乙酰化STAT3诱导的DIRAS2缺失促进三阴性乳腺癌细胞的增殖  

Acetylated STAT3-induced DIRAS2 deletion promotes the proliferation of triple-negative breast cancer cells

作　　者：张丽芬 王璐 赵琳[1] 罗敏娜 邵珊[1] 顾珊智[3] ZHANG Lifen;WANG Lu;ZHAO Lin;LUO Minna;SHAO Shan;GU Shanzhi(Department of Oncology,The First Affiliated Hospital of Xi’an Jiaotong University,Xi’an 710061;Department of Hematology,The First Affiliated Hospital of Xi’an Jiaotong University,Xi’an 710061;Department of Forensic Medicine,Xi’an Jiaotong University,Xi’an 710061,China)

机构地区：[1]西安交通大学第一附属医院肿瘤内科,陕西西安710061 [2]西安交通大学第一附属医院血液内科,陕西西安710061 [3]西安交通大学法医学系,陕西西安710061

出　　处：《西安交通大学学报（医学版）》2024年第5期741-747,共7页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：国家自然科学基金资助项目(No.81972473)。

摘　　要：目的探讨乙酰化STAT3对DIRAS2基因表达的调控及其在三阴性乳腺癌(triple-negative breast cancer,TNBC)细胞增殖中的作用。方法通过数据库查询、Western blotting和qRT-PCR分析TNBC组织和细胞中DIRAS2和乙酰化STAT3的表达水平。选取TNBC细胞系MDA-MB-231和SUM159,通过慢病毒或质粒构建DIRAS2过表达及STAT3野生或Lys685位点突变的细胞株。利用CCK-8实验评估DIRAS2和STAT3乙酰化对TNBC细胞增殖的影响。应用Western blotting、焦磷酸测序、ChIP和IP技术研究乙酰化STAT3对DIRAS2表达的调控作用及机制。结果DIRAS2在TNBC组织和细胞中表达较低。焦磷酸测序分析发现,与正常乳腺上皮细胞相比,TNBC细胞中DIRAS2启动子区域的CpG岛甲基化水平升高,并促进癌细胞增殖。此外,TNBC细胞中STAT3的乙酰化程度增加,而DIRAS2启动子甲基化状态随着STAT3乙酰化的增加而发生改变。ChIP和IP实验表明,乙酰化STAT3可与DIRAS2启动子结合,而STAT3 Lys685位点突变会破坏STAT3与DNMT1的相互作用。结论在TNBC中,乙酰化STAT3通过招募DNMT1诱导DIRAS2基因启动子甲基化,从而导致DIRAS2表达缺失和癌细胞增殖。Objective To explore the regulation of DIRAS2 gene expression by acetylated STAT3 and its involvement in the proliferation of triple-negative breast cancer(TNBC)cells.Methods The expression levels of DIRAS2 and acetylated STAT3 in TNBC tissues and cells were analyzed by database query,Western blotting,and qRT-PCR.TNBC cell lines MDA-MB-231 and SUM159 were selected,and lentivirus or plasmid was used to construct DIRAS2 overexpression and STAT3 wild or Lys685 mutation cell lines.The CCK-8 assay was used to evaluate the effect of DIRAS2 and STAT3 acetylation on the proliferation of TNBC cells.Western blotting,pyrosequencing,ChIP and IP were employed to investigate the regulatory effect and mechanism of acetylated STAT3 on DIRAS2 expression.Results The expression of DIRAS2 was decreased in TNBC tissues and cells.Pyrosequencing analysis found that the methylation level of CpG islands in the DIRAS2 promoter was increased in TNBC cells compared with normal breast epithelial cells,which promoted the growth of cancer cells.Furthermore,TNBC cells showed an increase in STAT3 acetylation,which was accompanied by a shift in the methylation status of the DIRAS2 promoter.ChIP and IP experiments showed that acetylated STAT3 could bind to the DIRAS2 promoter,and the STAT3 Lys685 mutation disrupted the interaction between STAT3 and DNMT1.Conclusion Acetylated STAT3 induces DIRAS2 promoter methylation by recruiting DNMT1,leading to loss of DIRAS2 expression and cancer cell proliferation in TNBC.

关 键 词：三阴性乳腺癌(TNBC) 乙酰化STAT3 DIRAS2 甲基化 增殖 

分 类 号：R737.9[医药卫生—肿瘤]