減數分裂後男性精細胞的核膜完整性維持需SEPT12/SPAG4/LAMINB1複合體參與
葉忠信1,2、汪雅雲3、吳盈羽3、江漢聲2,3、林盈宏2,3
1新光醫院 泌尿科 2輔仁大學 醫學院 3輔仁大學 基礎醫學研究所
SEPT12/SPAG4/LAMINB1 complexes are required for maintaining the integrity of the nuclear envelope in postmeiotic male germ cells
Chung-Hsin Yeh1,2、Ya-Yun Wan、Ying-Yu Wu、Han-Sun Chiang2,3、Ying-Hung Lin2,3
1Division of Urology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan
2School of Medicine, Fu-Jen Catholic University, New Taipei City, Taiwan
3Graduate Institute of Basic Medicine, Fu-Jen Catholic University
 
Purpose: Our previous studies based on a knockout mice model indicated that SEPT12 proteins are critical for the terminal morphological formation of sperm. SEPT12 mutations in men result in teratozospermia and oligozospermia. In addition, the spermatozoa exhibit morphological defects of the head and tail, premature chromosomal condensation, and nuclear damage. However, the molecular functions of SEPT12 during spermatogenesis remain unclear. To determine the molecular functions of SEPT12, we designed and carried out the present study.
  
Methods and Materials: We applied a yeast 2-hybrid system to identify SEPT12 interactors. The molecule-biological skills included the following items: Yeast 2-hybrid screening and β-galactosidase assay, Reverse transcription-PCR (RT-PCR), Preparation of human testicular spermatogenic cells and spermatozoa, Immunofluorescence assay, Cloning, Transfection, and Coimmunoprecipitation assay.
 
Results: Seven proteins that interact with SEPT12 were identified: SEPT family proteins (SEPT4 and SEPT6), nuclear or nuclear membrane proteins (PRM2, SPAG4, and NDC1), and sperm-related structural proteins (PCM1 and OBSL1). SPAG4; also known as SUN4 belongs to the SUN family of proteins and acts as a linker protein between nucleoskeleton and cytoskeleton proteins and localizes in the nuclear membrane. We determined that SEPT12 interacts with SPAG4 in a male germ cell line through coimmunoprecipitation. During human spermiogenesis, SEPT12 is colocalized with SPAG4 near the nuclear periphery in round spermatids and in the centrosome region in elongating spermatids. Furthermore, we observed that SEPT12/SPAG4/LAMINB1 formed complexes and were coexpressed in the nuclear periphery of round spermatids. In addition, mutated SEPT12 screened from an infertile man affected the integration of these nuclear envelope complexes through coimmunoprecipitation.
 
Conclusion: SEPT12 interactors are active at various stages during human spermiogenesis. One of the interactors, SPAG4, is critical for the formation of the nuclear membrane of sperm and DNA integration in the differentiation of male germ cells.
 
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    台灣泌尿科醫學會
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    2015-06-04 23:11:00
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    2015-06-04 23:13:46
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