DEVELOPMENT OF CELLULAR DIVERSITY IN THE COLLECTING DUCT

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The broad objectives of the proposed research are to understand the cellular and molecular mechanisms that lead to the development of cell heterogeneity both with and between segments of the collecting duct system. Fundamental to this project is our ability to isolate and culture various cell types of the collecting duct, using cell-specific antibodies and cell sorting, and the development of collecting duct cell lines Using these tools, we have recently established that beta-intercalated cells in culture exhibit a remarkable plasticity and can give rise to both alpha-intercalated and principal cells. This cellular conversion seems to involve formation of a hybrid cell, followed by an asymmetric cell division resulting in two antigenically different daughter cells. In this proposal we plan to expand these observations to the developing kidney. Aim (1) will test the hypothesis that the heterogeneity of the collecting duct system arises through cellular interconversion and the influence of neighboring nephron segments. We will examine the effect of cell-cell and cell-matrix interactions on the phenotype of cultured collecting duct cells, correlate proliferation rates with the sequence of expression of cell-specific antigens during kidney development; and follow the fate of genetically tagged cells reintroduced into developing kidneys. Aim (2) will test the hypothesis that asymmetric cell division is a novel mechanism for simultaneous cell differentiation and stem cell renewal and that it is, at least in part, mediated by electric field-induced polarization of membrane proteins. Aim (3) proposes to determine the role of beta-intercalated cells in kidney development, by cell-specific ablation in transgenic animals with toxic genes controlled by beta-intercalated cell-specific regulatory DNA sequences. It is anticipated that these studies will expand our knowledge of the basic mechanisms of kidney development and help to understand the pathogenesis of genetic disorders such as polycystic kidney disease. In addition, we believe that some of the fundamental mechanisms we plan to explore with kidney cells, like cell differentiation and stem cell renewal via cell polarization and asymmetric cell division, are broadly applicable to other pathological conditions such as developmental disorders and malignancies.