Cadmium favors F-actin depolymerization in rat renal mesangial cells by site-specific, disulfide-based dimerization of the CAP1 protein.

Cadmium is a toxic metal that produces oxidative stress and has been shown to disrupt the actin cytoskeleton in rat renal mesangial cells (RMC). In a survey of proteins that might undergo Cd2+-dependent disulfide crosslinking, we identified the adenylyl cyclase-associated protein, CAP1, as undergoing a dimerization in response to Cd2+ (5-40 µM) that was sensitive to disulfide reducing agents, was reproduced by the disulfide crosslinking agent diamide, and was shown by site-directed mutagenesis to involve the Cys29 residue of the protein. Reactive oxygen species are not involved in the thiol oxidation, and glutathione modulates background levels of dimer. CAP1 is known to enhance cofilin's F-actin severing activity through binding to F-actin and cofilin. F-actin sedimentation and GST-cofilin pulldown studies of CAP1 demonstrated enrichment of the CAP1 dimer's association with cofilin, and in the cofilin-F-actin pellet, suggesting that Cd2+-induced dimer increases the formation of a CAP1-cofilin-F-actin complex. Both siRNA-based silencing of CAP1 and overexpression of a CAP1 mutant lacking Cys29 (and therefore, incapable of dimerization in response to Cd2+) increased RMC viability and provided some protection of F-actin structures against Cd2+. It is concluded that Cd2+ brings about disruption of the RMC cytoskeleton in part through formation of a CAP1 dimer that increases recruitment of cofilin to F-actin filaments.

CAP2 in cardiac conduction, sudden cardiac death and eye development.

Sudden cardiac death kills 180,000 to 450,000 Americans annually, predominantly males. A locus that confers a risk for sudden cardiac death, cardiac conduction disease, and a newly described developmental disorder (6p22 syndrome) is located at 6p22. One gene at 6p22 is CAP2, which encodes a cytoskeletal protein that regulates actin dynamics. To determine the role of CAP2 in vivo, we generated knockout (KO) mice. cap2(-)/cap2(-) males were underrepresented at weaning and ~70% died by 12 weeks of age, but cap2(-)/cap2(-) females survived at close to the expected levels and lived normal life spans. CAP2 knockouts resembled patients with 6p22 syndrome in that mice were smaller and they developed microphthalmia and cardiac disease. The cardiac disease included cardiac conduction disease (CCD) and, after six months of age, dilated cardiomyopathy (DCM), most noticeably in the males. To address the mechanisms underlying these phenotypes, we used Cre-mediated recombination to knock out CAP2 in cardiomyocytes. We found that the mice developed CCD, leading to sudden cardiac death from complete heart block, but no longer developed DCM or the other phenotypes, including sex bias. These studies establish a direct role for CAP2 and actin dynamics in sudden cardiac death and cardiac conduction disease.