Comparative Analysis Of Cotton Anther Histone H3K4me3 And H3K27me3 Modification Under High Temperature Stress

Cotton is an important economic crop,and its fiber is a natural textile raw material,which has many uses in industry.Xinjiang is the main cotton-producing region in C hina,and there are often persistent high temperature(HT)weather in summer,and summer is an important period of cotton reproductive development,the male reproductive organs of cotton are more sensitive to HT stress than the female reproductive organs.HT stress caused abnormal development of cotton male reproductive organs,such as indehiscence anthers,reduced pollen vitality,etc.Our previous research showed that histone modification in epigenetics might participate in the HT stress response in cotton anthers.Thus,in the current study,HT-tolerant line ‘84021' and HT-sensitive line ‘H05' tapetal degradation stage(TDS,9-14 mm)and anther dehiscence stage(ADS,>24 mm)under normal temperature(N T)and HT were used as experimental materials through chromatin immunoprecipitation(Ch IP)to capture fragments modified by H3K4me3 and H3K27 me3 in anthers.Combined with methods such as Ch IP-Seq,RNA-Seq,q RT-PCR and bioinformatics analysis,the changes of whole genome histone H3K4me3 and H3K27me3 modifications were analyzed at TDS and ADS in ‘84021'and ‘H05' anthers under NT and HT conditions.The main findings are as follows: 1.Ch IP-Seq analysis showed that the pearson correlation coefficients of two biological replicates of H3K4me3 and H3K27me3 were above 0.7,most of which were above 0.9;MACS software was used to visualization ofthe H3K4me3 and H3K27me3 modification peaks on the genome,the results showed H3K4me3 peaks were mainly distributed in the first exon region,H3K27me3 peaks were mainly distributed in the intergenic region,and no significant differences in the peak distribution regions of the two modifications under NT and HT.2.According to RNA-Seq data,H3K4me3 modifications were enriched in high expression genes,while H3K27me3 modifications were enriched in low expression genes and the two were in a ‘heart-shaped' distribution.3.Genome-wide analysis showed that the H3K4me3 modification significantly increased during ADS anthers of ‘84021'and ‘H05' under HT,and there were no obvious differences at TDS.At TDS,the H3K27me3 modification significantly decreased in the ‘84021' anthers under HT,however,that was significantly increased in ‘H05'.At ADS,the H3K27me3 modification in ‘84021' anthers increased significantly under HT but unchanged in ‘H05'.Then,we obtained 33 histone H3K4me3 and H3K27me3 methytransferase and demethylase homologous genes in cotton.Tissue expression analysis showed 8 genes highly expression in anther,including 4 H3K4me3 methyltransferase genes,2 H3K4me3 demethylase genes and 2 H3K27me3 demethylase genes.The expression of these 8 genes was analyzed during anther development stages under NT and HT conditions,the results showed that 8 genes have lower expression at TDS compared to ADS.In addition,at ADS,the expression of 8 homologous genes was down-regulated by HT in ‘84021',and the expression of H3K4me3 demethylase genes showed more decreases than H3K4me3 methyltransferase genes after HT.In the ‘H05',the expression of H3K4me3 methyltransferase genes and H3K4me3 demethylase genes was up-regulated and down-regulated by HT,respectively.And the H3K27me3 demethylase genes showed low expression under NT were suppressed by HT.Therefore the expression changes of H3K4me3 methyltransferase genes,H3K4me3 demethylase genes,and H3K27me3 demethylase genes under HT in ‘84021' and ‘H05' at ADS,consistent with the changes of H3K4me3 and H3K27me3 modifications in the whole genome under HT in ‘84021' and ‘H05' at ADS.4.At the gene body region,H3K4me3 modification levels in anthers of ‘84021' and‘H05' were almost unchanged at TDS,and slightly decreased at ADS under HT;H3K27me3 modification levels were decreased in ‘84021' and ‘H05'under HT and the decline trends of the two materials were similar at TDS.However,the H3K27me3 modification in ‘84021' decreased significantly under HT at ADS,while ‘H05' decreased slightly.Further analysis of the number of H3K4me3 and H3K27me3 modified genes revealed that,more genes lost the H3K27me3 modification in anthers compares to H3K4me3 modification under HT at TDS and ADS.Then these genes have lost H3K27me3 modifications under HT at TDS and ADS of ‘84021' and ‘H05' were performed GO analysis,the results revealed that the different ial genes in ‘84021' two stages involved in many common pathways,there were common biological processes: cell wall modification,malate transport,and phosphatidylcholine metabolic process;the common cell component was the cell wall;the common molecular functions were transferase activity,pectinesterase activity,and protein serine/threonine kinase activity.Among these pathways,only the expression of malate transport pathway related geneswas up-regulated by HT in ‘84021' anthers at ADS.In summary,genome-wide histone H3K4me3 and H3K27me3 modifications,gene body,GO and the expression changes analysis in ‘84021' and ‘H05' anthers at TDS and ADS under NT and HT conditions were performed,we found HT induced obvious changes in two lines' H3K27me3 modification,but no significant changes in H3K4me3 modification.Then we found HT stress caused a large number of genes to lost H3K27me3 modification in TDS and ADS anthers of ‘84021',and the loss of H3K27me3 modified genes might mainly affect malate transport to maintain anther fertility of ‘84021' under HT.