Functional Analysis Of A Histone Deacetylases Gene (SlHDA5) In Tomato (Solanum Lycopersicum )

Histone deacetylation, one of the histone modifications, is a very important mean of epigenetic regulations. Histone deacetylation is involved in the regulations of DNA replication, transcription, DNA damage repair, gene silence, cell differentiation and cycle. In chromatin, histone deacetylation can not only make nucleosome stable, but also restore the interactions between histones and DNA, which can impede the specific combinations of DNA and protein factors. Histone deacetylases(HDACs) are the main enzymes in the process of deacetylation, which can remove acetyl from the lysine residues of histone, promote the interactions of histone and silencers, and supperss the transcription. HDACs play important roles in the process of not only growth and development but also hormone and abiotic stress. In recent years, many HDACs of Arabidopsis and rice have been isolated and identified. However, little is known about the function and mechanism of regulational HDACs in tomato.In this paper, we selected wild type(WT) tomato as research material and studied the member of HDACs, Sl HDA5.The experiment contents and results are as follows:(1) According to the serial number of Sl HDA5, we got the sequence of the gene in the solanaceae genome library, which encodes a protein with 347 amino acids. Compared the amino acids sequence of Sl HDA5 with HDACs whose functions were already known well in other species' HDACs, the result showed that Sl HDA5 contained a HDAC structure domain as well as other known proteins. Meanwhile, the phylogenetic tree analysis showed that Sl HDA5 was orthologous to At HDA2, and we can conclude that Sl HDA5 gene belongs to III class of RPD3 / HDA1 family.(2) Analysis of Sl HDA5 tissue-specific expression pattern in tomato showed that this gene expressed mainly in root and the expression level increased in the process of leaf development and fruit maturity. This suggested that Sl HDA5 may be involved in the development of ripening of tomato fruit.(3) The expression levels of Sl HDA5 in wild type(WT) tomato seedlings under hormone treatments(ABA, SA, Me JA and IAA) were analyzed. The q RT-PCR results showed that the expression of Sl HDA5 was significantly induced by exogenous hormone ABA and Me JA, while no obvious changes were observed under SA and IAA treatment, suggesting that Sl HDA5 may be involved in ABA and Me JA regulation.(4) The expression levels of Sl HDA5 gene in wild type(WT) tomato seedlings under abiotic treatments(high/low temperature, dehydration and salt) were also analyzed. The q RT-PCR results showed that the Sl HDA5 transcription level was also significantly induced by high/low temperature, dehydration and salt, indicating that Sl HDA5 may be involved in abiotic stresses.(5) The RNAi vector targeting Sl HDA5 was constructed using RNA interference technology, and introduced into WT tomato to generate the transgenic lines by agrobacterium-mediated transformation method. However, no difference was observed in the fruits of wild type and transgenic tomato lines.(6) We further investigated the effects of drought and salt stress on Sl HDA5-RNAi transgenic lines and WT. The Sl HDA5-RNAi lines showed wilting symptom much earlier than the WT when exposed to drought. The drought-related physiological indexes were detected in transgenic lines and WT. The results showed that transgenic lines had lower levels of water and chlorophyll contents, and higher levels of MDA contents in their leaves as compared to wild-type plants under the same drought-stressed conditions. Otherwise, the expression levels of two stress-related genes(PR1 and TAS14) were down-regulated in roots of RNAi plants after 24 and 48 hours of salt-stressed treatment. The leaf salt assay also showed that transgenic plants displayed less tolerance than wild-type plants demonstrated by retaining lower chlorophyll content. These results suggested that Sl HDA5 gene silencing results in decreased tolerance to drought and salt stress.(7) The sensitivity of transgenic T1 seedlings to salinity and ABA were conducted. The results demonstrated that the root and shoot growth of RNAi plants were more inhibited by salt stress and ABA than that of wild-type at post-germination stage, suggesting that repressed expression of Sl HDA5 lead transgenic seedlings more hypersusceptible to salinity and exogenous ABA than WT.In conclusion, Sl HDA5 gene is the member of III class of RPD3 / HDA1 family. Sl HDA5 mainly expressed in roots, sepals, flowers and fruits and was induced by exogenous hormone and abiotic stress, such as ABA, Me JA, salt, high temperature and dehydration. Moreover, silencing of Sl HDA5 resulted in decreased tolerance to drought and salt stress, and Sl HDA5-RNAi transgenic seedlings were more hypersusceptible to salinity and exogenous ABA than WT. These results indicate that Sl HDA5 might take part in the drought and salt stress responses.