Regulation of testis-specific poly(A) polymerase by protein-protein interaction

Abstract

Mammalian spermatogenesis shows a strict control of many specific molecular and cellular events. After two-step meiosis, DNA-protamine-interactions in elongating spermatids result in chromatin condensation causing cessation of transcription at a time when many proteins need to be synthesized and assembled for the complete morphological change. In this step, spermatogenesis occurs with translational control of mRNA which transcribes in early step. It is well-known that cytoplasmic polyadenylation is involved a translational activation of germ cell differentiation. Testis-specific poly(A) polymerase (TPAP), involved in adding poly(A) at the 3’ end of mRNA in haploid germ cells, is highly expressed in round spermatids. In TPAP-deficient mice, expression of haploid-specific genes required for morphogenesis of germ cells is impaired, and poly(A) tails of specific transcription factor mRNAs of round spermatids are not elongated completely. Consequently, these mice are infertile due to spermatogenesis arrest. In spermatogenesis, CPEB-dependent cytoplasmic polyadenylation is related to the formation of early spermatid and CPEB-deficient mice show a different aspect to TPAP-deficient mice. Mice overexpressing TPAP display normal spermatogenesis and fertility, and the mRNA sizes of the transcription factors are unaltered, suggestive of limiting regulatory factors that may act via interactions with TPAP. However, the regulation-mechanism of TPAP is unrevealed. In this thesis, regulatory proteins that would be involved in the regulation of TPAP functions via protein-protein interactions were screened using a yeast two-hybrid assay. Two proteins, germ-cell specific gene 1 protein (GSG1) and testis-specific adenine deaminase (ADAt) were identified as TPAP interaction partner proteins. Although the functions of GSG1 are not known yet, its expression coincides during mouse development. Coimmunoprecipitation assays revealed phosphorylation-independent interactions between TPAP ...