摘要
Soluble adenylyl cyclase (sAC) plays a critical role in male fertility in mammals by regulating sperm hyperactivation. We aimed to study the mechanism of sAC in this phenomenon and to explore potential target sites for male contraception. In this study, in vivo electroporation and rete testis microinjection-mediated short hairpin (sh)RNA plasmids were adopted to silence sAC gene expression in male rats. The results showed that high transfection efficiency (shRNA717, 49.0% and shRNA4205, 65.0%) was achieved by shRNA plasmids injected directly into the rete testis. When the sAC was downregulated, the cyclic adenosine mono-phosphate (cAMP) content and protein phosphorylation level of spermatozoa both declined with a significantly lower hyperactivation rate compared with negative controls. The highest transfection efficiency occurred at 15 d and was obviously time dependent. Bioinformatic and experimental results showed that sAC and tmAC both belong to the AC family and might have analogous functions. ShRNA717 and shRNA4205 were the best targets for the sAC gene, suggesting that they could be candidates for male contraception. Thus, it appears feasible to achieve male contraception by silencing the expression of sAC, affecting sperm hyper-activation via a cAMP-mediated signaling pathway.
Soluble adenylyl cyclase （sAC） plays a critical role in male fertility in mammals by regulating sperm hyperactivation. We aimed to study the mechanism of sAC in this phenomenon and to explore potential target sites for male contraception. In this study, in vivo electroporation and rete testis microinjection-mediated short hairpin （sh）RNA plasmids were adopted to silence sAC gene expression in male rats. The results showed that high transfection efficiency （shRNA717, 49.0% and shRNA4205, 65.0%） was achieved by shRNA plasmids injected directly into the rete testis. When the sAC was downregulated, the cyclic adenosine mono- phosphate （cAMP） content and protein phosphorylation level of spermatozoa both declined with a significantly lower hyperacti- vation rate compared with negative controls. The highest transfection efficiency occurred at 15 d and was obviously time depend- ent. Bioinformatic and experimental results showed that sAC and tmAC both belong to the AC family and might have analogous functions. ShRNA717 and shRNA4205 were the best targets for the sAC gene, suggesting that they could be candidates for male contraception. Thus, it appears feasible to achieve male contraception by silencing the expression of sAC, affecting sperm hyper- activation via a cAMP-mediated signaling pathway.
基金
supported by the National Natural Science Foundation of China (30471243)
National Supporting Projects for Science and Techniques (2011BAD28B02, 2012BAD12B10)
Provincial Project for Agricultural Techniques (SX2011-189, SXGC[2012]394)
