During the present funding period, we and others demonstrated that testicular POMC gene is expressed not only in Leydig cells but also in germ cells. the expression of this gene depends upon the interaction between these two cell types. The recent studies on inhibin have opened a new field for studying the regulation of reproductive physiology. Inhibin peptides synthesized by Sertoli cells were shown to modulate Leydig cell steroidogenesis in addition to their well-know function of regulating pituitary FSH secretion. Thus, inhibin is also considered to be an important regulator of spermatogenesis. In addition, POMC-derived peptides are important paracrine modulators of Sertoli cell function and inhibin production. These new observations suggest that the expression of inhibin genes in Sertoli cells and the expression POMC gene in Leydig and germ cells may be regulated by each other. We therefore propose to use inhibin and POMC as model systems to study the intratesticular regulation of gene expression in Sertoli, Leydig, and germ cells. The information on the structure of POMC gene and its regulation in the testis have been accumulated in the past years. However, the expressions of inhibin genes and their regulations in the Sertoli cells have not been well studied. Our first specific aim is to characterize the structure of mRNAs coding for inhibin subunits in rat testis. Secondly, we will investigate the in vitro differential regulation of inhibin subunit gene expression in rat Sertoli cell primary cultures by factors know to affect Sertoli cell functions, by paracrine regulators (POMC-derived peptides and Leydig cell proteins), or by cellular interactions. We will further investigate whether the above modulators affect the gene transcription or the mRNA stability. Using this information we will then investigate the intratesticular regulation of inhibin gene expression in animal systems under the influence of Leydig and/or germ cell functions or of POMC gene modulation, affected by pituitary hormones, androgens, or different stages of germ cells (Specific Aim A.3). finally, we will identify regulatory DNA elements, the promotor/regulatory regions and hormone-responsive elements, that are responsible for inhibin alpha-subunit gene transcription, and determine the molecular mechanism involved in the tissue-specific altered transcription of POMC gene in rat testis (Specific Aim A.4). The information we will obtain from these studies will facilitate our understanding of the intragonadal control of male reproduction.
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