Sun Wen-Juan, Zhu Ming, Wang Yuan-Li, Li Qing, Yang Hong-Dan, Duan Ze-Lin, He Lin, Wang Qun
School of Life Science, East China Normal University, Shanghai, China.
Mar Biotechnol (NY). 2015 Jun;17(3):305-16. doi: 10.1007/s10126-015-9619-y. Epub 2015 Feb 8.
Mitogen-activated protein kinases (MAPKs), also termed extracellular signal-regulated kinases (ERKs), are cytoplasmic and nuclear serine/threonine kinases involved in signal transduction of several extracellular effectors. In mammals, ERKs participate in the regulation of spermatogenesis, mature spermatozoa motility, hyperactivation, and the acrosome reaction. To investigate ERK functions in Eriocheir sinensis reproduction, we successfully cloned the full-length ERK from the testis of E. sinensis (ES-ERK). The 1098-nucleotide open reading frame encodes a 365-amino-acid protein with a predicted molecular weight of 42 kDa. Expressions of ES-ERK in different tissues and testis development stages were detected by the quantitative RT-PCR and Western blotting. ES-ERK is expressed relatively highly in the testis. The expression of ES-ERK protein gradually increased in the spermatid stage, reaching a peak in sperm stage. Western blotting showed a similar expression pattern for the total ES-ERK protein, but phospho-ERK (p-ERK) showed the higher expression in spermatid than sperm stage. We also used trypan blue and hematoxylin and eosin staining to identify structural changes in E. sinensis spermatozoa during the process of acrosome reaction (AR). After stimulating the process of AR, the ES-ERK has translocated from the nucleus to the acrosomal tubule. This result suggested that the ERK MAPK might be involved in the regulation of the E. sinensis acrosome reaction.
丝裂原活化蛋白激酶(MAPKs),也被称为细胞外信号调节激酶(ERKs),是细胞质和细胞核中的丝氨酸/苏氨酸激酶,参与多种细胞外效应器的信号转导。在哺乳动物中,ERKs参与精子发生、成熟精子活力、超活化以及顶体反应的调节。为了研究中华绒螯蟹繁殖过程中ERK的功能,我们成功地从中华绒螯蟹的睾丸中克隆了ERK的全长基因(ES-ERK)。1098个核苷酸的开放阅读框编码一个365个氨基酸的蛋白质,预测分子量为42 kDa。通过定量RT-PCR和蛋白质免疫印迹法检测ES-ERK在不同组织和睾丸发育阶段的表达。ES-ERK在睾丸中表达相对较高。ES-ERK蛋白的表达在精子细胞阶段逐渐增加,在精子阶段达到峰值。蛋白质免疫印迹法显示总ES-ERK蛋白的表达模式相似,但磷酸化ERK(p-ERK)在精子细胞中的表达高于精子阶段。我们还使用台盼蓝和苏木精-伊红染色来鉴定中华绒螯蟹精子在顶体反应(AR)过程中的结构变化。在刺激AR过程后,ES-ERK已从细胞核转移到顶体小管。这一结果表明ERK MAPK可能参与中华绒螯蟹顶体反应的调节。