UHRF2通过其RING结构域与TIP60作用进而调控组蛋白H3K9和H3K14乙酰化水平
[Abstract]:Background UHRF2 is a multi domain nuclear protein with E3 ubiquitin ligase, involved in cell cycle, DNA damage repair, epigenetics and ubiquitin proteasome system. In our previous work, our group confirmed that UHRF2 participates in the regulation of HBV transcription and replication and regulates the level of histone H3 acetylation with CCC DNA. Recent research We found that UHRF2 and histone H3K9ac and H3K14ac interact with each other. Based on the above research results, we continue to explore the regulatory mechanism between UHRF2 and histone acetylation. Methods this paper mainly through the Western blot test to verify the regulatory role of UHRF2 on H3K9ac, H3K14ac, TIP60, H2AK5ac and HDAC1, immunization. The interaction between UHRF2 and TIP60 and HDAC1 was verified by the precipitation (Co-IP) experiment; the immunofluorescence staining (IF) test verified the relationship between UHRF2 and the cell Sublocation of TIP60 and HDAC1; the half life experiment verified the regulation of UHRF2 on TIP60 half life; in vivo ubiquitination test verified the ubiquitination of UHRF2 to TIP60; immunohistochemical staining. IHC test verified the regulatory relationship between UHRF2 and H3K9ac, H3K14ac, TIP60 and H2AK5a. Results 1) Western blot experiments confirmed that overexpression of UHRF2 in normal cells can up regulate the expression of H3K9ac and H3K14ac. The blot experiment confirmed that the YDG domain or RING domain missing UHRF2 in the HEK293, LO2 and Hep G2 cells can abolish UHRF2's regulation of H3K9ac and H3K14ac. The deletion of these domains can significantly reduce the combination of UHRF2 and TIP60, but the combination of UHRF2 and HDAC1 has no significant effect on.4) Western blot experiments confirmed that the overexpression of UHRF2 in normal cells can up regulate the expression of TIP60 and the activity of enzymes, while UHRF2 in the tumor cells can inhibit TIP60 expression and enzyme activity. Sex has no significant regulation on the expression of HDAC1. And the regulation of UHRF2 on TIP60 will be abolished with the addition of the proteasome inhibitor MG132 and the half-life experiment confirms that the overexpression of UHRF2 in normal cells (HEK293 and LO2 cells) can prolong the half-life of TIP60, while the overexpression of UHRF2 in the tumor cells (Hep G2 cells) can accelerate the decline. The degradation rate of IP60, and the deletion of the UHRF2 protein RING domain can abolish this regulatory effect.6) in vivo ubiquitination experiments confirmed that the overexpression of UHRF2 in normal cells (HEK293 and LO2 cells) increased the ubiquitination of TIP60, while the overexpression in the tumor cells (Hep G2 cells) could inhibit TIP60 ubiquitination. The overexpression of TIP60 in HEK293, LO2 and Hep G2 cells can up regulate the expression of H3K9ac and H3K14ac, and the interference of TIP60 can inhibit the expression of H3K9ac and H3K14ac. The expression of AC and H3K14ac; when we used TIP60 inhibitor MG149 to treat the cells, no matter over expressed wild type UHRF2 or UHRF2 domain mutants could not continue to regulate.9 in H3K9ac and H3K14ac. The high expression of UHRF2 is associated with high expression of TIP60, H2AK5ac, H3K9ac and H3K14ac in the para cancerous tissues. Conclusion in normal cells (HEK293 and LO2 cells), UHRF2 prolongs the half-life of TIP60 and up regulates the protein expression and enzyme activity of TIP60, thus increasing H3K9ac and H3K14ac expression. The degradation rate of fast TIP60 inhibited the protein expression and enzyme activity of TIP60 and inhibited the expression of H3K9ac and H3K14ac.
【学位授予单位】:重庆医科大学
【学位级别】:硕士
【学位授予年份】:2017
【分类号】:R730.2
【参考文献】
相关期刊论文 前9条
1 彭城;李国亮;张红雨;阮一骏;;染色质三维结构重建及其生物学意义[J];中国科学:生命科学;2014年08期
2 周丹琳;胡斌;宣艳艳;段昌柱;;NIRF对HBV复制的影响及其对组蛋白H3乙酰化水平的修饰[J];中国细胞生物学学报;2013年03期
3 胡斌;周丹琳;宣艳艳;段昌柱;;NIRF在体内外可明显抑制乙型肝炎病毒抗原的表达[J];中国生物化学与分子生物学报;2012年11期
4 杨艳;金方敏;白露;王筱绘;段昌柱;;HBc蛋白与NIRF蛋白细胞外相互作用鉴定[J];生物学杂志;2012年01期
5 郑小梅;伍宁丰;;DNA甲基化作用的生物学功能[J];中国农业科技导报;2009年01期
6 王溪;朱卫国;;组蛋白甲基化酶及去甲基化酶的研究进展[J];癌症;2008年10期
7 PaolaGallinari;StefaniaDiMarco;PhillipJones;MichelePallaoro;ChristianSteinkühler;;HDACs,histone deacetylation and gene transcription: from molecular biology to cancer therapeutics[J];Cell Research;2007年03期
8 段昌柱;蒲淑萍;TSUTOMU MORI;HIDEO KOCHI;邱宗荫;;NIRF对P53蛋白泛素化作用的研究[J];生物化学与生物物理进展;2006年02期
9 黄百渠,曾庆华,毕晓辉,王玉红,李玉新;组蛋白和核小体在基因转录中的作用[J];科学通报;2000年19期
,本文编号:2141442
本文链接:https://www.wllwen.com/yixuelunwen/zlx/2141442.html