Strategies for the isolation of genes expressed meiotically during mouse spermatogenesis

The process of gametogenesis is of paramount importance for reproductive success in mammals Gamete production includes a specialized form of cell division termed meiosis whereby chromosomally diploid cells become haploid. Although meiosis is genetically essential for the formation of a normal gamete, very little is known of the molecular and cellular events constituting this phase of gametogenesis. An initial and reasonable approach to elucidating factors related to the meiotic process is to isolate genes expressed during meiosis. An overview of the literature, as it is related to this work, is presented in Part I of this dissertation. Part II of this dissertation outlines the construction of spermatogenic cell-specific cDNA libraries in order to establish a system for the molecular analysis of meiosis in male mammals. Cytological and molecular characterization of these libraries are also described. These libraries represent an important tool for the isolation of genes that are expressed meiotically. Part III of this dissertation describes the use of the meiotic cDNA libraries in a novel screen for meiotically-enriched cDNA clones. This screen utilizes RNA from meiotically-arrested male-sterile mutant mice for the successful identification of spermatocyte-enriched cDNA clones. The isolation of these clones serves as a foundation for further investigations into the molecular basis of meiosis. Part IV of this dissertation describes the use of homology screening by PGR to identify a meiotically-expressed cDNA. The isolation and characterization of a novel meiotically-expressed cDNA is outlined in this section. This cDNA is expressed most abundantly in pachytene spermatocytes and in the brain and thymus of mice and contains a putative leucine zipper motif. Additionally, although a molecular analysis of meiosis will potentially reveal new information, cytological approaches for understanding meiosis should lead to the discovery of different information concerning this process. In this regard. Part V reports the development of a spermatocyte in vitro culture system for the analysis of meiosis and interesting observations that have come from in vitro manipulation of spermatocytes. Finally, Part VI serves to summarize the various conclusions and significance of this work and postulates future directions for continued investigation into the molecular basis of meiosis.