Analysis Of Histone Deacetylase Gene Families In Arabidopsis Thaliana And The Molecular Mechanism Of AtSRT2 Controlling Seed Germination Under Salt Stress

Accumulated evidences indicated that epigenetics modification including DNA methylation and histone methylation played a key role in plant growth, development and the adaptation to the changing environments. However, few results about the biological functions and regulation mechanism of histone acetylation was reported so far. Therefore, in this study, we analyzed the histone deacetylases(HDACs) in Arabidopsis thaliana including their phylogenetic relationship, genomic structure, tissue-specific expression patterns, expression pattern under stresses and phenotype of the mutants, which would be useful to understand their biological functions.At the same time, the sirtuin-like histone deacetylases from human were involved in some processes including chromatin silent, DNA damage repair, cell cycle regulation and autophagy. However, biological functions of sirtuin-like histone deacetylases in plants still remained unknown. Therefore, a sirtuin-like histone deacetylase, At SRT2 was selected to identify its gene function and molecular mechanism in Arabidopsis. The major results were showed as following:1. Characteristics analysis of histone deacetylases family in Arabidopsis thaliana. Phylogenetic tree analysis of HDACs family in Arabidopsis thaliana(At HDACs) showed that At HDACs family was grouped into three types. Analysis of the genomic structure of At HDACs showed that their intron was abundant in their genomes. 18 members of At HDACs were unevenly located on 5 different chromosome locations, and most members containing 5 and 9. Tissue-specific expression analysis showed higher expression of At HDACs were detected in leaf, silique and seed compared with other tissues. The expression of At HDACs were not sharply up or down under hormones and abotic stress treatment. Mutants of members of At HDACs were various during plant development compared with wild type, some mutants of At HDACs displayed abnormal leaf,flower morphology and flowering time.2. Sirtuin like histone deacetylases At SRT2 controlled seed germination under salt stress. Under normal condition, the srt2 plants displayed similar phenotype with wild type plants, but srt2 showed a salt-sensitive phynotype during seed germination. One splice variants of At SRT2, At SRT2.7 had a highest expression level in seed compared with other splice variants, and the expression of At SRT2.7 was induced by salt stress during seed germination. Overexpression of At SRT2.7 in srt2-1 partly rescued the salt-sensitive phenotype of srt2-1, which indicated that At SRT2.7 was responsive for seed germination under salt stress.3. At SRT2 alleviated damage impact on plant genome and reduced NAD+ accumulation caused by salt stress during seed germination. SCGE showed that during seed germination stage, salt stress induced DNA damage in Arabidopiss, and srt2 was more sensitive than WT. Compared with wild type srt2 was more sensitive to MMS treatment. On the other hand compared with wild type, srt2 had a higher NAD+ content during seed germination under salt stress, suggested that accumulation of NAD+ inhibited seed germination. and srt2 was more sensitive to NAD+ treatment. These results indicated that At SRT2 involved in the process of NAD+ metabolism, and the salt-sensitive phenotype of srt2 was caused by DNA damage and NAD+ accumulation.4. Expression of a salt stress responsive gene At VAMP714 was repressed by At SRT2.7 through deacetylation of H4K8 location. At SRT2.7 located in nucleus and membrane, and showed a NAD+ dependent histone deacetylation activity in vivo and in vitro, specifically deacetylated H3K4, H3K19 and H4K8. H4K8 acetylation in WT was gradually reduced during seed germination under salt stress, whereas no obvious H4K8 acetylation change was observed in srt2. These results indicated that At SRT2.7 controlled seed germination by H4K8 deacetylaion under salt stress. The Ch IP-Seq and Ch IP-q PCR assay showed that the expression of a salt stress responsive gene At VAMP714 was repressed by At SRT2.7 through H4K8 deacetylation, and then reduced damage impact on tonoplast caused by H2O2, finally promoted seed germination under salt stress.In short, At HDACs were widely involved in plant growth, development and adaptation to environment, and At HDACs have various biological functions in Arabidopsis thaliana. A Sirtuin like protein, At SRT2 mediated seed germination under salt stress in different ways. First, At SRT2 alleviated the damage impact on DNA caused by salt stress during seed germination; Second, At SRT2 catalyzed NAD+ metabolized to nicotinamide, and reduced NAD+ accumulation that represses seed germination. This research enriched our knowledge about the biological functions of At HDACs and paved a way to study epigenetic related network in plants.