MGCRABGAP經由調控Rap1表現影響精子成型
柯智群1,2、林盈宏3
恩主公醫院 泌尿科1；輔仁大學 食品營養博士學位學程2、生醫藥學研究所3
Modulation of Rap1 by MGCRABGAP regulate mammalian spermiogenesis
Chih-Chun Ke1, 2, Ying-Hung Lin3
1Department of Urology, En Chu Kong Hospital, New Taipei City; 2Ph.D. Program in Nutrition and Food Sciences, Fu-Jen University, New Taipei City, Taiwan; 3Graduate Institute of Biomedical and Pharmaceutical Science, Fu-Jen Catholic University, New Taipei City, Taiwan
Purpose: To investigate whether MGCRABGAP regulate Rap1 during mammalian spermiogenesis, and the expression pattern of Rap1 in different developmental stages.
Materials and Methods: To identify proteins that interact with MGCRABGAP, co-immunoprecipitation (co-IP) and nano liquid chromatography-mass spectrometry/mass spectrometry (nano LC-MS/MS) was performed in NT2D1 lysates. By using active Rap1 pull-down and detection assay, activity of Rap1 in wild-type cells and pFLAG-MGCRABGAP transfected NT2D1 cells was compared with or without 8-pCPT-2’-O-Me-cAMP stimulation. The expression patterns and localization of Rap1 during different stages of spermatogenesis was evaluated with immunofluorescence detection on murine testicular sections.
Results: Rap1 was identified as one of MGCRABGAP interactors, the binding ability between Rap1 and MGCRABGAP was further confirmed via co-IP. Overexpression of MGCRABGAP downregulated Rap1 activity under 8-pCPT stimulation. Immunofluorescence detection demonstrated Rap1 express exclusively in elongating/elongated spermatids of stage VI~VIII.
Conclusions: We previously identified MGCRABGAP as a male germ cell-specific protein expressed in elongating and elongated spermatids. In the present study, our findings indicate that the regulatory role of MGCRABGAP in spermiogenesis assembly depend on Rap1 modulation.
Key words: Male Germ Cells Rab GTPase-Activating Protein (MGCRABGAP), Ras-related protein 1 (Rap1), spermiogenesis