University of Texas Southwestern Medical Center
The focus of my research has been on mechanisms of steroid action with emphases on: 1) structure-activity relationships of ligand-steroid receptor interactions, and 2) steroid metabolism. My early work involved interactions of the estrogen receptor with several non-steroidal antiestrogens (triaryl-ethylene and -ethane; TAEs) in order to define the ligand binding site on this receptor. These studies showed that these ligands bound to the same site on the estrogen receptor as estradiol-17b. The TAEs, however, have an "accessory" binding site which is responsible for anti-estrogenic activity. Data on binding of ligands to the accessory site could improve anti-estrogen activity for use as anti-tumor agents or fertility regulators. I carried out additional studies demonstrating that estrogen-receptor complexes needed to be constantly present at nuclear acceptor sites in order to maintain the transcriptional activity of estrogen. Specificity of steroid action in target tissues may be increased by biochemically modifying the steroid. For example, in male genital skin, testosterone is converted by 5a-reductase to dihydrotestosterone which is a more potent androgen than testosterone itself. Some analogs of testosterone, however, are maximally active with no further metabolism. One such synthetic analog of testosterone is 7a- methyl-19nor-testosterone. Using rat liver and a prostate microsomal fraction, I showed that this steroid was not metabolized by 5a-reductase and thus it might be useful for treatment of 5-alpha reductase deficient patients. It is currently being evaluated for its androgenic potency in primates. My work over the past decade has concentrated on the conversion of cortisol (the main glucocorticoid in humans) to its inactive metabolite, cortisone. This conversion is mediated by 11b- hydroxysteroid dehydrogenase (11-HSD). Specific defects in this enzyme lead to high levels of cortisol in the kidney and spurious activation of mineralocorticoid receptors by cortisol, an inherited form of hypertension termed apparent mineralocorticoid excess (AME). I cloned two isoforms of 11-HSD (types 1 and 2) and expressed them using a number of different systems including Xenopus oocytes, vaccinia virus and transfection of expression plasmids in mammalian cells. I showed that the 11-HSD1 isozyme catalyzes both oxidation of cortisol (and corticosterone) and reduction of cortisone (and 11-dehydrocorticosterone), requires NADP+ or NADPH as a cofactor, and requires glycosylation for full activity. In contrast, 11-HSD2 catalyzes only oxidation, requires NAD+ as a cofactor, has a much higher affinity for steroids, and apparently doesn’t require glycosylation. Together with my colleagues, I showed that mutations of the 11-HSD2 gene cause AME. My current focus is to determine if there is an association between polymorphisms in the 11-HSD2 gene and essential hypertension or salt sensitivity and to examine the transcriptional regulation of this gene in placenta and kidney. Our most recent work relates to congenital generalized lipodystrophy (CGL), an autosomal recessive disorder characterized by extreme lack of body fat since birth, severe insulin resistance, hypertriglyceridemia, hepatic steatosis and early onset of diabetes. Through positional cloning, we identified disease=causing mutations in the AGPAT2 gene located on chromosome 9q34, encoding 1-acylglycerol-3-phosphate-O-acyltransferase 2, in affected subjects from 26 of the 42 pedigrees of various ethnicities. The affected individuals were either homozygous or compound heterozygous for various mutations including, deletions, nonsense, missense, splice-site and those in the 3’UTR. The AGPAT2 catalyzes the acylation of the lysophosphatidic acid at the sn-2 position to form phosphatidic acid, a key intermediate in the biosynthesis of triacylglycerol (TG) and glycerophospholipids, which are involved in signal transduction.
Functional aspects of acyltransferases Molecular aspects of lipodystrophy Molecular aspects of premature aging