HORMONE RESISTANCE IN PATIENTS WITH PSEUDOHYPOPARATHYROIDISM

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The action of many hormones requires their interaction with specific cell surface receptors that are coupled by the guanine nucleotide- binding protein Gs to stimulation of adenylyl cyclase. Expression of hormones, receptors, or Gs molecules with altered biological activity can result in gain or loss of hormone action. This protocol examines the molecular basis for altered hormone action in signal transduction pathways that utilize Gs protein as a signal transducer. These disorders include pseudohypoparathyroidism type la and McCune Albright syndrome due to altered expression or function of Gs, hypoparathyroidism due to defects in the calcium-sensing receptor or PTH gene, and short stature due to defects in the growth hormone-releasing hormone receptor. Biochemical and clinical characterization of the patient's phenotype facilitates a laboratory-based approach to elucidating the molecular mechanism of disease. Over the past year we have focused primarily on elucidation of the molecular basis for PTH resistance in subjects with pseudohypoparathyroidism (PHP) type lb, an autosomal syndrome in which hormone resistance is limited to PTH target tissues and associated with normal G protein activity. We have examined and characterized the biochemical phenotype of subjects with PTH resistance and their unaffected relatives from several multiplex kindreds. Peripheral blood lymphoblasts were transformed with Epstein-Barr virus, and genomic DNA and total RNA were isolated from cultures of the transformed B- lymphocytes. Our previous work had excluded the PTH receptor gene as the basis for PTH resistance in these families. To identify the gene that accounts for PTH resistance in PHP type lb, we have performed a genome-wide scan using semi-automated fluorescence-based PCR technology to analyze linkage of microsatellite marker loci. We have completed a linkage map at 20 cM intervals of all autosomal chromosomes, and using an affected-only analysis we found linkage of PHP type lb to a locus on chromosome 10q that has a LOD score of 2.94. We have recently begun to analyze another large, multiplex family, and we anticipate that the addition of informative affected subjects from this family will increase our LOD score to greater than 3, thus confirming linkage to within 5 cm of the correct locus. Future studies will focus on identification of the candidate gene in this region.