Androgen receptors (AR) bind testosterone and dihydrotestosterone to form transacting transcriptional regulatory complexes necessary for induction of specific genes during normal male sex differentiation and development and in maintenance of normal male physiology. X-linked inheritance of the clinical syndromes of androgen insensitivity (AIS) provide a human model for understanding the mechanisms of androgen action and the molecular lesions that lead to abnormalities of sex differentiation. Recent isolation and cloning of the human AR cDNA and generation of antibodies against AR peptide provide the molecular tools for the proposed studies. The goal of this research proposal is to study the molecular genetics of human AIS as a means to understand the structure and function of AR. We will investigate AR gene structure at the level of nucleotide sequence for DNA, gene transcription for RNA and gene expression via translation into AR peptide. AR function will be determined by its ability to bind specific androgen response elements and to regulate reporter gene expression. AR gene mutations will be detected by restriction analyses and denaturing gradient gel electrophoresis of exons amplified by the polymerase chain reaction prior to nucleotide sequencing. Northern blots, nuclear run-on assays and solution hybridization assays will be used to detect the level of AR gene transcription and the mRNA transcript size. The structure- function relationship between AR gene mutations, AR gene expression and AR regulation of gene transcription will be determined by correlating AR mRNA levels, radioligand binding activity of AR an the interaction of AR with androgen specific transcriptional regulatory elements. The biological function of AR will be determined by androgen response elements. We will identify heterozygous carriers of AIS from affect pedigrees and perform prenatal diagnosis of infants from heterozygous mothers of this pedigrees and perform prenatal diagnosis of infants from heterozygous mothers of this linked trait. These tools will permit the evaluation of infants with ambiguous genitalia and other abnormalities of male sex differentiation for which the diagnosis of AIS was and other abnormalities of male sex differentiation for which the diagnosis of AIS was often equivocal. The long-term objectives are to elucidate how AR interacts specifically with its ligand and to understand how this binding leads to in vivo activation of gene transcription through specific protein-DNA interactions within the genome.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK043147-02
Application #
3244477
Study Section
Biochemical Endocrinology Study Section (BCE)
Project Start
1990-08-01
Project End
1995-07-31
Budget Start
1991-08-01
Budget End
1992-07-31
Support Year
2
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Johns Hopkins University
Department
Type
Schools of Medicine
DUNS #
045911138
City
Baltimore
State
MD
Country
United States
Zip Code
21218
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