Interaction of the mTERT telomerase catalytic subunit with the c-Abl tyrosine kinase in mouse granulosa cells

<b>Context:</b> Oocyte and granulosa cells (GCs) have bidirectional communication and GCs play an important role in folliculogenesis and proliferation of GCs is very important for the development of ovulatory follicle. DNA double-strand breaks activate c-Abl protein tyrosine kinase and c-Abl has a functional role in repairement of DNA and control of telomere. <b>Objective:</b> In this study, we hypothesized that c-Abl has a regulative role on mTERT in mouse ovarian granulosa cells (GCs) and we aimed to detect c-Abl and mTERT interaction in mouse primary culture of GCs. <b>Materials and methods:</b> Mouse ovarian granulosa cell were cultured and siRNA-mediated knockdown approach was used to knockdown c-Abl expression. <b>Results:</b> We showed c-Abl and mTERT immunolocalization <i>in vivo</i> and <i>in vitro</i> mouse GCs. c-Abl and mTERT were constitutively expressed in mouse granulosa cells and c-Abl presented more intense expression in granulosa cells than mTERT expression. The interaction of the c-Abl-mTERT is supported by the exhibition that c-Abl siRNA knockdown cells show decreased mTERT expression. We also present an interaction between c-Abl and mTERT by immunoprecipitation. In addition, our results indicated that the down-regulation of c-Abl was also accompanied by reduced expression of proliferating cell nuclear antigen (PCNA) in GCs. <b>Conclusions:</b> We suggest that mTERT may associate with the c-Abl in mouse GCs and the interactions between c-Abl and mTERT suggest a role for c-Abl in the regulation of telomerase function and proliferation in mouse granulosa cells.

**背景：** 卵母细胞与颗粒细胞（GCs）之间存在双向交流，颗粒细胞在卵泡发生过程中发挥关键作用，而颗粒细胞的增殖对于排卵卵泡的发育至关重要。DNA双链断裂可激活c-Abl蛋白酪氨酸激酶，c-Abl在DNA修复及端粒调控中具有功能性作用。
**目的：** 本研究假设c-Abl对小鼠卵巢颗粒细胞（GCs）中的小鼠端粒酶逆转录酶（mTERT）具有调控作用，并旨在探究小鼠原代培养颗粒细胞中c-Abl与mTERT的相互作用。
**材料与方法：** 体外培养小鼠卵巢颗粒细胞，并采用siRNA介导的基因敲低技术敲低c-Abl的表达。
**结果：** 本研究明确了c-Abl与mTERT在小鼠颗粒细胞体内及体外的免疫定位情况。二者在小鼠颗粒细胞中均呈组成型表达，且c-Abl的表达强度高于mTERT。经c-Abl siRNA敲低的细胞中mTERT表达水平显著下调，这一结果为c-Abl与mTERT的相互作用提供了支持依据；此外，本研究还通过免疫共沉淀实验验证了c-Abl与mTERT的相互作用。进一步研究发现，c-Abl表达下调同时伴随颗粒细胞中增殖细胞核抗原（PCNA）表达水平的降低。
**结论：** 本研究表明，mTERT可与小鼠颗粒细胞中的c-Abl发生相互作用；c-Abl与mTERT的相互作用提示，c-Abl可调控小鼠颗粒细胞中的端粒酶功能及细胞增殖过程。