Visualization of Cytoskeletal Dynamics in Podocytes Using Adenoviral Vectors

Glomerular visceral epithelial cells (podocytes) play a key role in maintaining selective protein filtration in the kidney. Podocytes have a complex cell shape characterized by the presence of numerous actin-rich processes, which cover the surface of glomerular capillaries and are connected by specialized cell-cell adhesion complexes (slit diaphragms). Human genetic studies and experiments in knockout mouse models show that actin filaments and actin-associated proteins are indispensable for the maintenance of podocyte shape, slit diaphragm integrity, and normal glomerular filtration. The ability to examine cytoskeletal protein organization and dynamics in podocytes and to test the effects of disease-associated mutations on protein localization provides valuable information for researchers aiming to dissect the molecular mechanisms of podocyte dysfunction. We describe how adenovirus-mediated transduction of cultured podocytes with DNA constructs can be used to reliably introduce fluorescently tagged cytoskeletal markers for live cell imaging with high efficiency and low toxicity. This technique can be used to study the dynamic reorganization of the podocyte cytoskeleton and to test the effects of novel mutations on podocyte cytoskeletal dynamics. (c) 2014 Wiley Periodicals, Inc.