8-氯腺苷诱导的组蛋白H3K79双甲基化促进NBS1和p21的基因转录激活

8-Chloro-adenosine-induced H3K79me2 Promotes Transcriptional Activation of NBS1 and p21 Genes in Response to DNA Double-stranded Breaks

组蛋白H3第79位赖氨酸甲基化(H3K79me)修饰有单甲基、双甲基及三甲基3种形式,是常染色质的标志.然而,对于组蛋白H3K79三种甲基化各自在基因转录、DNA损伤修复中所起的作用尚不十分清楚.本研究以8-氯腺苷(8-Cl-Ado)为DNA双链断裂(DNAdouble-stranded breaks,DSB)诱导剂,采用Western印迹,在人肺癌细胞H1299检测出了DNA修复分子NBS1、细胞周期检验点相关分子p21,并发现H3K79me1、H3K79me2和H3K79me3三种甲基化修饰的组蛋白明显增加;染色质免疫共沉淀结合实时定量PCR实验显示,只有H3K79me2与DNA损伤检验点分子p21、DNA修复分子NBS1的启动子区域相结合,说明H3K79双甲基化修饰与这些基因的转录激活有关.结果提示,在8-氯腺苷引起DSB时,是H3K79me2、而不是H3K79me1和H3K79me3参与NBS1和p21基因转录激活时的染色质重塑.8-氯腺苷诱导H3K79双甲基化增强、促进H3K79me2所在染色质区域的NBS1和p21基因转录激活可能是8-Cl-Ado抑制肿瘤细胞生长作用机制之一.

Methylation of H3K79, including mono, di and trimethyled H3K79 （H3K79me1, H3K79me2 and H3K79me3）, is one of the landmark modifications associated with euehromatin. However, the roles of methylated H3K79 in gene regulation, DNA damage and repair are unclear. In this study, we showed that the levels of NBS1 （a DNA damage response protein）, p21 （a DNA damage checkpoint protein）, and three methylated H3K79 were markedly up-regulated in 8-Cl-Ado-exposed H1299 cells, evidenced by Western blotting. The combination of chromatin immunoprecipitation （CHIP） with real-time quantitative PCR （RT-qPCR） revealed that only H3K79me2 was associated with the NBS1 and p21 promoters, indicating the contribution of H3K79me2, to transcriptional activation of these genes. These results suggest that H3K79me2, but not H3K79me1 and H3K79me3, is involved in the chromatin remodeling associated with the transcription activation of the NBS1 and p21 genes. Induction of H3K79me2 by 8-Cl-Ado in the chromatin surrounding the NBS1 and p21 genes may be a way by that 8-Cl-Ado inhibits tumor cell proliferation.