The trxG family histone methyltransferase SET DOMAIN GROUP 26 promotes flowering via a distinctive genetic pathway

Histone methylation is a major component in numerous processes such as determination of flowering time, which is fine‐tuned by multiple genetic pathways that integrate both endogenous and environmental signals. Previous studies identified SET DOMAIN GROUP 26 (SDG26) as a histone methyltransferase involved in the activation of flowering, as loss of function of SDG26 caused a late‐flowering phenotype in Arabidopsis thaliana. However, the SDG26 function and the underlying molecular mechanism remain largely unknown. In this study, we undertook a genetic analysis by combining the sdg26 mutant with mutants of other histone methylation enzymes, including the methyltransferase mutants Arabidopsis trithorax1 (atx1), sdg25 and curly leaf (clf), as well as the demethylase double mutant lsd1‐like1 lsd1‐like2 (ldl1 ldl2). We found that the early‐flowering mutants sdg25, atx1 and clf interact antagonistically with the late‐flowering mutant sdg26, whereas the late‐flowering mutant ldl1 ldl2 interacts synergistically with sdg26. Based on microarray analysis, we observed weak overlaps in the genes that were differentially expressed between sdg26 and the other mutants. Our analyses of the chromatin of flowering genes revealed that the SDG26 protein binds at the key flowering integrator SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1/AGAMOUS‐LIKE 20 (SOC1/AGL20), and is required for histone H3 lysine 4 trimethylation (H3K4me3) and histone H3 lysine 36 trimethylation (H3K36me3) at this locus. Together, our results indicate that SDG26 promotes flowering time through a distinctive genetic pathway, and that loss of function of SDG26 causes a decrease in H3K4me3 and H3K36me3 at its target gene SOC1, leading to repression of this gene and the late‐flowering phenotype.     

Chromatin interacting factor OsVIL2 increases biomass and rice grain yield

Grain number is an important agronomic trait. We investigated the roles of chromatin interacting factor Oryza sativa VIN3‐LIKE 2 (OsVIL2), which controls plant biomass and yield in rice. Mutations in OsVIL2 led to shorter plants and fewer grains whereas its overexpression (OX) enhanced biomass production and grain numbers when compared with the wild type. RNA‐sequencing analyses revealed that 1958 genes were up‐regulated and 2096 genes were down‐regulated in the region of active division within the first internodes of OX plants. Chromatin immunoprecipitation analysis showed that, among the downregulated genes, OsVIL2 was directly associated with chromatins in the promoter region of CYTOKININ OXIDASE/DEHYDROGENASE2 (OsCKX2), a gene responsible for cytokinin degradation. Likewise, active cytokinin levels were increased in the OX plants. We conclude that OsVIL2 improves the production of biomass and grain by suppressing OsCKX2 chromatin.     

Regulation of flowering time by the histone deacetylase HDA5 in Arabidopsis

The acetylation level of histones on lysine residues regulated by histone acetyltransferases and histone deacetylases plays an important but under‐studied role in the control of gene expression in plants. With the aim of characterizing the Arabidopsis RPD3/HDA1 family histone deacetylase HDA5, we present evidence showing that HDA5 displays deacetylase activity. Mutants defective in the expression of HDA5 displayed a late‐flowering phenotype. Expression of the flowering repressor genes FLC and MAF1 was up‐regulated in hda5 mutants. Furthermore, the gene activation markers, histone H3 acetylation and H3K4 trimethylation on FLC and MAF1 chromatin were increased in hda5‐1 mutants. Chromatin immunoprecipitation analysis showed that HDA5 binds to the chromatin of FLC and MAF1. Bimolecular fluorescence complementation assays and co‐immunoprecipitation assays showed that HDA5 interacts with FVE, FLD and HDA6, indicating that these proteins are present in a protein complex involved in the regulation of flowering time. Comparing gene expression profiles of hda5 and hda6 mutants by RNA‐seq revealed that HDA5 and HDA6 co‐regulate gene expression in multiple development processes and pathways.     

Mutagenesis of GmFT2a and GmFT5a mediated by CRISPR/Cas9 contributes for expanding the regional adaptability of soybean

Flowering time is a key agronomic trait that directly influences the successful adaptation of soybean (Glycine max) to diverse latitudes and farming systems. GmFT2a and GmFT5a have been extensively identified as flowering activators and integrators in soybean. Here, we identified two quantitative trait loci (QTLs) regions harbouring GmFT2a and GmFT5a, respectively, associated with different genetic effects on flowering under different photoperiods. We analysed the flowering time of transgenic plants overexpressing GmFT2a or GmFT5a, ft2a mutants, ft5a mutants and ft2aft5a double mutants under long‐day (LD) and short‐day (SD) conditions. We confirmed that GmFT2a and GmFT5a are not redundant, they collectively regulate flowering time, and the effect of GmFT2a is more prominent than that of GmFT5a under SD conditions whereas GmFT5a has more significant effects than GmFT2a under LD conditions. GmFT5a, not GmFT2a, was essential for soybean to adapt to high latitude regions. The ft2aft5a double mutants showed late flowering by about 31.3 days under SD conditions and produced significantly increased numbers of pods and seeds per plant compared to the wild type. We speculate that these mutants may have enormous yield potential for the tropics. In addition, we examined the sequences of these two loci in 202 soybean accessions and investigated the flowering phenotypes, geographical distributions and maturity groups within major haplotypes. These results will contribute to soybean breeding and regional adaptability.     

Arabidopsis O‐GlcNAc transferase SEC activates histone methyltransferase ATX1 to regulate flowering

Post‐translational modification of proteins by O‐linked β‐N‐acetylglucosamine (O‐GlcNAc) is catalyzed by O‐GlcNAc transferases (OGTs). O‐GlcNAc modification of proteins regulates multiple important biological processes in metazoans. However, whether protein O‐GlcNAcylation is involved in epigenetic processes during plant development is largely unknown. Here, we show that loss of function of SECRET AGENT (SEC), an OGT in Arabidopsis, leads to an early flowering phenotype. This results from reduced histone H3 lysine 4 trimethylation (H3K4me3) of FLOWERING LOCUS C (FLC) locus, which encodes a key negative regulator of flowering. SEC activates ARABIDOPSIS HOMOLOG OF TRITHORAX1 (ATX1), a histone lysine methyltransferase (HKMT), through O‐GlcNAc modification to augment ATX1‐mediated H3K4me3 histone modification at FLC locus. SEC transfers an O‐GlcNAc group on Ser947 of ATX1, which resides in the SET domain, thereby activating ATX1. Taken together, these results uncover a novel post‐translational O‐GlcNAc modification‐mediated mechanism for regulation of HKMT activity and establish the function of O‐GlcNAc signaling in epigenetic processes in plants.     

Distinct roles of the histone chaperones NAP1 and NRP and the chromatin‐remodeling factor INO80 in somatic homologous recombination in Arabidopsis thaliana

Homologous recombination (HR) of nuclear DNA occurs within the context of a highly complex chromatin structure. Despite extensive studies of HR in diverse organisms, mechanisms regulating HR within the chromatin context remain poorly elucidated. Here we investigate the role and interplay of the histone chaperones NUCLEOSOME ASSEMBLY PROTEIN1 (NAP1) and NAP1‐RELATED PROTEIN (NRP) and the ATP‐dependent chromatin‐remodeling factor INOSITOL AUXOTROPHY80 (INO80) in regulating somatic HR in Arabidopsis thaliana. We show that simultaneous knockout of the four AtNAP1 genes and the two NRP genes in the sextuple mutant m123456‐1 barely affects normal plant growth and development. Interestingly, compared with the respective AtNAP1 (m123‐1 and m1234‐1) or NRP (m56‐1) loss‐of‐function mutants, the sextuple mutant m123456‐1 displays an enhanced plant hypersensitivity to UV or bleomycin treatments. Using HR reporter constructs, we show that AtNAP1 and NRP act in parallel to synergistically promote somatic HR. Distinctively, the AtINO80 loss‐of‐function mutation (atino80‐5) is epistatic to m56‐1 in plant phenotype and telomere length but hypostatic to m56‐1 in HR determinacy. Further analyses show that expression of HR machinery genes and phosphorylation of H2A.X (γ‐H2A.X) are not impaired in the mutants. Collectively, our study indicates that NRP and AtNAP1 synergistically promote HR upstream of AtINO80‐mediated chromatin remodeling after the formation of γ‐H2A.X foci during DNA damage repair.     

Phenotypic reversion in fas mutants of Arabidopsis thaliana by reintroduction of FAS genes: variable recovery of telomeres with major spatial rearrangements and transcriptional reprogramming of 45S rDNA genes

Arabidopsis thaliana mutants dysfunctional in the evolutionarily conserved protein complex chromatin assembly factor‐1 (CAF‐1), which deposits the canonical histone H3 variant H3.1 during DNA synthesis‐dependent chromatin assembly, display complex phenotypic changes including meristem and growth alterations, sensitivity to DNA‐damaging agents, and reduced fertility. We reported previously that mutants in the FAS1 subunit of CAF‐1 progressively lose telomere and 45S rDNA repeats. Here we show that multiple aspects of the fas phenotype are recovered immediately on expression of a reintroduced FAS1 allele, and are clearly independent of the recovery of rDNA copy‐numbers and telomeres. In reverted lines, 45S rDNA genes are recovered to diverse levels with a strikingly different representation of their variants, and the typical association of nucleolar organizing region 4 with the nucleolus is perturbed. One of 45S rDNA variants (VAR1), which is silenced in wild‐type (WT) plants without mutation history (Col‐0 WT), dominates the expression pattern, whereas VAR2 is dominant in Col‐0 WT plants. We propose an explanation for the variability of telomere and 45S rDNA repeats associated with CAF‐1 function, suggesting that the differences in nuclear partitioning and expression of the rDNA variants in fas mutants and their revertants provide a useful experimental system to study genetic and epigenetic factors in gene dosage compensation.