This project aims to determine the molecular genetic bases of the various forms of cogenital adrenal hyperplasia (CAH). CAH results from an inherited metabolic defect in cortisol biosynthesis. It may cause abnormalities in sexual differentiation, somatic growth, and salt metabolism. While any of the five metabolic steps in the pathway from cholesterol to cortisol may be affected, 95% of cases involve the 21-hydroxylation of 17-hydroxyprogesterone tp 11-deoxycortisol (21-hydroxylase deficiency), and in most of the remainder, 11B-hydroxylation is defective (11B-hydroxylase deficiency.) Both of these hydroxylation steps are mediated by specific adrenal cytochromes P-450 (respectively termed P-450 and P-450). The project therefore concentrates on the manner in which mutations in genes for these P-450 enzymes result in CAH. Previously, cDNA clones have been isolated which encode bovine and human P-450 as well as cosmid clones containing both normal P-450 genomic genes. One of these two genes may be a pseudogene, while many of the most severe cases of 21-OHase deficiency results from a deletion of the other, normally active gene. Milder forms of 21-OHase deficiency probably result from various point mutations. These genes are located in the HLA major histocompatibility complex, and different mutations appear to be associated with characteristic HLA antigens. Such mutations will be defined by producing genomic libraries using DNA from patients with 21-OHase deficiency who are homozygous for characeristic HLA antigens, and isolating and sequencing mutant genes. Previously isolated bovine cDNA clones encoding P-450 and P-450 (cholesterol side-chain cleavage enzyme) will be used to probe a human fetal adrenal cDNA library to identify the corresponding human genes, which will then be used to probe a genomic library to identify the genomic genes. The DNA sequences of these genes will be determined, and the chromosomal locations of these genes determined by in situ hybridization or hybridization to somatic cell hybrids. These studies should provide reagents which will be useful in haplotype identification for purposes of prenatal diagnosis of these defects.
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