Abstract

Mutations in the gene ACTN4 encoding the actin bundling protein-alpha-actinin-4 underlie an inherited form of kidney lesions known as focal segmental glomerulosclerosis (FSGS). Previously, we developed a model for this condition by generating mice with podocyte-specific overexpression of a disease-causing mutant alpha-actinin-4 (K256E-ACTN4 (pod+)). However, whether alpha-actinin-4 overexpression artifacts and not the gain of affinity effects of the mutation accounted for the robust FSGS phenotype in these mice was unclear. To address this question, we developed a control line of mice with podocyte-specific overexpression of wildtype alpha-actinin-4 (wt-ACTN4 (pod+)). An 8.3 kb fragment of the mouse nephrin promoter (NPHS1) was used to drive expression of a hemagglutinin (HA)-tagged wildtype alpha-actinin-4 coding sequence in mice. Five founder lines expressing the HA-tagged alpha-actinin-4 protein in a podocyte-specific manner were obtained, as determined by co-immunofluorescence with HA and synaptopodin antibodies. Quantitative PCR revealed that renal transgene mRNA levels of wt-ACTN4 (pod+) mice are similar to K256E-ACTN4 (pod+) mice. In contrast to K256E-ACTN4 (pod+) mice which exhibit albuminuria, podocyte foot process effacement and glomerular scarring, wt-ACTN4 (pod+) mice are healthy and indistinguishable from non-transgenic littermates. These findings suggest that the K256E mutation itself and not overexpression of alpha-actinin-4 protein per se accounts for the FSGS phenotype in our transgenic model.