The long term objective of this project it to understand what genes influence steroid hormone synthesis, and to understand how they work. Steroid hormones are ubiquitous physiologic regulators: mineralcorticoids regulate salt and water metabolism and blood pressure; glucocorticoids regulate carbohydrate metabolism and other functions; sex steroids regulate numerous aspects of reproduction. These steroids are made by sequential conversions of cholesterol to a series of steroid hormones; most of these conversions are made by four specific P450 enzymes: P450scc converts cholesterol to pregnenolone; P450c17 mediates both 17, 20 lyase activity and 17 hydroxylase activity; P450c21 mediates the 21-hydroxylation of both glucocorticoids and mineralcorticoids; P450c11 mediates 11 hydroxylase, 18 hydroxylase, and 18 methyloxidase activities. The first phase of this project, funded by this grant since 1987, was to obtain cDNA and gene clones for the steroidogenic P450 enzymes, to examine their regulation, and to study their mutations causing congenital adrenal hyperplasia (CAH). This work has been highly successful to date. In the course of studying CAH, we have discovered two previously unknown genes lying on the opposite strand of DNA from the P450c21 genes. The gene overlapping the P450c21A pseudogene is termed XA and that overlapping the functional P450c21B gene is termed XB. We have also discovered a third gene termed Y, lying within XB, on the same strand as P450c21B and 3' to it. Sequencing studies indicate that XA and XB probably encode extracellular matrix proteins related to tenascin; the function of Y is unknown. XA and Y are expressed solely in the adrenal, and hence may be involved in steroidogenesis; XB is expressed in virtually all fetal tissues suggesting a more general role in development. The present application for competing renewal focuses on establishing the structure and function Genes XA, XB and Y and on continuing our studies of the mutations in the steroidogenic P450 genes causing various forms of congenital adrenal hyperplasia. To do this we propose to: 1) Complete the structural characterization of the XA, XB and Y loci; 2) characterize the mRNA and protein products encoded by these genes 3) determine the tissue distribution of their expression 4) characterize the biologic activities of the products encoded by these genes 5) continue our studies on the molecular basis of the congenital adrenal hyperplasia. Successful completion of this work will greatly expand our understanding of the roles of these newly discovered genes in adrenal function.
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