1 Endocrinology Division, CSIR-Central Drug Research Institute, Lucknow, India.
Reprod Sci. 2017 May;24(5):738-752. doi: 10.1177/1933719116669057. Epub 2016 Sep 22.
The function of RHOG, a RAC1 activator, was explored in the ovary during ovarian follicular development and pathological conditions. With the help of immunoblotting and immunolocalization, we determined the expression and localization of RHOG in normal (estrous cycle) and polycystic ovaries using Sprague Dawley (SD) rat model. Employing polymerase chain reaction and flow cytometry, we analyzed the transcript and expression levels of downstream molecules of RHOG, DOCK1, and RAC1 in the polycystic ovarian syndrome (PCOS) ovary along with normal antral follicular theca and granulosa cells after dehydroepiandrosterone (DHEA) supplementation. The effect of RHOG knockdown on DOCK1, VAV, and RAC1 expression was evaluated in the human ovarian cells (SKOV3), theca cells, and granulosa cells from SD rats with the help of flow cytometry. Oocyte at secondary follicles along with stromal cells showed optimal expression of RHOG. Immunoblotting of RHOG revealed its maximum expression at diestrus and proestrus, which was downregulated at estrus stage. Mild immunostaining of RHOG was also present in the theca and granulosa cells of the secondary and antral follicles. Polycystic ovary exhibited weak immunostaining for RHOG and that was corroborated by immunoblotting-based investigations. RHOG effectors DOCK1 and ELMO1 were found reduced in the ovary in PCOS condition/DHEA. RHOG silencing reduced the expression of DOCK1 and RAC1 in the theca and granulosa cells from SD rat antral follicles and that was mirrored in the human ovarian cells. Collectively, RHOG can mediate signaling through downstream effectors DOCK1 and RAC1 during ovarian follicular development (theca and granulosa cells and oocyte), but DHEA downregulated them in the PCOS ovary.
RHOG 是 RAC1 的激活剂，其功能在卵巢卵泡发育和病理条件下得到了探索。我们使用 Sprague Dawley（SD）大鼠模型，通过免疫印迹和免疫定位法，确定了正常发情周期和多囊卵巢中的 RHOG 表达和定位。我们采用聚合酶链反应和流式细胞术，分析了多囊卵巢综合征（PCOS）卵巢中 RHOG 下游分子 DOCK1 和 RAC1 的转录和表达水平，以及脱氢表雄酮（DHEA）补充后正常窦前卵泡的膜和颗粒细胞。我们还使用流式细胞术评估了 RHOG 敲低对人卵巢细胞（SKOV3）、SD 大鼠膜细胞和颗粒细胞中 DOCK1、VAV 和 RAC1 表达的影响。次级卵泡的卵母细胞和基质细胞显示出 RHOG 的最佳表达。RHOG 的免疫印迹显示其在动情前期和动情前期表达最高，在动情期下调。次级和窦前卵泡的膜和颗粒细胞中也存在 RHOG 的轻度免疫染色。多囊卵巢的 RHOG 免疫染色较弱，免疫印迹研究也证实了这一点。在 PCOS 条件/DHEA 下，卵巢中的 RHOG 效应物 DOCK1 和 ELMO1 减少。RHOG 沉默降低了 SD 大鼠窦前卵泡的膜和颗粒细胞中 DOCK1 和 RAC1 的表达，这在人卵巢细胞中也得到了反映。总之，RHOG 可以在卵巢卵泡发育过程中（膜和颗粒细胞和卵母细胞）通过下游效应物 DOCK1 和 RAC1 介导信号转导，但 DHEA 在 PCOS 卵巢中下调了它们。
