Follicle-stimulating Hormone Regulates Pro-apoptotic Protein Bcl-2-interacting Mediator of Cell Death-Extra Long (BimEL)-induced Porcine Granulosa Cell Apoptosis*
Follicle-stimulating Hormone Regulates Pro-apoptotic Protein Bcl-2-interacting Mediator of Cell Death-Extra Long (BimEL)-induced Porcine Granulosa Cell Apoptosis*
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DOI：
10.1074/jbc.m111.293274
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发表时间：
2012-01
期刊：
影响因子：
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通讯作者：
Xian-long Wang;Yi Wu;L. Tan;Zhen Tian;Jing-hao Liu;De-Sheng Zhu;S. Zeng
中科院分区：
文献类型：
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作者：
Xian-long Wang;Yi Wu;L. Tan;Zhen Tian;Jing-hao Liu;De-Sheng Zhu;S. Zeng
背景：在哺乳动物卵巢中，99%的卵泡是通过颗粒细胞凋亡引起的卵泡闭锁而被摘除的。结果：BimEL可诱导猪颗粒细胞凋亡，其表达受FSH通过PI3K/Akt/FOXO3a途径调节。结论：促凋亡蛋白BimEL参与了猪颗粒细胞的凋亡过程。意义：这为卵泡闭锁的分子机制提供了新的见解。促凋亡蛋白Bim(B细胞淋巴瘤-2(Bcl2)-细胞死亡的相互作用调节剂)最近被发现，并被证明在几种刺激下促进细胞死亡。在本报告中，我们研究了BIM在猪卵泡闭锁中的作用。首先，从猪卵巢组织中克隆并鉴定了Bim基因。猪Bim有三种不同的剪接变异体(Bim-Extra Long、Bim-Long和Bim-Short)，均含有共识的Bcl2同源3结构域。然后我们发现Bim-Extra Long(BimEL)蛋白是Bim最丰富的异构体，它强烈表达并与来自闭锁卵泡的凋亡(TUNEL阳性)颗粒细胞共存。此外，BimEL的过表达还引发了颗粒细胞的凋亡。在基础条件下的原代颗粒细胞培养中，我们观察到BimEL的表达被卵泡刺激素(FSH)抑制。通过使用PI3K抑制剂LY294002和Akt siRNA的转染，阐明了PI3K/Akt通路在调节抑制中的作用。Bim转录激活因子Forkhead Box O3a(FOXO3a)在Ser-253处被磷酸化，FSH刺激后失活。此外，FSH还取消了FOXO3a核积累，以回应LY294002。染色质免疫沉淀实验证明FOXO3a直接结合并激活了bim启动子。综上所述，BimEL可诱导卵泡闭锁过程中猪颗粒细胞的凋亡，其表达受FSH通过PI3K/Akt/FOXO3a途径调节。
Background: In the mammalian ovary, 99% of follicles are removed through follicular atresia caused by granulosa cell apoptosis. Results: BimEL can induce porcine granulosa cell apoptosis, and its expression is regulated by FSH via the PI3K/Akt/FoxO3a pathway. Conclusion: The pro-apoptotic protein BimEL is involved in porcine granulosa cell apoptosis. Significance: This provides novel insights into the molecular mechanisms underlying follicular atresia. The pro-apoptotic protein Bim (B-cell lymphoma-2 (Bcl-2)-interacting modulator of cell death) has recently been identified and shown to promote cell death in response to several stimuli. In this report, we investigated the role of Bim in porcine follicular atresia. Initially, Bim cDNA was cloned and characterized from porcine ovarian tissue. Porcine Bim had three alternative splicing variants (Bim-extra long, Bim-long, and Bim-short), all containing the consensus Bcl-2 homology 3 domain. We then found the Bim-extra long (BimEL) protein, the most abundant isoform of Bim, was strongly expressed and co-localized with apoptotic (TUNEL-positive) granulosa cells from porcine atretic follicles. Furthermore, overexpression of BimEL triggered apoptosis in granulosa cells. In primary granulosa cell cultures under basal conditions, we observed that BimEL expression was dampened by treatment with follicle-stimulating hormone (FSH). The role of the PI3K/Akt pathway in the regulation of repression was clarified by the use of the PI3K inhibitor, LY294002, and by transfection with Akt siRNA. Forkhead Box Protein O3a (FoxO3a), a well defined transcriptional activator of Bim, was phosphorylated at Ser-253 and inactivated after FSH stimulation. Also, FSH abolished FoxO3a nuclear accumulation in response to LY294002. Finally, chromatin immunoprecipitation assays demonstrated that FoxO3a directly bound and activated the bim promoter. Taken together, we conclude that BimEL induces porcine granulosa cell apoptosis during follicular atresia, and its expression is regulated by FSH via the PI3K/Akt/FoxO3a pathway.