Evaluation of a new tool for exploring podocyte biology: Mouse Nphs1 5 ' flanking region drives LacZ expression in podocytes

Study of podocyte biology has been hampered by limitations in available exp erimental models that both recapitulate the in vivo phenotypes of this cell and can be readily and specifically manipulated at the molecular level. Tr ansgenic manipulation of the podocyte represents one approach that might ci rcumvent these limitations. The purpose of this study was to identify a pro moter-enhancer that would direct the expression of transgenes in a podocyte -specific manner. The nephrin (Nphs1) promoter was considered a good candid ate for this purpose, because nephrin was thought to be expressed exclusive ly in podocytes. Two independent BAC clones that contained the murine Nphs1 gene were identified. An 8.3-kb and a 5.4-kb fragment containing the 5' fl anking promoter sequence were identified and characterized. Two constructs were generated by placing a bacterial lacZ reporter with a nuclear localiza tion signal under the control of these two DNA fragments. Mice transgenic f or both constructs were generated. Using a chemiluminescence assay, beta -g alactosidase activity significantly above control was detected only in tiss ue homogenates of kidneys and brain of transgenic mice. In X-gal stained se ctions of transgenic adult kidneys, only podocyte nuclei expressed beta -ga lactosidase, In adult brain examined by tissue sectioning, beta -galactosid ase activity was confined to a discrete area in the medulla. Identical patt erns of beta -galactosidase expression were observed in multiple transgenic founders, suggesting that the expression pattern observed was independent of the site of transgene integration. The developmental expression of beta -galactosidase in transgenic embryos was also analyzed. Transgenes regulate d by this promoter should be useful for studying the biology of gene produc ts that regulate podocyte phenotype and function.