Histone Deacetylases Play An Important Role In Bacterial Wilt Resistance Of Tomato

ObjectiveBacterial wilt which caused by Ralstonia solanacearum was a major constraint on the production of tomato. In plants, HDACs modulated the expression of developmental, stress-response and defense against pathogens. In this research, 15 genes of histone deacetylases family were found in tomato through bioinformatics analysis. The results of tissue expression pattern and oxidative stress showed that 15 genes were involved in bacterial wilt resistance. These studies mainly focus on the separation, identification and molecular mechanisms of HDACs, and will lay the foundation for improving resistance to bacterial wilt through molecular biology and genetic engineering techniques.MethodThe expression of HDACs under stress treatment in different organizations of tomato were analyzed by GEO microarray, and were verified by Q-PCR. Western-blot was used to analyze the differences in expression of histone deacetylateses between resistant cultivars and susceptible cultivars. It also detected the different expression level of resistance marker genes by Ch IP assay. The vector of p Can Gmyc-Sl HDT3 and p YLRNAi 5-35S-Sl HDT3 were constructed. The concentration of kanamycin and hygromycin were optimized. The conventional PCR was applied to detect Henz 1706 tomato genome exogenous gene neomycin phosphotransferase(NptⅡ)and hygromycin phosphotransferase(HPT) which are both exogenous in transgenic plants.Results1. Bioinformatics analysis indicated that there were 15 tomato histone deacetylases genes.2. Sl HDA6 、 Sl HDA9 and Sl HDA10 were upregulated in resistant cultivar, while down-regulated in susceptible cultivars. Sl HDA8 、Sl HDT1 、Sl HDT3 and Sl SRT1 were doun-regulated in resistant cultivars, while in susceptible cultivars they were upregulated.The changes in expression levels of other genes were similar in resistant and susceptible cultivars.3. Sl ERF transcription factors and hormone pathways marker genes were upregulated in resistant cultivars, and down-regulated in susceptible cultivars; many transcription factor, such as Sl CBF1、Sl Pti5、Sl WRKY39、Sl WRKY80 had more higher response to bacterial wilt.4. Acetylation levels of the transcription factor Sl ERF(H3ac, H4ac) increased in resistant cultivars, which indicated that acetylation was involved in bacterial wilt resistance.5. The susceptible cultivars wereincreased resistant to bacterial wilt afer treatment with histone deacetylase inhibitors(TSA) treatment.6. The optimal concentration of kanamycin and hygromycin in Henz 1706 tomato genetic transformate system was as follows: 20 mg/L Kan and 4 mg/L Hyg.7. p Can Gmyc-Sl HDT3 and p YLRNAi 5-35S-Sl HDT3 expression vectors have been constructed; there are 5 and 6 transgenic plants which were transformed by p Can Gmyc-Sl HDT3 and p YLRNAi 5-35S-Sl HDT3, respectively.ConclusionIn this study, 15 HDAC genes were identified from tomato genome through bioinformatics, and play an important role in response to oxidative stress and bacterial wilt, which was preliminary clarified through comparing between histone deacetylation levels by western-blot and expression of the defense marker genes, and TSA treatment can improve the resistant of bacterial in susceptible cultivars.