The Effect Of DZNep On The Methylation Status Of Histone H3-lysine9at The Promoters Of MOB1A/B Genes And Its Effect On The Gene Expression In Leukemia Molt-4Cell Line

Objective：To detect the expression of MOB1A/B in de novo AL patients andinvestigate its clinical significance. To explore the effect of DZNep on themethylation of H3K9at the promoters of MOB1A/B genes and the expression ofMOB1A/B, and then provide a new direction for the treatment of acute leukemia.Methods：The expression of MOB1A/B was detected and its clinical significancewas analysised in clinical samples with quantitative real-time polymerase chainreaction and Western-blot method. The CCK-8assay methods was used to evaluatethe effect on situation of cell proliferation in Molt-4cells after disposal of DZNep;Apoptosis ratio was detected by flow cytometry with Annexin V，FITC/PI; Real-time qPCR and Western blot were applied to identify the expression level ofMOB1A/B; The methylation status of MOB1A/B gene in Molt-4cell line wasmeasured by bisulfite sequencing PCR. The methylation status of H3K9at thepromoters of MOB1A/B genes was measured before and after treated with DZNepusing chromatin immunoprecipitation (ChIP) followed by qPCR.Results：1. The expression of MOB1A/B in de novo ALL patients wassignificantly decreased (P<0.05).2. CCK-8assay demonstrated that DZNep couldinhibit the proliferation of Molt-4cells, which also increase apoptosis by flowcytometry (P <0.05). The expression of MOB1B gene increased after treated withDZNep (P<0.05), but the increase of MOB1A was not significant.3. Highmethylation rates of MOB1A were presented in Molt-4cell line, however, themethylation levels of MOB1B were very low.4. Treatment with DZNep inhibitedmethylation of lysine9on histone H3in the cultured human leukemia Molt-4cells.Conclusion: The core components of Hippo signaling pathway, MOB1A/B, weresignificantly decreased in de novo AL patients, which is closely related to themethylation of H3K9at the promoters of MOB1A/B genes and/or high methylationlevel of CpG island. The histone methylase inhibitor, DZNep, reversed histone H3K9methylation levels at the promoters of MOB1B genes, and reversed the expression of MOB1B gene, significantly inhibited the proliferation and increased apoptosis of ofMolt-4cell. Our study laid the experimental foundation for the role of the Hipposignaling pathway in the pathogenesis and development of acute leukemia, andprovided a new direction for the treatment of acute leukemia.