Polycystic kidney disease (PKD) can be inherited as an autosomal dominant (ADPKD) or autosomal recessive (ARPKD) trait, or can be provoked by environmental factors (acquired cystic disease). The disease is characterized by the growth of large epithelial-lined cysts from the nephrons and collecting ducts of affected kidneys. In all types of inherited PKD, cyst formation is initiated early in development, in the fetal kidney, suggesting that all forms of the disease are caused by abnormalities in early kidney organogenesis. Cysts are thought to initiate as small dilations in renal tubules, which then expand into fluid-filled cavities of relatively large size. Cyst formation appears to require both increased cell proliferation and epithelial fluid secretion, while more advanced cystic kidneys display a number of progressive cell growth abnormalities, including changes in cell morphology, formation of micropolyps, and development of adenomas and adenocarcinomas. The experiments in this proposal will make use of two murine models of PKD. Disease development in the C57BL/6J-cpk mouse appears similar to human ARPKD, and in the DBA-pcy mouse appears similar to human ADPKD. We have evidence that the cpk and pcy mutations profoundly affect gene expression in cystic kidneys. In particular, there is markedly elevated expression of several proto-oncogenes, and there is persistent expression of the developmentally expressed SGP-2 (sulfated glycoprotein-2) gene. Differential cDNA library construction/screening will be carried out to isolate, identify and characterize over- and underexpressed genes associated with PKD in cpk and pcy mice. These genes will be studied by examining their expression by Northern and in situ hybridization, and by expressing them in cultured cells to explore their function. The cDNAs will also be used to find families of genes that are misregulated, whose promoters will be experimentally analyzed to determine how they are controlled and therefore what may be responsible for initiating the cascade of genetic events leading to cyst formation. Kidney-specific genes that fail to be expressed in cystic kidneys are candidates for the cpk and pcy genes themselves. The regulation of the SGP-2 gene will be investigated in the kidney and in cell culture to determine why it is not turned off during cyst formation. These studies will make use of promoter/reporter constructs that will be mutated and expressed in cells and in transgenic mice. We also plan to isolate kidney-specific growth inhibitory genes by differential cDNA library screening of growth-arrested kidney cells in culture. These cDNAs will then be expressed under the control of an inducible heterologous promoter to determine if they inhibit cell proliferation in vitro. PKD may be caused by uncontrolled cell proliferation because of (or leading to) an inability of affected cells to undergo terminal differentiation, and this failure could be due to a defective kidney-specific growth inhibitory gene whose lack of expression would initiate the genetic events leading ultimately to the polycystic phenotype.
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