Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways.
Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways.
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Hsp90 通过 Erk1/2 和 p38 MAPK 信号通路调节人类精子获能。
DOI：
10.1186/s12958-021-00723-2
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发表时间：
2021-03-04
期刊：
影响因子：
--
通讯作者：
Ni Y
中科院分区：
文献类型：
--
作者：
Sun P;Wang Y;Gao T;Li K;Zheng D;Liu A;Ni Y
热休克蛋白90（Heat shock protein 90，Hsp 90）是一种高丰度的真核生物分子伴侣，在客户蛋白的成熟、折叠和降解以及信号转导等过程中发挥重要作用。以前，我们发现热休克蛋白90和它的辅助分子伴侣细胞分裂周期蛋白37（Cdc 37）在人类精子中表达。已知Hsp 90通过未知的潜在机制参与人类精子获能。由于Cdc 37是Hsp 90的一种激酶特异性辅伴侣，因此Hsp 90可能通过其他激酶调节人精子获能。据报道，两种主要的丝裂原活化蛋白激酶（MAPK），细胞外信号调节激酶1/2（Erk 1/2）和p38，在人类精子中表达的位置与Hsp 90和Cdc 37相同。磷酸化的Erk 1/2可促进精子的超活化运动和顶体反应，而磷酸化的p38可抑制精子的运动。因此，在本研究中，我们探讨是否热休克蛋白90调节人类精子获能通过Erk 1/2和p38 MAPK信号通路。在获能过程中，用Hsp 90特异性抑制剂17-烯丙基氨基-17-去甲氧基格尔德霉素（17-AAG）处理人精子。采用计算机辅助精子分析仪（CASA）检测精子活动率和超活化率。采用异硫氰酸荧光素标记的豌豆凝集素（PSA-FITC）染色法分析精子顶体反应。采用免疫共沉淀法（Co-IP）检测Hsp 90、Cdc 37、Erk 1/2和p38之间的相互作用。Western blotting分析用于评估蛋白表达和磷酸化水平。17-AAG能抑制人精子的超活化和顶体反应，提示Hsp 90参与人精子获能。此外，Co-IP实验表明，17-AAG降低了Hsp 90和Cdc 37之间的相互作用，导致Erk 1/2从Hsp 90-Cdc 37蛋白复合物中解离。蛋白质印迹分析显示，Erk 1/2及其磷酸化形式的水平随后下降。Hsp 90-Cdc 37复合物的减少也影响了Hsp 90与p38的相互作用。然而，p38从Hsp 90蛋白复合物中解离，并通过自磷酸化被激活。综上所述，我们的研究结果表明，热休克蛋白90参与人类精子的超激活和顶体反应。特别地，Hsp 90及其共伴侣Cdc 37与Erk 1/2和p38形成蛋白复合物以调节它们的激酶活性。这些结果表明，热休克蛋白90调节人类精子获能通过Erk 1/2和p38 MAPK信号通路。
Heat shock protein 90 (Hsp90) is a highly abundant eukaryotic molecular chaperone that plays important roles in client protein maturation, protein folding and degradation, and signal transduction. Previously, we found that both Hsp90 and its co-chaperone cell division cycle protein 37 (Cdc37) were expressed in human sperm. Hsp90 is known to be involved in human sperm capacitation via unknown underlying mechanism(s). As Cdc37 was a kinase-specific co-chaperone of Hsp90, Hsp90 may regulate human sperm capacitation via other kinases. It has been reported that two major mitogen-activated protein kinases (MAPKs), extracellular signal-regulated kinase 1/2 (Erk1/2) and p38, are expressed in human sperm in the same locations as Hsp90 and Cdc37. Phosphorylated Erk1/2 has been shown to promote sperm hyperactivated motility and acrosome reaction, while phosphorylated p38 inhibits sperm motility. Therefore, in this study we explored whether Hsp90 modulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways. Human sperm was treated with the Hsp90-specific inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) during capacitation. Computer-assisted sperm analyzer (CASA) was used to detect sperm motility and hyperactivation. The sperm acrosome reaction was analyzed by using fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (PSA-FITC) staining. The interactions between Hsp90, Cdc37, Erk1/2 and p38 were assessed using co-immunoprecipitation (Co-IP) experiments. Western blotting analysis was used to evaluate the levels of protein expression and phosphorylation. Human sperm hyperactivation and acrosome reaction were inhibited by 17-AAG, suggesting that Hsp90 is involved in human sperm capacitation. In addition, Co-IP experiments revealed that 17-AAG reduced the interaction between Hsp90 and Cdc37, leading to the dissociation of Erk1/2 from the Hsp90-Cdc37 protein complex. Western blotting analysis revealed that levels of Erk1/2 and its phosphorylated form were subsequently decreased. Decreasing of Hsp90-Cdc37 complex also affected the interaction between Hsp90 and p38. Nevertheless, p38 dissociated from the Hsp90 protein complex and was activated by autophosphorylation. Taken together, our findings indicate that Hsp90 is involved in human sperm hyperactivation and acrosome reaction. In particular, Hsp90 and its co-chaperone Cdc37 form a protein complex with Erk1/2 and p38 to regulate their kinase activity. These results suggest that Hsp90 regulates human sperm capacitation via the Erk1/2 and p38 MAPK signaling pathways.
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