Congenital nephrogenic diabetes insipidus (CNDI) is an inherited X-linked disorder of water homeostasis. Affected individuals fail to concentrate their urine during periods of low water intake because the kidney is insensitive to the effects of the water-sparing, pituitary-derived hormone, antidiuretic hormone (ADH). As a result, they quickly become dehydrated and their body fluids hyperosmolar when water intake is restricted. In infants, the hyperosmolar dehydration may result in permanent brain damage or death. With time, the polyuria resulting from the increased water intake needed to avoid dehydration may cause permanent anatomic dilatation of the urinary tract structures, increasing the likelihood of urinary tract infections and traumatic rupture. Renal failure may also be a late complication. A cDNA for the human kidney-specific ADH receptor, the V2 ADH receptor (V2 ADHR), was recently cloned and sequenced. Its predicted protein product has seven hydrophobic alpha-helical domains suggesting membership in the large superfamily of seven-transmembrane-segment (STM) receptors that usually modulate second messengers via association with heterotrimeric guanine-nucleotide binding complexes (G proteins). In most patients with CNDI studied, mutations in the V2 ADHR gene are present and likely explain the resistance to ADH. Over six- mutations are known to date. The proposed research is intended to clarify how the genetic defects in the V2 ADHR molecule affect its ability to transduce an ADH mediated signal, and thus should provide a genetic approach to the modeling of structure-function relationships in STM receptors.
The specific aims of the proposed research are: l. To determine the genetic mutations in the V2 ADHR in 20 or more unrelated families with CNDI: 2. To assay the amount of residual function of each mutant receptor; and 3. To determine how each defect results in diminished receptor function, i.e. whether membrane targeting, ligand binding, G protein coupling, or simply signaling is altered.
These aims will be accomplished by sequencing the V2ADHR genes from affected families expressing the mutated genes in cultured cells to assay signal transduction capacity by a sensitive measure of cAMP generation in response to ligand. Subcellular localization of the mutant receptors will be evaluated by immunohistology, and ligand binding by standard radioactive ligand techniques. G protein coupling will be detected by demonstrating GTP-dependent dual ADH affinity states and by co- immunoprecipitation. Understanding the spectrum of naturally occurring receptor defects causing CNDI may facilitate the design of rational ameliorative therapies for CNDI and related disorders. The structure-function implications of the V2 ADHR mutation analysis will also contribute to a fuller understanding of STM receptors in general.
| Smyk-Pearson, S K; Bakke, A C; Held, P K et al. (2003) Rescue of the autoimmune scurfy mouse by partial bone marrow transplantation or by injection with T-enriched splenocytes. Clin Exp Immunol 133:193-9 |
| Wildin, R S; Smyk-Pearson, S; Filipovich, A H (2002) Clinical and molecular features of the immunodysregulation, polyendocrinopathy, enteropathy, X linked (IPEX) syndrome. J Med Genet 39:537-45 |
