α-Actinin-4 Is Required for Normal Podocyte Adhesion

Abstract

Mutations in the ␣-actinin-4 gene ACTN4 cause an autosomal dominant human kidney disease. Mice deficient in ␣-actinin-4 develop a recessive phenotype characterized by kidney failure, proteinuria, glomerulosclerosis, and retraction of glomerular podocyte foot processes. However, the mechanism by which ␣-actinin-4 deficiency leads to glomerular disease has not been defined. Here, we examined the effect of ␣-actinin-4 deficiency on the adhesive properties of podocytes in vivo and in a cell culture system. In ␣-actinin-4-deficient mice, we observed a decrease in the number of podocytes per glomerulus compared with wild-type mice as well as the presence of podocyte markers in the urine. Podocyte cell lines generated from ␣-actinin-4-deficient mice were less adherent than wild-type cells to glomerular basement membrane (GBM) components collagen IV and laminin 10 and 11. We also observed markedly reduced adhesion of ␣-actinin-4-deficient podocytes under increasing shear stresses. This adhesion deficit was restored by transfecting cells with ␣-actinin-4-GFP. We tested the strength of the integrin receptor-mediated linkages to the cytoskeleton by applying force to microbeads bound to integrin using magnetic pulling cytometry. Beads bound to ␣-actinin-4-deficient podocytes showed greater displacement in response to an applied force than those bound to wild-type cells. Consistent with integrin-dependent ␣-actinin-4-mediated adhesion, phosphorylation of ␤1-integrins on ␣-actinin-4-deficient podocytes is reduced. We rescued the phosphorylation deficit by transfecting ␣-actinin-4 into ␣-actinin-4-deficient podocytes. These results suggest that ␣-actinin-4 interacts with integrins and strengthens the podocyte-GBM interaction thereby stabilizing glomerular architecture and preventing disease.

Wnt/β-Catenin Pathway in Podocytes Integrates Cell Adhesion, Differentiation, and Survival

“…We used the following primary antibodies for the experiments: total Ctnnb1 (sc-7199 Santa Cruz Biotechnology); active Ctnnb1 (05-665, Millipore); and ␤-actin (ab8226, Abcam). For Wt1 detection of urinary cells, spot urine was collected and centrifuged, and the sediment was dissolved in loading buffer (32).…”

Mechanisms of angiotensin II signaling on cytoskeleton of podocytes

“…Recently, a experimental study showed that α actinin-4 null mice have severe glomerular disease, 91 and α actinin-4 is required for normal podocyte adhesion. 92 Little is known about the effects of α actinin-4 on Ang II-related actin cytoskeletal change in podocytes. This work examined the function of α actinin-4 in AT1R signaling and the associated mechanisms leading to FSGS in Neph-hAT1 TGRs.…”

“…92 To explore further the mechanisms linking chronically increased AT1R signaling in podocytes and proteinuria, expression levels of α actinin-4 in Neph-hAT1 TGRs (Fig. 4.6.1 a) were determined.…”

Ultrastructural Features and Expression of Cytoskeleton Proteins of Podocyte from Patients with Minimal Change Disease and Focal Segmental Glomerulosclerosis

“…[17,24] An in vitro study had proved that podocytes generated from α-actinin-4-deficient mice showed aggregates and increased degradation of the mutant proteins, as well as decreased adhesive ability to GBM components. [25,26] Both in vivo and in vitro studies suggested that the breakdown of cytoskeleton filaments was the final common pathway lead to foot process effacement and aggregated microfilaments of podocytes and its dysfunction. Goode et al [14] reported that the expression of α-actinin-4 in podocytes of MCD was similar to that of normal kidney, while an elevated α-actinin-4 expression could be found in the areas of subepithelial deposits of podocytes in membranous nephropathy.…”