Floral Transition In Arabidopsis Research Articles

ATX1 and HUB1/2 promote recruitment of the transcription elongation factor VIP2 to modulate the floral transition in Arabidopsis.

Histone 2B ubiquitination (H2Bub) and trimethylation of H3 at lysine 4 (H3K4me3) are associated with transcription activation. However, the function of these modifications in transcription in plants remains largely unknown. Here, we report that coordination of H2Bub and H3K4me3 deposition with the binding of the RNA polymerase-associated factor VERNALIZATION INDEPENDENCE2 (VIP2) to FLOWERING LOCUS C (FLC) modulates flowering time in Arabidopsis. We found that RING domain protein HISTONE MONOUBIQUITINATION1 (HUB1) and HUB2 (we refer as HUB1/2), which are responsible for H2Bub, interact with ARABIDOPSIS TRITHORAX1 (ATX1), which is required for H3K4me3 deposition, to promote the transcription of FLC and repress the flowering time. The atx1-2 hub1-10 hub2-2 triple mutant in FRIGIDIA (FRI) background displayed early flowering like FRI hub1-10 hub2-2 and overexpression of ATX1 failed to rescue the early flowering phenotype of hub1-10 hub2-2. Mutations in HUB1 and HUB2 reduced the ATX1 enrichment at FLC, indicating that HUB1 and HUB2 are required for ATX1 recruitment and H3K4me3 deposition at FLC. We also found that the VIP2 directly binds to HUB1, HUB2, and ATX1 and that loss of VIP2 in FRI hub1-10 hub2-2 and FRI atx1-2 plants resulted in early flowering like that observed in FRI vip2-10. Loss of function of HUB2 and ATX1 impaired VIP2 enrichment at FLC, and reduced the transcription initiation and elongation of FLC. In addition, mutations in VIP2 reduced HUB1 and ATX1 enrichment and H2Bub and H3K4me3 levels at FLC. Together, our findings revealed that HUB1/2, ATX1, and VIP2 coordinately modulate H2Bub and H3K4me3 deposition, FLC transcription, and flowering time.

Genetic and epigenetic basis of phytohormones control of floral transition in plants.

The timing of the developmental transition from the vegetative to the reproductive stages is critical for angiosperm and fine-tuned by the integration of endogenous factors and external environmental cues to ensure proper and successful reproduction. Plants have evolved sophisticated mechanisms to response to diverse environmental or stress signals, which may be mediated by plant hormones which coordinate their flowering time. Endogenous and exogenous phytohormones such as gibberellin (GA), auxin, cytokinin (CK), jasmonate (JA), abscisic acid (ABA), ethylene (ET), brassinosteroids (BR) and the cross-talk among them are critical for the precise regulating of flowering time. Recent studies on the model flowering plant Arabidopsis thaliana revealed that diverse transcription factors and epigenetic regulators play key roles in the phytohormones that regulate floral transition. This review aims to summarize current knowledge on the genetic and epigenetic mechanisms that underlying the phytohormone control of floral transition in Arabidopsis, offering insights into how these processes are regulated and their implications for plant biology.

MED8 regulates floral transition in Arabidopsis by interacting with FPA.

Success in plant reproduction is highly dependent on the correct timing of the floral transition, which is tightly regulated by the flowering pathways. In the model plant Arabidopsis thaliana, the central flowering repressor FLOWERING LOCUS C (FLC) is precisely regulated by multiple flowering time regulators in the vernalization pathway and autonomous pathway, including FPA. Here we report that Arabidopsis MEDIATOR SUBUNIT 8 (MED8) promotes floral transition in Arabidopsis by recruiting FPA to the FLC locus to repress FLC expression. Loss of MED8 function leads to a significant late-flowering phenotype due to increased FLC expression. We further show that MED8 directly interacts with FPA in the nucleus and recruits FPA to the FLC locus. Moreover, MED8 is indispensable for FPA's function in controlling flowering time and regulating FLC expression. Our study thus reveals a flowering mechanism by which the Mediator subunit MED8 represses FLC expression by facilitating the binding of FPA to the FLC locus to ensure appropriate timing of flowering for reproductive success.

FLK is an mRNA m6A reader that regulates floral transition by modulating the stability and splicing of FLC in Arabidopsis

N6-methyladenosine (m6A), which is added, removed, and interpreted by m6A writers, erasers, and readers, respectively, is the most abundant modification in eukaryotic mRNAs. The m6A marks play a pivotal role in the regulation of floral transition in plants. FLOWERING LOCUS K (FLK), an RNA-binding protein harboring K-homology (KH) motifs, is known to regulate floral transition by repressing the levels of a key floral repressor FLOWERING LOCUS C (FLC) in Arabidopsis. However, the molecular mechanism underlying FLK-mediated FLC regulation remains unclear. In this study, we identified FLK as a novel mRNA m6A reader protein that directly binds the m6A site in the 3ʹ-untranslated region of FLC transcripts to repressing FLC levels by reducing its stability and splicing. Importantly, FLK binding of FLC transcripts was abolished in vir-1, an m6A writer mutant, and the late-flowering phenotype of the flk mutant could not be rescued by genetic complementation using the mutant FLKm gene, in which the m6A reader encoding function was eliminated, indicating that FLK binds and regulates FLC expression in an m6A-dependent manner. Collectively, our study has addressed a long-standing question of how FLK regulates FLC transcript levels and established a molecular link between the FLK-mediated recognition of m6A modifications on FLC transcripts and floral transition in Arabidopsis.

ALKBH10B-mediated m6A demethylation is crucial for drought tolerance by affecting mRNA stability in Arabidopsis

N6-methyladenosine (m6A) is the most abundant modification found in eukaryotic mRNAs. The methyltransferases (“writers”) and demethylases (“erasers”) add and remove m6A marks, respectively, which controls transcriptome-wide m6A levels and plays crucial roles in plant development and response to environmental cues. The alpha-ketoglutarate-dependent dioxygenase homolog 10 (ALKBH10) is the first confirmed plant m6A eraser and is involved in floral transition in Arabidopsis (Arabidopsis thaliana). Recent studies showed that ALKBH10B is also involved in the Arabidopsis response to salt, drought or ABA. However, the molecular mechanism underlying the ALKBH10B-guided drought tolerance has not yet been explored. In this study, we investigated how ALKBH10B-mediated m6A demethylation confers drought tolerance in Arabidopsis. The alkbh10b mutants were sensitive to drought stress, whereas the ALKBH10B-overexpressing plants were tolerant to drought stress. Notably, under dehydration stress, the m6A levels of several drought stress positive effectors were elevated in the alkbh10b mutants, and their transcript abundance was lower in the mutants but higher in the transgenic lines. Importantly, the decay rates of these m6A-modified transcripts were significantly higher in the alkbh10b relative to the wild-type under dehydration stress. Collectively, these findings establish a molecular link between ALKBH10B-guided m6A demethylation and mRNA stability control in drought stress tolerance.

Negative regulation of floral transition in Arabidopsis by HOS15-PWR-HDA9 complex.

Arabidopsis HOS15/PWR/HDA9 repressor complex, which is similar to the TBL1/NcoR1/HDAC complex in animals, plays a well-known role in epigenetic regulation. PWR and HDA9 have been reported to interact with each other and modulate the flowering time by repressing AGL19 expression, whereas HOS15 and HDA9, together with the photoperiodic evening complex, regulate flowering time through repression of GI transcription. However, the role of the HOS15/PWR/HDA9 core repressor complex as a functional unit in the regulation of flowering time is yet to be explored. In this study, we reported that the loss-of-function hos15-2/pwr/hda9 triple mutant accumulates higher transcript levels of AGL19 and exhibits an early flowering phenotype similar to those of hos15, pwr, and hda9 single mutants. Interestingly, the accumulation of HOS15 in the nucleus was drastically reduced in pwr and hda9 mutants. As a result, HOS15 could not perform its role in histone deacetylation or interaction with H3 in the nucleus. Furthermore, HOS15 is also associated with the same region of the AGL19 promoter known for PWR-HDA9 binding. The acetylation level of the AGL19 promoter was increased in the hos15-2 mutant, similar to the pwr and hda9 mutants. Therefore, our findings reveal that the HOS15/PWR/HDA9 repressor complex deacetylates the promoter region of AGL19, thereby negatively regulating AGL19 transcription, which leads to early flowering in Arabidopsis.

Arabidopsis N6-methyladenosine methyltransferase FIONA1 regulates floral transition by affecting the splicing of FLC and the stability of floral activators SPL3 and SEP3.

N 6-methyladenosine (m6A) RNA methylation has been shown to play a crucial role in plant development and floral transition. Two recent studies have identified FIONA1 as an m6A methyltransferase that regulates the floral transition in Arabidopsis through influencing the stability of CONSTANS (CO), SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1), and FLOWERING LOCUS C (FLC). In this study, we confirmed that FIONA1 is an m6A methyltransferase that installs m6A marks in a small group of mRNAs. Furthermore, we show that, in addition to its role in influencing the stability of CO, SOC1, and FLC, FIONA1-mediated m6A methylation influences the splicing of FLC, a key floral repressor, and the stability of SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE 3 (SPL3) and SEPALLATA3 (SEP3), floral activators, which together play a vital role in floral transition in Arabidopsis. Our study confirms the function of FIONA1 as an m6A methyltransferase and suggests a close molecular link between FIONA1-mediated m6A methylation and the splicing of FLC and the destabilization of SPL3 and SEP3 in flowering time control.

Decoding Gene Expression Signatures Underlying Vegetative to Inflorescence Meristem Transition in the Common Bean

The tropical common bean (Phaseolus vulgaris L.) is an obligatory short-day plant that requires relaxation of the photoperiod to induce flowering. Similar to other crops, photoperiod-induced floral initiation depends on the differentiation and maintenance of meristems. In this study, the global changes in transcript expression profiles were analyzed in two meristematic tissues corresponding to the vegetative and inflorescence meristems of two genotypes with different sensitivities to photoperiods. A total of 3396 differentially expressed genes (DEGs) were identified, and 1271 and 1533 were found to be up-regulated and down-regulated, respectively, whereas 592 genes showed discordant expression patterns between both genotypes. Arabidopsis homologues of DEGs were identified, and most of them were not previously involved in Arabidopsis floral transition, suggesting an evolutionary divergence of the transcriptional regulatory networks of the flowering process of both species. However, some genes belonging to the photoperiod and flower development pathways with evolutionarily conserved transcriptional profiles have been found. In addition, the flower meristem identity genes APETALA1 and LEAFY, as well as CONSTANS-LIKE 5, were identified as markers to distinguish between the vegetative and reproductive stages. Our data also indicated that the down-regulation of the photoperiodic genes seems to be directly associated with promoting floral transition under inductive short-day lengths. These findings provide valuable insight into the molecular factors that underlie meristematic development and contribute to understanding the photoperiod adaptation in the common bean.

Replication protein RPA2A regulates floral transition by cooperating with PRC2 in Arabidopsis.

RPA2A is a subunit of the conserved heterotrimeric replication protein A (RPA) in Arabidopsis, which is an essential replisome component that binds to single-stranded DNA during DNA replication. RPA2A controls a set of developmental processes, but the underlying mechanism is largely unknown. Here we show that RPA2A represses key flowering genes including FLOWERING LOCUS T (FT), AGAMOUS (AG) and AGAMOUS LIKE 71 (AGL71) to suppress floral transition by cooperating with the PRC2 complex. RPA2A is vigorously expressed in dividing cells and required for correct DNA replication. Mutation of RPA2A leads to early flowering, which is dependent on ectopic expression of key flowering genes including FT molecularly and genetically. RPA2A and PRC2 have common target genes including FT, AG and AGL71 supported using genetic analysis, transcriptome profiling and H3K27me3 ChIP-seq analysis. Furthermore, RPA2A physically interacts with PRC2 components CLF, EMF2 and MSI1, which recruits CLF to the chromatin loci of FT, AG and AGL71. Together, our results show that the replication protein RPA2A recruits PRC2 to key flowering genes through physical protein interaction, thereby repressing the expression of these genes to suppress floral transition in Arabidopsis.

Phase separation of HRLP regulates flowering time in Arabidopsis.

RNA binding proteins mediate posttranscriptional RNA metabolism and play regulatory roles in many developmental processes in eukaryotes. Despite their known effects on the floral transition from vegetative to reproductive growth in plants, the underlying mechanisms remain largely obscure. Here, we show that a hitherto unknown RNA binding protein, hnRNP R-LIKE PROTEIN (HRLP), inhibits cotranscriptional splicing of a key floral repressor gene FLOWERING LOCUS C (FLC). This, in turn, facilitates R-loop formation near FLC intron I to repress its transcription, thereby promoting the floral transition in Arabidopsis thaliana. HRLP, together with the splicing factor ARGININE/SERINE-RICH 45, forms phase-separated nuclear condensates with liquid-like properties, which is essential for HRLP function in regulating FLC splicing, R-loop formation, and RNA Polymerase II recruitment. Our findings reveal that inhibition of cotranscriptional splicing of FLC by nuclear HRLP condensates constitutes the molecular basis for down-regulation of FLC transcript levels to ensure the reproductive success of Arabidopsis.

The long road to bloom in conifers

More than 600 species of conifers (phylum Pinophyta) serve as the backbone of the Earth’s terrestrial plant community and play key roles in global carbon and water cycles. Although coniferous forests account for a large fraction of global wood production, their productivity relies largely on the use of genetically improved seeds. However, acquisition of such seeds requires recurrent selection and testing of genetically superior parent trees, eventually followed by the establishment of a seed orchard to produce the improved seeds. The breeding cycle for obtaining the next generation of genetically improved seeds can be significantly lengthened when a target species has a long juvenile period. Therefore, development of methods for diminishing the juvenile phase is a cost-effective strategy for shortening breeding cycle in conifers. The molecular regulatory programs associated with the reproductive transition and annual reproductive cycle of conifers are modulated by environmental cues and endogenous developmental signals. Mounting evidence indicates that an increase in global average temperature seriously threatens plant productivity, but how conifers respond to the ever-changing natural environment has yet to be fully characterized. With the breakthrough of assembling and annotating the giant genome of conifers, identification of key components in the regulatory cascades that control the vegetative to reproductive transition is imminent. However, comparison of the signaling pathways that control the reproductive transition in conifers and the floral transition in Arabidopsis has revealed many differences. Therefore, a more complete understanding of the underlying regulatory mechanisms that control the conifer reproductive transition is of paramount importance. Here, we review our current understanding of the molecular basis for reproductive regulation, highlight recent discoveries, and review new approaches for molecular research on conifers.

Chrysanthemum lavandulifolium homolog ClMAD1 modulates the floral transition during temperature shift

Flower initiation and development in chrysanthemum requires accurate cell division and expansion, which are regulated by a complex network involved with many cell cycle regulators. Mitotic arrest deficiency 1 (MAD1) is a key component of spindle assembly checkpoint, which was reported to regulate floral transition in Arabidopsis through the interaction with Suppressor of FRIGIDA 4 (SUF4) to modulate Flowering Locus C (FLC) transcription. So far, no MAD1, SUF4 and FLC homologous genes, and related studies were reported in chrysanthemum. Here we first isolated and clone ClMAD1 and ClSUF4 homologs in Chrysanthemum lavandulifolium. Both ClMAD1 and ClSUF4 localized in nuclei, and their expression varied in different tissues. Overexpression of ClMAD1 in Arabidopsis promoted endopolyploidization in leaves, and restored cell cycle and flowering phenotypes of mad1 mutants to different extents. Overexpression of ClSUF4 in Arabidopsis resulted in significantly late flowering, which was correlated with enhanced transcription of FLC and reduced transcription of FT gene. ClMAD1 was predicted to have three coiled-coil (CC) domains, which is one more than that in Arabidopsis. Similar to AtSUF4, ClSUF4 was predicted to have two Zn-C2H2 domains. Protein-protein interaction assays revealed that ClSUF4 could interact with AtMAD1, and the third coiled-coil domain of ClMAD1 not the full-length protein. All these data suggest that both ClMAD1 and ClSUF4 are functional homologs. We further isolated their downstream signaling component ClFLC-like (ClFLCl) genes. ClFLCl had high expression in leaves at vegetative growth stage, and significantly reduced after the bolting suggesting a negative role in the floral transition. Long-term cold treatment significantly repressed the transcription of ClFLCl genes. Temperature shift promoted floral transition of chrysanthemum, which is correlated with the fluctuation of ClMAD1, ClSUF4 and ClFLCl transcription. Taken together, we reported a novel machinery on the regulation of floral transition in chrysanthemum that ClMAD1 interacts with ClSUF4 to modulate ClFLCl-mediated floral transition during the temperature shift.

Discovering candidate genes related to flowering time in the spring panel of Camelina sativa

Camelina sativa is a promising oilseed and industrial crop that benefits sustainable food, feed and fuel industries. Early flowering is critical for local adaptation as well as maximizing yield in Camelina sativa. Even though the preliminary data indicated wide variation in flowering time in the spring camelina germplasm, our understanding of underlying genes and their roles in regulating flower development is still limited. The current study combined genotypic data and flowering time from the spring panel, followed by genome-wide association study (GWAS) and whole-genome prediction to identify significant trait-associated markers and evaluate the predictive capability of the entire marker set. The analysis of phenotypic data showed significant genotypic and environmental effects on flowering time. A high heritability of 0.893 in flowering time suggests effectiveness of breeding early flowering camelina varieties. The GWAS analysis identified 20 significant trait-associated single nucleotide polymorphisms (SNPs) that colocalized within/or near a variety of transcription factors (e.g. SUPPRESSOR of PHYA-1/SPA1, BES1-INTERACTING MYC-LIKE 1/BIM1) or protein families containing specific functional domains (e.g. CCCH zinc finger protein family and B3-DNA binding domain containing protein family). These transcription factors were known to interact with key regulatory genes in the four major pathways (i.e. photoperiod, autonomous, vernalization and gibberellic acid pathways) to cooperatively regulate floral transition in arabidopsis. Whole-genome prediction showed a low-to-moderate predictive ability (0.559) to improve early flowering trait in camelina. This study is the first step for future in-depth exploration and genetic improvements of flower development and timing in camelina for breeding.

C-terminal domain phosphatase-like 1 (CPL1) is involved in floral transition in Arabidopsis

BackgroundRNA polymerase II plays critical roles in transcription in eukaryotic organisms. C-terminal Domain Phosphatase-like 1 (CPL1) regulates the phosphorylation state of the C-terminal domain of RNA polymerase II subunit B1, which is critical in determining RNA polymerase II activity. CPL1 plays an important role in miRNA biogenesis, plant growth and stress responses. Although cpl1 mutant showes delayed-flowering phenotype, the molecular mechanism behind CPL1’s role in floral transition is still unknown.ResultsTo study the role of CPL1 during the floral transition, we first tested phenotypes of cpl1-3 mutant, which harbors a point-mutation. The cpl1-3 mutant contains a G-to-A transition in the second exon, which results in an amino acid substitution from Glu to Lys (E116K). Further analyses found that the mutated amino acid (Glu) was conserved in these species. As a result, we found that the cpl1-3 mutant experienced delayed flowering under both long- and short-day conditions, and CPL1 is involved in the vernalization pathway. Transcriptome analysis identified 109 genes differentially expressed in the cpl1 mutant, with 2 being involved in floral transition. Differential expression of the two flowering-related DEGs was further validated by qRT-PCR.ConclusionsFlowering genetic pathways analysis coupled with transciptomic analysis provides potential genes related to floral transition in the cpl1-3 mutant, and a framework for future studies of the molecular mechanisms behind CPL1’s role in floral transition.

TEM1 combinatorially binds to FLOWERING LOCUS T and recruits a Polycomb factor to repress the floral transition in Arabidopsis

Arabidopsis TEMPRANILLO 1 (TEM1) is a transcriptional repressor that participates in multiple flowering pathways and negatively regulates the juvenile-to-adult transition and the flowering transition. To understand the molecular basis for the site-specific regulation of FLOWERING LOCUS T (FT) by TEM1, we determined the structures of the two plant-specific DNA-binding domains in TEM1, AP2 and B3, in complex with their target DNA sequences from the FT gene 5'-untranslated region (5'-UTR), revealing the molecular basis for TEM1 specificity for its DNA targets. In vitro binding assays revealed that the combination of the AP2 and B3 binding sites greatly enhanced the overall binding of TEM1 to the FT 5'-UTR, indicating TEM1 combinatorically recognizes the FT gene 5'-UTR. We further showed that TEM1 recruits the Polycomb repressive complex 2 (PRC2) to the FT 5'-UTR. The simultaneous binding of the TEM1 AP2 and B3 domains to FT is necessary for deposition of H3K27me3 at the FT 5'-UTR and for the flowering repressor function of TEM1. Overall, our data suggest that the combinatorial recognition of FT 5'-UTR by TEM1 ensures H3K27me3 deposition to precisely regulate the floral transition.

The histone variant H3.3 promotes the active chromatin state to repress flowering in Arabidopsis.

The histone H3 family in animals and plants includes replicative H3 and nonreplicative H3.3 variants. H3.3 preferentially associates with active transcription, yet its function in development and transcription regulation remains elusive. The floral transition in Arabidopsis (Arabidopsis thaliana) involves complex chromatin regulation at a central flowering repressor FLOWERING LOCUS C (FLC). Here, we show that H3.3 upregulates FLC expression and promotes active histone modifications histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 36 trimethylation (H3K36me3) at the FLC locus. The FLC activator FRIGIDA (FRI) directly mediates H3.3 enrichment at FLC, leading to chromatin conformation changes and further induction of active histone modifications at FLC. Moreover, the antagonistic H3.3 and H2A.Z act in concert to activate FLC expression, likely by forming unstable nucleosomes ideal for transcription processing. We also show that H3.3 knockdown leads to H3K4me3 reduction at a subset of particularly short genes, suggesting the general role of H3.3 in promoting H3K4me3. The finding that H3.3 stably accumulates at FLC in the absence of H3K36me3 indicates that the H3.3 deposition may serve as a prerequisite for active histone modifications. Our results reveal the important function of H3.3 in mediating the active chromatin state for flowering repression.

DEK domain-containing proteins control flowering time in Arabidopsis.

Evolutionarily conserved DEK domain-containing proteins have been implicated in multiple chromatin-related processes, mRNA splicing and transcriptional regulation in eukaryotes. Here, we show that two DEK proteins, DEK3 and DEK4, control the floral transition in Arabidopsis. DEK3 and DEK4 directly associate with chromatin of related flowering repressors, FLOWERING LOCUS C (FLC), and its two homologs, MADS AFFECTING FLOWERING4 (MAF4) and MAF5, to promote their expression. The binding of DEK3 and DEK4 to a histone octamer in vivo affects histone modifications at FLC, MAF4 and MAF5 loci. In addition, DEK3 and DEK4 interact with RNA polymerase II and promote the association of RNA polymerase II with FLC, MAF4 and MAF5 chromatin to promote their expression. Our results show that DEK3 and DEK4 directly interact with chromatin to facilitate the transcription of key flowering repressors and thus prevent precocious flowering in Arabidopsis.

Dual Recognition of H3K4me3 and DNA by the ISWI Component ARID5 Regulates the Floral Transition in Arabidopsis.

Chromatin remodeling and histone modifications are important for development and floral transition in plants. However, it is largely unknown whether and how these two epigenetic regulators coordinately regulate the important biological processes. Here, we identified three types of Imitation Switch (ISWI) chromatin-remodeling complexes in Arabidopsis (Arabidopsis thaliana). We found that AT-RICH INTERACTING DOMAIN5 (ARID5), a subunit of a plant-specific ISWI complex, can regulate development and floral transition. The ARID-PHD dual domain cassette of ARID5 recognizes both the H3K4me3 histone mark and AT-rich DNA. We determined the ternary complex structure of the ARID5 ARID-PHD cassette with an H3K4me3 peptide and an AT-containing DNA. The H3K4me3 peptide is combinatorially recognized by the PHD and ARID domains, while the DNA is specifically recognized by the ARID domain. Both PHD and ARID domains are necessary for the association of ARID5 with chromatin. The results suggest that the dual recognition of AT-rich DNA and H3K4me3 by the ARID5 ARID-PHD cassette may facilitate the association of the ISWI complex with specific chromatin regions to regulate development and floral transition.

Mutagenesis of a Quintuple Mutant Impaired in Environmental Responses Reveals Roles for CHROMATIN REMODELING4 in the Arabidopsis Floral Transition.

Several pathways conferring environmental flowering responses in Arabidopsis (Arabidopsis thaliana) converge on developmental processes that mediate the floral transition in the shoot apical meristem. Many characterized mutations disrupt these environmental responses, but downstream developmental processes have been more refractory to mutagenesis. Here, we constructed a quintuple mutant impaired in several environmental pathways and showed that it possesses severely reduced flowering responses to changes in photoperiod and ambient temperature. RNA-sequencing (RNA-seq) analysis of the quintuple mutant showed that the expression of genes encoding gibberellin biosynthesis enzymes and transcription factors involved in the age pathway correlates with flowering. Mutagenesis of the quintuple mutant generated two late-flowering mutants, quintuple ems1 (qem1) and qem2 The mutated genes were identified by isogenic mapping and transgenic complementation. The qem1 mutant is an allele of the gibberellin 20-oxidase gene ga20ox2, confirming the importance of gibberellin for flowering in the absence of environmental responses. By contrast, qem2 is impaired in CHROMATIN REMODELING4 (CHR4), which has not been genetically implicated in floral induction. Using co-immunoprecipitation, RNA-seq, and chromatin immunoprecipitation sequencing, we show that CHR4 interacts with transcription factors involved in floral meristem identity and affects the expression of key floral regulators. Therefore, CHR4 mediates the response to endogenous flowering pathways in the inflorescence meristem to promote floral identity.

Brassinosteroid Signaling Recruits Histone 3 Lysine-27 Demethylation Activity to FLOWERING LOCUS C Chromatin to Inhibit the Floral Transition in Arabidopsis

The steroid hormone brassinosteroid (BR) plays a broad role in plant growth and development. As the retarded growth in BR-insensitive and BR-deficient mutants causes a strong delay in days to flowering, BR signaling has been thought to promote the floral transition in Arabidopsis. In this study, using a developmental measure of flowering time, we show that BR signaling inhibits the floral transition and promotes vegetative growth in the Arabidopsis accessions Columbia and Enkheim-2. We found that BR signaling promotes the expression of the potent floral repressor FLOWERING LOCUS C (FLC) and three FLC homologs to inhibit flowering. In the presence of BR, the transcription factor BRASSINAZOLE-RESISTANT1 (BZR1), together with BES1-INTERACTING MYC-like proteins (BIMs), specifically binds a BR- responsive element in the first intron of FLC and further recruits a histone 3 lysine 27 (H3K27) demethylase to downregulate levels of the repressive H3K27 trimethylation mark and thus antagonize Polycomb silencing at FLC, leading to its activation. Taken together, our findings demonstrate that BR signaling inhibits the floral transition in Arabidopsis by a novel molecular mechanism in which BR signals are transduced into FLC activation and consequent floral repression.