Project description:Combinatorial activities of SHORT VEGETATIVE PHASE and FLOWERING LOCUS C define distinct modes of flowering regulation in Arabidopsis
Project description:The transition to flowering in plants is controlled by a regulatory network that responds to both developmental and environmental signals. The MADS-box genes FLOWERING LOCUS C (FLC) and SHORT VEGETATIVE PHASE (SVP) are major flowering repressors that enhance responses to environmental cues such as winter temperatures, high ambient temperatures and photoperiod. FLC and SVP physically interact in vivo and mutation of each gene causes early flowering while the double mutant is more extreme. The molecular mechanisms underlying these genetic interactions are mostly unknown. We addressed the regulatory input of these two key transcription factors (TFs) both individually and as a complex at the genome-wide level through ChIP-seq and microarray expression analysis in single and double mutants. Analysis of each TF demonstrated that the complex acts predominantly via functional redundancy in the repression of flowering. SVP and FLC bind to the same regions of the flowering genes SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) and FLOWERING LOCUS T (FT) but do not require the presence of the other to bind. However, genome-wide identification of SVP and FLC occupancy events revealed that their binding scenarios are quantitatively and qualitatively affected by the presence of the cognate partner. A subgroup of genes whose regulation by these TFs depends exclusively on combinatorial binding of both proteins was identified, demonstrating a qualitatively essential role of the SVP-FLC complex. Some of these genes are involved in the control of flowering through direct and indirect regulation of Gibberellin-related processes. Cis-regulatory elements enriched only at such complex-bound sites were identified. Thus the regulatory output mediated by SVP and FLC reveals substantial flexibility, leading to dependent and independent DNA binding that enables additive, cooperative and repressive modes of co-regulation. In total 48 samples; 24 leaf samples corresponding to two different growing conditions, 24 apex samples corresponding to two different growing conditions.
Project description:The MADS-box transcription factors FLOWERING LOCUS C (FLC) and SHORT VEGETATIVE PHASE (SVP) are major transcriptional repressors controlling flowering time. They enhance responses to environmental cues such as winter temperatures, high ambient temperatures and photoperiod, acting at least in part by blocking transcription of the floral pathway integrators. As other MADS-box transcription factors, FLC and SVP can interact in vivo forming multimeric complexes. Mutations in either FLC or SVP lead to similar early flowering, suggesting that FLC-SVP interaction might be the major control unit. Here we have analyzed the coordinated regulatory modes of these two key transcription factors at a genome-wide level through ChIP-seq and gene expression microarrays. Genome-wide identification of SVP and FLC DNA-binding occupancy events revealed that their binding scenarios are strongly yet differently affected by the presence of the cognate partner both at a quantitative as well as a qualitative level. Also, we identified a subgroup of genes whose regulation exclusively depends on the combinatorial binding of these two proteins, strengthening the role of the SVP-FLC complex. Some of these genes are involved in the control of flowering through direct and indirect regulation of Gibberellin-related genes such as GA2ox8, DDF1 and TEM1. Interestingly, we identified cis-regulatory elements enriched uniquely at complex-bound sites. This study decoded the regulatory code mediated by the major flowering repressors SVP and FLC.
Project description:The transition to flowering in plants is controlled by a regulatory network that responds to both developmental and environmental signals. The MADS-box genes FLOWERING LOCUS C (FLC) and SHORT VEGETATIVE PHASE (SVP) are major flowering repressors that enhance responses to environmental cues such as winter temperatures, high ambient temperatures and photoperiod. FLC and SVP physically interact in vivo and mutation of each gene causes early flowering while the double mutant is more extreme. The molecular mechanisms underlying these genetic interactions are mostly unknown. We addressed the regulatory input of these two key transcription factors (TFs) both individually and as a complex at the genome-wide level through ChIP-seq and microarray expression analysis in single and double mutants. Analysis of each TF demonstrated that the complex acts predominantly via functional redundancy in the repression of flowering. SVP and FLC bind to the same regions of the flowering genes SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) and FLOWERING LOCUS T (FT) but do not require the presence of the other to bind. However, genome-wide identification of SVP and FLC occupancy events revealed that their binding scenarios are quantitatively and qualitatively affected by the presence of the cognate partner. A subgroup of genes whose regulation by these TFs depends exclusively on combinatorial binding of both proteins was identified, demonstrating a qualitatively essential role of the SVP-FLC complex. Some of these genes are involved in the control of flowering through direct and indirect regulation of Gibberellin-related processes. Cis-regulatory elements enriched only at such complex-bound sites were identified. Thus the regulatory output mediated by SVP and FLC reveals substantial flexibility, leading to dependent and independent DNA binding that enables additive, cooperative and repressive modes of co-regulation.
Project description:How plants control the transition to flowering in response to ambient temperature is only beginning to be understood. In Arabidopsis thaliana, the MADS-box transcription factor genes FLOWERING LOCUS M (FLM) and SHORT VEGETATIVE PHASE (SVP) have key roles in this process. FLM is subject to temperature-dependent alternative splicing, producing two splice variants, FLM-β and FLM-δ, which compete for interaction with the floral repressor SVP. The SVP/FLM-β complex is predominately formed at low temperatures and prevents precocious flowering. In contrast, the competing SVP FLM-δ complex is impaired in DNA binding and acts as a dominant negative activator of flowering at higher temperatures. Our results demonstrate the importance of temperature-dependent alternative splicing in modulating the timing of the floral transition in response to environmental change.
Project description:How plants control the transition to flowering in response to ambient temperature is only beginning to be understood. In Arabidopsis thaliana, the MADS-box transcription factor genes FLOWERING LOCUS M (FLM) and SHORT VEGETATIVE PHASE (SVP) have key roles in this process. FLM is subject to temperature-dependent alternative splicing, producing two splice variants, FLM-M-NM-2 and FLM-M-NM-4, which compete for interaction with the floral repressor SVP. The SVP/FLM-M-NM-2 complex is predominately formed at low temperatures and prevents precocious flowering. In contrast, the competing SVP FLM-M-NM-4 complex is impaired in DNA binding and acts as a dominant negative activator of flowering at higher temperatures. Our results demonstrate the importance of temperature-dependent alternative splicing in modulating the timing of the floral transition in response to environmental change. ChIP-seq A. thaliana FLM (3 replicates for gFLM and 2 replicates for FLM splice variants)
Project description:Many plants dramatically elongate their vegetative stems following flowering, yet it is unclear how this response is linked to the reproductive phase. We show that microRNA (miRNA) control of a deeply conserved phase change transcription factor, APETALA (AP2), is required for rapid and complete elongation of stem internodes in barley, especially of the final 'peduncle' internode directly underneath the inflorescence. Disrupted miR172-targeting of AP2 causes leads to short peduncles in the Zeo barley mutant due to reduced cell number and late stage cell expansion, which was associated with lower mitotic activity, delayed growth dynamics and premature lignification. In agreement, multiple, stage and tissue specific comparative transcriptomics revealed decreased expression of proliferation-associated genes, and ectopic expression of maturation-related genes in Zeo1.b. In addition, Zeo1.b peduncles showed persistent, elevated expression of genes associated with jasmonate (JA) and stress responses. Reproductive development including stem elongation in Zeo1.b was hypersensitive to inhibition by methyl JA (MeJA) but less responsive to promotion via gibberellin (GA). Based on these data, we propose that miR172- restriction of AP2 during flowering may release JA-associated vegetative growth restraint in order to facilitate GA-promoted stem growth in the reproductive phase. Microarray experiment M1. Comparison of control Bowman (Bw) and mutant (Zeo1.b) lines in peduncle initials tissue.
Project description:We were interested in changes in small RNA abundance changes in response to developmental transitions in Arabidopsis thaliana shoots, with special focus on vegetative phase change. We specifically wanted to separate the temporal changes in gene expression that result from vegetative phase change and those from flowering. Because of the close timing between the juvenile-to-adult and adult-to-reproductive developmental transitions in Arabidopsis grown under long day conditions, we used the late-flowering genotype FRI;FLC developed by the lab of Richard Amasino by introgressing the FRI allele from Sf-2 into the Col-0 genetic background, which is fri;FLC. For the early flowering genotype, we used FRI;flc-3, also developed by the Amasino lab by EMS-mutagenizing FRI;FLC, identifying early flowering mutants, and backcrossing multiple times to eliminate other EMS-induced mutations. The onset of vegetative phase change in FRI;FLC and FRI;flc-3 under our growth conditions was identical, but the progression was slower in FRI;FLC. By sequencing small RNAs from shoot apices at different time points and fully-expanded leaves at different positions on the shoot and comparing the results between the two genotypes, we were able to obtain a clear picture of changes in small RNA abundance in response to vegetative phase change and flowering in Arabidopsis. For the small RNA samples, we performed two replicates using two different indices in the 5'-adapter and ran each replicate pair on the same sequencing lane. For the cotyledon and leaf samples we only performed one replicate using the same index for all samples because we obtained significantly different results with the two adapters used for the shoot apices, preventing us from using them as true replicates.
Project description:Plants of three different genotypes (FRI FLC, FRI flc and fri flc) were induced to flowering by shifting from short day conditions to long day conditions. FRI=FRIGIDA, FLC=FLOWERING LOCUS C.
Project description:Many plants dramatically elongate their vegetative stems following flowering, yet it is unclear how this response is linked to the reproductive phase. We show that microRNA (miRNA) control of a deeply conserved phase change transcription factor, APETALA (AP2), is required for rapid and complete elongation of stem internodes in barley, especially of the final 'peduncle' internode directly underneath the inflorescence. Disrupted miR172-targeting of AP2 causes leads to short peduncles in the Zeo barley mutant due to reduced cell number and late stage cell expansion, which was associated with lower mitotic activity, delayed growth dynamics and premature lignification. In agreement, multiple, stage and tissue specific comparative transcriptomics revealed decreased expression of proliferation-associated genes, and ectopic expression of maturation-related genes in Zeo1.b. In addition, Zeo1.b peduncles showed persistent, elevated expression of genes associated with jasmonate (JA) and stress responses. Reproductive development including stem elongation in Zeo1.b was hypersensitive to inhibition by methyl JA (MeJA) but less responsive to promotion via gibberellin (GA). Based on these data, we propose that miR172- restriction of AP2 during flowering may release JA-associated vegetative growth restraint in order to facilitate GA-promoted stem growth in the reproductive phase. Microarray experiment MS. Comparison of control Bowman (Bw) and mutant (Zeo1.b) lines in spike (developing flower) tissue at 21 days post-anthesis.