Involvement of Parkin in the ubiquitin proteasome system-mediated degradation of N-type voltage-gated Ca(2+) channels

N-type calcium (Ca(V)2.2) channels are widely expressed in the brain and the peripheral nervous system, where they play important roles in the regulation of transmitter release. Although Ca(V)2.2 channel expression levels are precisely regulated, presently little is known regarding the molecules that mediate its synthesis and degradation. Previously, by using a combination of biochemical and functional analyses, we showed that the complex formed by the light chain 1 of the microtubule-associated protein 1B (LC1-MAP1B) and the ubiquitin-proteasome system (UPS) E2 enzyme UBE2L3, may interact with the Ca(V)2.2 channels promoting ubiquitin-mediated degradation. The present report aims to gain further insights into the possible mechanism of degradation of the neuronal Ca(V)2.2 channel by the UPS. First, we identified the enzymes UBE3A and Parkin, members of the UPS E3 ubiquitin ligase family, as novel Ca(V)2.2 channel binding partners, although evidence to support a direct protein-protein interaction is not yet available. Immunoprecipitation assays confirmed the interaction between UBE3A and Parkin with Ca(V)2.2 channels heterologously expressed in HEK-293 cells and in neural tissues. Parkin, but not UBE3A, overexpression led to a reduced Ca(V)2.2 protein level and decreased current density. Electrophysiological recordings performed in the presence of MG132 prevented the actions of Parkin suggesting enhanced channel proteasomal degradation. Together these results unveil a novel functional coupling between Parkin and the Ca(V)2.2 channels and provide a novel insight into the basic mechanisms of Ca(V) channels protein quality control and functional expression.