国际生殖健康/计划生育杂志

国际生殖健康/计划生育 ›› 2017, Vol. 36 ›› Issue (2): 97-102.

• 论著 • 上一篇    下一篇

SET蛋白对精母细胞株（GC-2 spd）增殖和组蛋白乙酰化的影响

崔毓桂1,张蓓1,朱倩2,许波群3,徐文丹1,杨晓玉4,高超1,高莉5,刘嘉茵6   

  1. 1. 南京医科大学第一附属医院
    2. 南京医科大学第一附属医院生殖医学中心
    3. 南京医科大学第二附属医院
    4. 江苏省人民医院生殖医学科
    5.
    6. 江苏省人民医院
  • 收稿日期:2017-01-20 修回日期:2017-02-10 出版日期:2017-03-15 发布日期:2017-03-15
  • 通讯作者: 崔毓桂 E-mail:cuiygnj@njmu.edu.cn

Up- Regulation of SET Protein in the Proliferation of GC-2 spd cells and Histone Acetylation

  • Received:2017-01-20 Revised:2017-02-10 Published:2017-03-15 Online:2017-03-15
  • Contact: Yugui Cui E-mail:cuiygnj@njmu.edu.cn

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摘要： 【摘要】目的：观察SET蛋白对GC-2spd精母细胞株增殖和组蛋白乙酰化的影响，探讨SET蛋白调节精子形成的作用。方法：体外培养GC-2spd细胞，通过精母细胞标志物乳酸脱氢酶C（LDHC）和睾丸特异激酶1（TESK1）鉴定。琼脂糖珠免疫共沉淀试剂盒检测SET蛋白与组蛋白H4的结合。分别转染空载腺病毒（AdH1-siRNA/NS，对照组）和SET干涉腺病毒（AdH1-siRNA/ SET），比较细胞增殖情况，荧光显微镜观察GC-2spd细胞中SET蛋白的表达；用Realtime RT-PCR和Western Blot检测mRNA以及蛋白的表达。结果：免疫荧光显示精母细胞株GC-2spd阳性表达LDHC和TESK1；SET蛋白表达于GC-2细胞的胞核和胞浆中，免疫共沉淀提示SET蛋白能与组蛋白H4结合。转染SET干涉腺病毒后，核内和胞浆中SET蛋白表达明显降低（**P＜0.01）。干涉组细胞增殖明显降低，组蛋白H4乙酰化水平（1.236±0.2549）明显增高（*P＜0.05）；但组蛋白乙酰化酶和去乙酰化酶mRNA表达水平无明显差异（P>0.05）。结论：精母细胞核内和胞浆中均表达SET蛋白；SET蛋白降调，则抑制GC-2spd细胞增殖。SET蛋白通过与组蛋白H4相互作用调节其乙酰化，参与调节精子形成。

关键词: SET蛋白, 精子形成, 精母细胞, 组蛋白, 组蛋白乙酰化

Abstract: Objective: To investigate the effects of SET protein on the regulation of spermiogenesis, we observe the proliferation and histone acetylation of spermatocyte cell line GC-2 spd. Methods: GC-2 spd cells were cultured in vitro and identified by the of spermatocytes’ marker LDHC(lactate dehydrogenase C) and TESK1(Testis-Specific Kinase 1). Immunoprecipitation Kit detect combination of SET and H4. The cells in control group were transfected with AdH1-siRNA/NS and experimental group were transfected with AdH1-siRNA/SET, then compare the proliferation between them. Immunoflourscence microscopy detect the and location of SET protein; Use Realtime RT-PCRand Western Blot to detect the of mRNA and protein. Results: GC-2 spd cells espress the spermatocyte marker LDHC and TESK1. SET protein is expressed in both cytoplasm and nucleus of GC-2 spg cells. Immunoprecipitation showed that SET can combine with H4. The of SET mRNA and protein transfected with AdH1-siRNA/SET was significantly decreased. Both of the cytoplasm and nucleusa SET decreased after infection by immunoflourscence microscopy; Proliferation was significantly inhibited and acetylation of H4 was increased after transfection (1.236±0.2549)( *P＜0.05); There was no significantly difference in mRNA of HATs and HDACs. Conclusions: SET protein was expressed in both cytoplasm and nucleus of GC-2 spd cells; Knockdown of SET protein inhibited proliferation; SET participated in the regulation of spermiogenesis by combining with H4 then regulated its acetylation level.

Key words: SET protein, spermiogenesis, spermatocyte, histone, histone acetylation