| Summary: | 碩士 === 國立臺灣大學 === 植物科學研究所 === 97 === Acetylation and deacetylation of nucleosomal core histones play important roles in regulation of eukaryotic transcriptional activity. Histone acetylation levels are determined by the action of histone acetyltransferases and histone deacetylases. Acetylation of the histone is often associated with increased gene activity, whereas deacetylation of histones is correlated with transcriptional repression. Recent studies indicated that histone acetylation is involved in plant response to ABA and abiotic stresses including salt, drought and cold stresses. However, little is known about the molecular mechanisms of how histone acetylation and deacetylation are involved in stress response in plants. In this study, I focus on ABA, salt, drought and cold stresses to study the role of Arabidopsis histone deacetylases HDA6 and HDA19 in ABA response and abiotic stress signaling. HDA6 and HDA19 are members of RPD3/HDA1 histone deacetylases family, and they have similar amino acid sequence and gene expression patterns in different development stages. A hda6 mutant line, axe1-5, and a HDA6 RNA interfering line, CS24039, as well as a HDA19 T-DNA insertion line, athd1-t1, were used to study the role of HDA6 and HDA19 in ABA and abiotic stress signaling. axe1-5 is a splicing mutant which carries a point mutation in the HDA6 splicing site, athd1-t1 has a T-DNA inserted in second exon of HDA19.
Compared with wild-type, axe1-5, CS24039 and athd1-t1 displayed higher sensitivity to ABA during the seed germination stage, and lower expression level of ABA-responsive genes (ABI1, ABI2, KAT1, KAT2 and RD29B). In high salinity stress, axe1-5 and CS24039 displayed lower germination rates and survival rates compared with wild-type. However, athd1-t1 has no significant difference with wild-type in high salinity stress. Compared with wild-type, axe1-5 and CS24039 displayed lower expression of salt-responsive genes (MYB2, RD29B, DREB2A and RD29A) under salt stress, but athd1-t1 displayed lower expression of RD29B and DREB2A only under salt stress. These results suggested that HDA6 might plat an important role in salt stress signaling pathway. Phenotypic comparison and RT-PCR analysis of cold stress responsive genes indicated that there was no significant difference between wild types, hda6 and hda19 mutants, suggesting that HDA6 and HDA19 are not involved in cold stress signaling pathways.
I further analyzed the H3 acetylation and methylation levels of the salt and ABA response genes by chromatin immunoprecipitation assay. It was found that ABA and salt can increase the H3 acetylation and methylation level of ABA and salt response genes (ABI1, ABI2, KAT1, KAT2, DREB2A, RD29A and RD29B), and wild-type and axe1-5 displayed different level of H3 acetylation and methylation. These results suggested that ABA and salt can induces the response gene expression by altering the H3 acetylation and methylaiton level, and HDA6 is required for this process.
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