There is considerable evidence that in normal, mature testes, developing male gametes interact with surrounding Sertoli cells and that these interactions are essential for spermatogenesis. In rats, one result of these interactions is the expression by Sertoli cells of specific genes at particular stages of the cycle of the seminiferous epithelium. It is proposed that these genes' products regulate the development of germ cells present in those stages or other important testis functions. To investigate stage-specific gene expression by Sertoli cells, we have focused on Cyclic Protein-2, the pro-enzyme form of the cysteine protease, Cathepsin L. This protein is secreted by Sertoli cells within stage V-VIII tubules. Recent and published results propose three different in vivo functions for this protein in rat testes: [1] Degradation of residual bodies that have been engulfed by Sertoli cells, preventing the release of the germ cell-specific antigens into the general physiological environment. [2] Cleavage of adhesive functions between compacted spermatids and Sertoli cells in stage VI tubules, facilitating the movement of spermatids towards the lumen of the tubule. [3] Stimulation of testosterone secretion by Leydig cells. Experiments in the first specific aim test, in vivo, these proposed functions. The second specific aim examines whether the stage-specific expression of CP-2/Cathepsin L mRNA by Sertoli cells also occurs in humans, rabbits, boar and bulls. Such expression in other species would indicate that our proposed studies in rodents are applicable to those species and in the future might provide insight into causes of human idiopathic fertility. This application also proposes to define the molecular mechanisms which regulate the stage- specific transcription of the gene encoding CP-2/Cathepsin L. We have previously demonstrated that with progression of the stages of the cycle, germ cells sequentially trigger the repression, derepression (or loss of repression), stimulation and re-repression of transcription of this gene. Such regulation by germ cells may be mediated by an enhancer and by repressor and derepressor regions that we have identified in the CP- 2/Cathepsin L gene. We, therefore, propose to identify the cis-acting elements in these regions which bind specific transacting factors in a stage-specific manner. We also propose to determine, by studying mice carrying specific CP-2/Cathepsin L-beta-galactosidase transgenes, whether these regions are responsible for stage-specific gene transcription. Results from these studies will begin to elucidate the molecular basis for a stage-specific germ cell-Sertoli cell interaction.
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