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.     

A histone methyltransferase inhibits seed germination by increasing PIF1 mRNA expression in imbibed seeds

Phytochrome‐interacting factor 1 (PIF1) inhibits light‐dependent seed germination. The specific function of PIF1 in seed germination is partly due to its high level of expression in imbibed seeds, but the associated regulatory factors have not been identified. Here we show that mutation of the early flowering in short days (EFS) gene, encoding an H3K4 and H3K36 methyltransferase, decreases the level of H3K36me2 and H3K36me3 but not H3K4me3 at the PIF1 locus, reduces the targeting of RNA polymerase II to the PIF1 locus, and reduces mRNA expression of PIF1 in imbibed seeds. Consistently, the efs mutant geminated even under the phyB_off condition, and had an expression profile of PIF1 target genes similar to that of the pif1 mutant. Introduction of an EFS transgene into the efs mutant restored the level of H3K36me2 and H3K36me3 at the PIF1 locus, the high‐level expression of PIF1 mRNA, the expression pattern of PIF1 target genes, and the light‐dependent germination of these seeds. Introduction of a PIF1 transgene into the efs mutant also restored the expression pattern of PIF1 target genes and light‐dependent germination in imbibed seeds, but did not restore the flowering phenotype. Taken together, our results indicate that EFS is necessary for high‐level expression of PIF1 mRNA in imbibed seeds.     

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.     

An Arabidopsis E3 ligase HUB2 increases histone H2B monoubiquitination and enhances drought tolerance in transgenic cotton

The HUB2 gene encoding histone H2B monoubiquitination E3 ligase is involved in seed dormancy, flowering timing, defence response and salt stress regulation in Arabidopsis thaliana. In this study, we used the cauliflower mosaic virus (CaMV) 35S promoter to drive AtHUB2 overexpression in cotton and found that it can significantly improve the agricultural traits of transgenic cotton plants under drought stress conditions, including increasing the fruit branch number, boll number, and boll‐setting rate and decreasing the boll abscission rate. In addition, survival and soluble sugar, proline and leaf relative water contents were increased in transgenic cotton plants after drought stress treatment. In contrast, RNAi knockdown of GhHUB2 genes reduced the drought resistance of transgenic cotton plants. AtHUB2 overexpression increased the global H2B monoubiquitination (H2Bub1) level through a direct interaction with GhH2B1 and up‐regulated the expression of drought‐related genes in transgenic cotton plants. Furthermore, we found a significant increase in H3K4me3 at the DREB locus in transgenic cotton, although no change in H3K4me3 was identified at the global level. These results demonstrated that AtHUB2 overexpression changed H2Bub1 and H3K4me3 levels at the GhDREB chromatin locus, leading the GhDREB gene to respond quickly to drought stress to improve transgenic cotton drought resistance, but had no influence on transgenic cotton development under normal growth conditions. Our findings also provide a useful route for breeding drought‐resistant transgenic plants.     

Extensive allele‐level remodeling of histone methylation modification in reciprocal F1 hybrids of rice subspecies

Epigenetic mechanisms play a major role in heterosis, partly as a result of the remodeling of epigenetic modifications in F₁ hybrids. Based on chromatin immunoprecipitation‐sequencing (ChIP‐Seq) analyses, we show that at the allele level extensive histone methylation remodeling occurred for a subset of genomic loci in reciprocal F₁ hybrids of Oryza sativa (rice) cultivars Nipponbare and 93‐11, representing the two subspecies japonica and indica. Globally, the allele modification‐altered loci in leaf or root of the reciprocal F₁ hybrids involved ?12–43% or more of the genomic regions carrying either of two typical histone methylation markers, H3K4me3 (>21 000 genomic regions) and H3K27me3 (>11 000 genomic regions). Nevertheless, at the total modification level, the majority (from ?43 to >90%) of the modification‐altered alleles lay within the range of parental additivity in the hybrids because of concerted alteration in opposite directions, consistent with an overall attenuation of allelic differences in the modifications. Importantly, of the genomic regions that did show non‐additivity in total modification level by either marker in the two tissues of hybrids, >80% manifested transgressivity, which involved genes enriched in specific functional categories. Extensive allele‐level alteration of H3K4me3 alone was positively correlated with genome‐wide changes in allele‐level gene expression, whereas at the total level, both H3K4me3 and H3K27me3 remodeling, although affecting just a small number of genes, contributes to the overall non‐additive gene expression to variable extents, depending on tissue/marker combinations. Our results emphasize the importance of allele‐level analysis in hybrids to assess the remodeling of epigenetic modifications and their relation to changes in gene expression.     

Haplotype analysis of key genes governing grain yield and quality traits across 3K RG panel reveals scope for the development of tailor‐made rice with enhanced genetic gains

Though several genes governing various major traits have been reported in rice, their superior haplotype combinations for developing ideal variety remains elusive. In this study, haplotype analysis of 120 previously functionally characterized genes, influencing grain yield (87 genes) and grain quality (33 genes) revealed significant variations in the 3K rice genome (RG) panel. For selected genes, meta‐expression analysis using already available datasets along with co‐expression network provided insights at systems level. Also, we conducted candidate gene based association study for the 120 genes and identified 21 strongly associated genes governing 10‐grain yield and quality traits. We report superior haplotypes upon phenotyping the subset of 3K RG panel, SD1‐H8 with haplotype frequency (HF) of 30.13% in 3K RG panel, MOC1‐H9 (HF: 23.08%), IPA1‐H14 (HF: 6.64%), DEP3‐H2 (HF: 5.59%), DEP1‐H2 (HF: 37.53%), SP1‐H3 (HF: 5.05%), LAX1‐H5 (HF: 1.56%), LP‐H13 (3.64%), OSH1‐H4 (5.52%), PHD1‐H14 (HF: 15.21%), AGO7‐H15 (HF: 3.33%), ROC5‐H2 (31.42%), RSR1‐H8 (HF: 4.20%) and OsNAS3‐H2 (HF: 1.00%). For heading date, Ghd7‐H8 (HF: 3.08%), TOB1‐H10 (HF: 4.60%) flowered early, Ghd7‐H14 (HF: 42.60%), TRX1‐H9 (HF: 27.97%), OsVIL3‐H14 (HF: 1.72%) for medium duration flowering, while Ghd7‐H6 (HF: 1.65%), SNB‐H9 (HF: 9.35%) were late flowering. GS5‐H4 (HF: 65.84%) attributed slender, GS5‐H5 (HF: 29.00%), GW2‐H2 (HF: 4.13%) were medium slender and GS5‐H9 (HF: 2.15%) for bold grains. Furthermore, haplotype analysis explained possible genetic basis for superiority of selected mega‐varieties. Overall, this study suggests the possibility for developing next‐generation tailor‐made rice with superior haplotype combinations of target genes suiting future food and nutritional demands via haplotype‐based breeding.     

Long non‐coding RNA NR2F1‐AS1 promoted proliferation and migration yet suppressed apoptosis of thyroid cancer cells through regulating miRNA‐338‐3p/CCND1 axis

Thyroid cancer (TC) is a prevalent endocrine malignant cancer whose pathogenic mechanism remains unclear. The aim of the study was to investigate the roles of long non‐coding RNA (lncRNA) NR2F1‐AS1/miRNA‐338‐3P/CCND1 axis in TC progression. Differentially expressed lncRNAs and mRNAs in TC tissues were screened out and visualized by R program. Relative expression of NR2F1‐AS1, miRNA‐338‐3p and cyclin D1 (CCND1) was determined by quantitative real time polymerase chain reaction. In addition, Western blot analysis was adopted for evaluation of protein expression of CCND1. Targeted relationships between NR2F1‐AS1 and miRNA‐338‐3p, as well as miRNA‐338‐3p and CCND1 were predicted using bioinformatics analysis and validated by dual‐luciferase reporter gene assay. Besides, tumour xenograft assay was adopted for verification of the role of NR2F1‐AS1 in TC in vivo. NR2F1‐AS1 and CCND1 were overexpressed, whereas miRNA‐338‐3p was down‐regulated in TC tissues and cell lines. Down‐regulation of NR2F1‐AS1 and CCND1 suppressed proliferation and migration of TC cells yet greatly enhanced cell apoptotic rate. Silence of NR2F1‐AS1 significantly suppressed TC tumorigenesis in vivo. NR2F1‐AS1 sponged miRNA‐338‐3p to up‐regulate CCND1 expression to promote TC progression. Our study demonstrated that up‐regulation of NR2F1‐AS1 accelerated TC progression through regulating miRNA‐338‐3P/CCND1 axis.     

A photo‐responsive F‐box protein FOF2 regulates floral initiation by promoting FLC expression in Arabidopsis

Floral initiation is regulated by various genetic pathways in response to light, temperature, hormones and developmental status; however, the molecular mechanisms underlying the interactions between different genetic pathways are not fully understood. Here, we show that the photoresponsive gene FOF2 (F‐box of flowering 2) negatively regulates flowering. FOF2 encodes a putative F‐box protein that interacts specifically with ASK14, and its overexpression results in later flowering under both long‐day and short‐day photoperiods. Conversely, transgenic plants expressing the F‐box domain deletion mutant of FOF2 (FOF2ΔF), or double loss of function mutant of FOF2 and FOL1 (FOF2‐LIKE 1) present early flowering phenotypes. The late flowering phenotype of the FOF2 overexpression lines is suppressed by the flc‐3 loss‐of‐function mutation. Furthermore, FOF2 mRNA expression is regulated by autonomous pathway gene FCA, and the repressive effect of FOF2 in flowering can be overcome by vernalization. Interestingly, FOF2 expression is regulated by light. The protein level of FOF2 accumulates in response to light, whereas it is degraded under dark conditions via the 26S proteasome pathway. Our findings suggest a possible mechanistic link between light conditions and the autonomous floral promotion pathway in Arabidopsis.