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J Am Soc Nephrol 11:2306-2314, 2000
© 2000 American Society of Nephrology

Evaluation of a New Tool for Exploring Podocyte Biology: Mouse Nphs1 5' Flanking Region Drives LacZ Expression in Podocytes

MARCUS J. MOELLER*, IULIA A. KOVARI* and LAWRENCE B. HOLZMAN*,{dagger}

* Division of Nephrology, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, Michigan.
{dagger} Department of Veterans Affairs, Ann Arbor, Michigan.

Correspondence to Dr. Lawrence B. Holzman, University of Michigan Medical School, 1560 Medical Science Research Building II, Ann Arbor, MI 48109-0676. Phone: 734-764-3157; Fax: 734-763-0982; E-mail: lholzman{at}umich.edu

Abstract. Study of podocyte biology has been hampered by limitations in available experimental models that both recapitulate the in vivo phenotypes of this cell and can be readily and specifically manipulated at the molecular level. Transgenic manipulation of the podocyte represents one approach that might circumvent these limitations. The purpose of this study was to identify a promoter-enhancer that would direct the expression of transgenes in a podocyte-specific manner. The nephrin (Nphs1) promoter was considered a good candidate for this purpose, because nephrin was thought to be expressed exclusively 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' flanking promoter sequence were identified and characterized. Two constructs were generated by placing a bacterial lacZ reporter with a nuclear localization signal under the control of these two DNA fragments. Mice transgenic for both constructs were generated. Using a chemiluminescence assay, ß-galactosidase activity significantly above control was detected only in tissue homogenates of kidneys and brain of transgenic mice. In X-gal stained sections of transgenic adult kidneys, only podocyte nuclei expressed ß-galactosidase. In adult brain examined by tissue sectioning, ß-galactosidase activity was confined to a discrete area in the medulla. Identical patterns of ß-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 ß-galactosidase in transgenic embryos was also analyzed. Transgenes regulated by this promoter should be useful for studying the biology of gene products that regulate podocyte phenotype and function.




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