and provide the framework for a detailed dissection of the factors controlling this organization.
Homeotic mutants have been useful for the study of animal development. Such mutants are also known in plants. The isolation and molecular analysis of several homeotic genes in Antirrhinum majus provide insights into the underlying molecular regulatory mechanisms of flower development. A model is presented of how the characteristic sequential pattern of developing organs, comprising the flower, is established in the process of morphogenesis.
The identity of floral organs in Arabidopsis thaliana is determined by homeotic genes, which are expressed in specific regions of the developing flower. The initial activation
of homeotic genes is accomplished at least in part by the products of two earlier acting genes with overlapping functions.
These are the floral meristem—identity genes LEAFY and APETALA1. The requirements of LEAFY and APETALA1 activity vary for different homeotic genes.
The genus as a genetic resource In Nicotiana: Procedures for Experimental Use
- H H Smith
Smith, H.H. (1979). The genus as a genetic resource. In Nicotiana: Procedures for Experimental Use, R.B. Durbin, ed (U.S. Department of Agriculture Technical Bulletin 1586), pp. 1-16
Genetic interactions that regulate inflorescence development in Arabidopsis Floral determination in the terminal and axillary buds of Nicotiana tabacum L The genus as a genetic resource
- S Shannon
- D R S R Meeks-Wagner
- C N Mcdaniel
Shannon, S., and Meeks-Wagner, D.R. (1993). Genetic interactions
that regulate inflorescence development in Arabidopsis. Plant Cell
Singer, S.R., and McDaniel, C.N. (1986). Floral determination in the
terminal and axillary buds of Nicotiana tabacum L. Dev. Biol. 118,
Smith, H.H. (1979). The genus as a genetic resource. In Nicotiana:
Procedures for Experimental Use, R.B. Durbin, ed (U.S. Department
of Agriculture Technical Bulletin 1586), pp. 1-16.