Project description:Plants regulate their time to flowering by gathering information from the environment. Photoperiod and temperature are among the most important environmental variables. Suboptimal, but not near-freezing, temperatures regulate flowering through the thermosensory pathway, which overlaps with the autonomous pathway. Here we show that ambient temperature regulates flowering by two genetically distinguishable pathways, one that requires TFL1 and another that requires ELF3. The delay in flowering time observed at lower temperatures was partially suppressed in single elf3 and tfl1 mutants, whereas double elf3 tfl1 mutants were insensitive to temperature. tfl1 mutations abolished the temperature response in cryptochrome mutants that are deficient in photoperiod perception, but not in phyB mutants that have a constitutive photoperiodic response. Contrary to tfl1, elf3 mutations were able to suppress the temperature response in phyB mutants, but not in cryptochrome mutants. The gene expression profile revealed that the tfl1 and elf3 effects are due to the activation of different sets of genes and identified CCA1 and SOC1/AGL20 as being important cross talk points. Finally, genome-wide gene expression analysis strongly suggests a general and complementary role for ELF3 and TFL1 in temperature signalling. Three genotypes, WT (Columbia), elf3-7 and tfl1-1 mutants. Three biological replicates for each condition (genotype X temperature combination). RNA prepared independently for each sample.
Project description:Plants regulate their time to flowering by gathering information from the environment. Photoperiod and temperature are among the most important environmental variables. Suboptimal, but not near-freezing, temperatures regulate flowering through the thermosensory pathway, which overlaps with the autonomous pathway. Here we show that ambient temperature regulates flowering by two genetically distinguishable pathways, one that requires TFL1 and another that requires ELF3. The delay in flowering time observed at lower temperatures was partially suppressed in single elf3 and tfl1 mutants, whereas double elf3 tfl1 mutants were insensitive to temperature. tfl1 mutations abolished the temperature response in cryptochrome mutants that are deficient in photoperiod perception, but not in phyB mutants that have a constitutive photoperiodic response. Contrary to tfl1, elf3 mutations were able to suppress the temperature response in phyB mutants, but not in cryptochrome mutants. The gene expression profile revealed that the tfl1 and elf3 effects are due to the activation of different sets of genes and identified CCA1 and SOC1/AGL20 as being important cross talk points. Finally, genome-wide gene expression analysis strongly suggests a general and complementary role for ELF3 and TFL1 in temperature signalling.
Project description:The epigenetic silencing of the Arabidopsis floral repressor gene FLOWERING LOCUS C (FLC) is induced by a prolonged period of cold and promotes the developmental transition to flowering post-cold. FLC silencing requires the function of VRN1, a non-sequence-specific DNA binding protein. Here, we used Arabidopsis seedlings growing at ambient temperature carrying a FLAG-tagged VRN1 under its endogenous promoter in the vrn1FRI mutant background. We performed FLAG immunoprecipitation followed by mass spectrometry (IP-MS) on crosslinked tissue. The proteomics analysis revealed several proteins that support the role of VRN1 as a chromatin regulator. The identified proteins associated with VRN1 in vivo also support its contribution to gene silencing.
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:Dynamic trimethylation of histone H3 at Lys27 (H3K27me3) affects gene expression and controls plant development and environmental responses. In Arabidopsis thaliana, RELATIVE OF EARLY FLOWERING 6/JUMONJI DOMAIN-CONTAINING PROTEIN 12 (REF6/JMJ12) demethylates H3K27me3 by recognizing a specific DNA motif; however, little is known about how REF6 activates target gene expression after recognition, especially in environmental responses. In response to warm ambient temperature, plants undergo thermomorphogenesis, which involves accelerated growth, early flowering, and changes in morphology. Here we show that REF6 regulates thermomorphogenesis and cooperates with the transcription factor PHYTOCHROME INTERACTING FACTOR 4 (PIF4) to synergistically activate thermo-responsive genes under warm ambient temperature. The ref6 loss-of-function mutants exhibited attenuated hypocotyl elongation at warm temperature, partially due to down-regulation of GIBBERELLIN 20-OXIDASE2 (GA20ox2) and BASIC HELIX-LOOP-HELIX 87 (bHLH87). REF6 enzymatic activity is necessary for warm ambient temperature responses. Together, our results provide direct evidence of an epigenetic modifier and a transcription factor working together to respond to the environment.
Project description:Plants integrate seasonal cues such as temperature and day length to optimally adjust their flowering time to the environment. Compared to the control of flowering before and after winter by the vernalization and day length pathways, mechanisms that delay or promote flowering during a transient cool or warm period, especially during spring, are less well known. Due to global warming, understanding this ambient temperature pathway has become increasingly important. FLOWERING LOCUS M (FLM) is one critical flowering regulator of this pathway in Arabidopsis thaliana. We identified the Arabidopsis accession Kil-0 as an early flowering strain when compared to the common reference accession Col-0. Genetic mapping of this trait identified a causative region of around 31 kb at the bottom of chromosome one. Within this region, only FLOWERING LOCUS M (FLM) was expressed at a significantly lower level in Kil-0 when comparing RNA-seq (RNA sequencing) data from 10-day old Kil-0 and Col-0 plants grown at 21C. Furthermore, FLM was also the gene with the greatest reduction in gene expression between Kil-0 and Col-0 when we specifically analyzed 267 genes with a role in flowering time regulation what strongly suggested that FLM is the major locus that results in accelerated flowering in Kil-0.
Project description:Plants coordinate their growth and developmental programs with various endogenous signals and environmental challenges such as seasonal and diurnal temperature fluctuations. The bHLH transcription factor PIF4 plays critical roles in thermoresponsive hypocotyl growth in Arabidopsis, and the evening complex component ELF3 negatively regulates PIF4's activity for downstream gene expression and hypocotyl elongation at elevated temperature. However, how warm temperature signal is transmitted to ELF3 is not known. Here, we report the identification of two B-Box protein BBX18/BBX23 as new regulators of thermomorphogenesis in Arabidopsis. Mutations of BBX18/BBX23 confer reduced thermoresponsive hypocotyl elongation. Overexpression of BBX18 enhances the sensitivity of hypocotyl growth to elevated temperature, which is dependent on the function of PIF4 and RING E3 ligase COP1, respectively. Both BBX18 and BBX23 interact with ELF3 or COP1, relegating the protein abundance of ELF3 at warm temperature. Further, the expression of multiple thermoresponsive genes is impaired in both the PIF4 single mutant and BBX18/BBX23 double mutant. In addition, both the transcription and protein levels of BBX18/BBX23 are up-regulated by elevated ambient temperature. Thus, our findings reveal the important roles of B-Box proteins in plant thermomorphogenesis, and build a new connection from warm temperature information to ELF3 and its downstream signaling components.
Project description:The timing of flowering is a critical agronomic trait governed by a number of different internal and external events, such as temperature, light and photoperiod. Despite a firm understanding of flowering time from a genetic standpoint, there exists a limited understanding of how post-translational modifications may influence flowering, and what molecular PTM machinery is responsible for catalyzing these events. Here, we identify and characterize a novel B4 Raf-like MAPKKK protein kinase called RAF24, which exhibits an early flowering phenotype. Analysis of closely-related B4 Raf-like MAPKKK protein kinases RAF18 and RAF20, indicate that RAF24 is the only B4 RAF to possess an early flowering phenotype. Through quantitative proteomics, we then found that RAF24 impacts the phosphorylation status of numerous flowering related proteins related to the autonomous flowering pathway, with changes in the phosphorylation status of HISTONE MONO-UBIQUITINATION 2 (HUB2) representing the largest fold-change. Subsequent analysis of the HUB2 protein interactome by quantitative affinity purification mass spectrometry (AP-MS) revealed a relationship between HUB2 phosphorylation at S314 phosphorylation status and HUB2 function that manifest as changes in activity and protein-protein interactions. Use of phospho-memetic and -ablative mutants validate the importance of S314, directly linking this HUB2 phosphorylation event to the proper control of flowering time. Collectively, our findings implicate RAF24 as a higher-order regulator of flowering, while further implicating HUB2 as a centerpiece of flowering time regulation in plants.
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)