组蛋白修饰对喉癌细胞系中 RASSF1A 基因表达的影响

摘要

Objective: To explore the correlation between RASSF1A gene expression and gene histone modification. Methods: ChIP and Realtime - PCR were used to detect H3 - K9 methylation,H3 - K4 methylation,and H3 -K9 acetylation of RASSF1A gene promoter region and gene expression before and after treatment with drugs 5 - Aza — 2'- deoxyeytidine (5 - Aza-2'- deoxyeytidine,5 - Aza - dC, DNA methyltransferase inhibitor 5 - Aza - dC) and TriChostatinA (TriChostatinA,TSA,histone deacetylase inhibitor)in laryngeal carcinoma Hep-2 cell line. Results: TSA was able to reduce H3 - K9 methylation slightly, increase H3 - K4 methylation slightly, and increase H3 - K9 acetylation significantly of RASSF1A gene promoter region histone. 5 - Aza - dC was able to reduce H3 - K9 methylation significantly, increase H3 - K4 methylation significantly, increase H3 - K9 acetylation. Synergistic effect was observed with combination of 5 - Aza — dC and TSA. The reducing of H3 - K9 methylation and increasing of H3 - K4 methylation and H3 - K9 acetylation were able to up - regulate gene expression. Conclusion: Down - regulation of RASSF1A tumor suppressor gene expression in laryngeal carcinoma cells is correlated with H3 - K9 methylation ,H3 — K4 methylation and H3 - K9 acetylation in its promoter region. Both methylation inhibitor and acetylating agent are able to up — regulate the down - regulated gene expression.%目的:探讨喉癌细胞中RASSF1A基因表达水平与基因组蛋白修饰的关系.方法:应用染色质免疫沉淀技术(ChIP)和实时定量逆转录聚合酶链反应(Realtime-PCR)分析喉癌Hep-2细胞系在以下药物:5-氮杂-2 '-脱氧胞苷(5-Aza-2'-deoxyeytidine,5-Aza-dC,DNA甲基转移酶抑制剂,5-AZa-dC)；曲古抑菌素A(TriChostatinA,TSA,组蛋白去乙酰化酶抑制剂)作用前后RASSF1A基因启动子区域组蛋白H3-K9甲基化、H3-K4甲基化、H3-K9乙酰化和RASSF1A基因表达情况.结果:TSA能轻度降低RASSF1A基因启动子区域组蛋白H3-K9甲基化水平、轻度提高H3-K4甲基化水平、明显提高H3-K9乙酰化水平.5-Aza-dC明显降低启动子区域H3-K9甲基化程度、明显提高H3-K4甲基化水平、提高H3-K9乙酰化水平,联合给予5-Aza-dC和TSA起协同增效作用.H3-K9甲基化水平降低、H3-K4甲基化和H3-K9乙酰化水平提高可使RASSF1A基因表达上调.结论:喉癌细胞中RASSF1A肿瘤抑制基因表达下调与其启动子区H3-K9甲基化、H3-K4甲基化和H3-K9乙酰化相关.甲基化抑制剂及乙酰化制剂均可使表达下调的基因表达上调.