TARGETED KNOCKOUT OF PKD1 GENE IN RENAL COLLECTING DUCT

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DESCRIPTION (Adapted from the Applicant's Abstract): An understanding of the role of PKD1 in the development and maintenance of renal architecture would be advanced by developing an animal model with a PKD1 gene mutation that develop renal cysts and survives into adulthood. PKD1 exon 34 null mice develop renal cysts but die in the perinatal period. This proposal aims to develop an animal model of ADPKD with a mutation in the PKD1 gene that results in limited renal cyst formation and does not effect other organs. The Cre/loxP recombination system will be utilized to selectively truncate the PKD-1 gene in renal collecting duct cells with the human aquaporin-2 promoter driving expression of an epitope tagged Cre recombinase. Transgenic mice in which Cre is expressed selectively in the renal collecting duct have been prepared. Gene targeting techniques will be used to create mice homozygous for floxed exon 34 of the murine PKD1 gene. These mice will be mated with AQP2-CreTag mice to produce mice heterozygous for AQP2-CreTag and either heterozygous or homozygous for the floxed PKD1 gene. We predict that CreTag expressed exclusively in the collecting duct will excise floxed PKD1 exon 34. The resulting mutant PKD1 transcript should be truncated, and non-functional PKD1 protein expressed selectively in the collecting duct. Phenotype will be determined, anticipating that cysts will only form in the medullary collecting duct. RT-PCR will be used to identify mutant PKD1 mRNA, loss of polycstin expression demonstrated by immunohistochemistry, and histological techniques employed to analyze cyst formation. We predict that animals harboring collecting duct specific PKD1 gene truncation will develop renal cysts, yet survive into adulthood. This animal model could then be used to study cystogenesis and therapies relevant to human ADPKD.