alpha-Actinin Promotes Surface Localization and Current Density of the Ca2+ Channel Ca(V)1.2 by Binding to the IQ Region of the alpha 1 Subunit

作者：Tseng Pang Yen; Henderson Peter B; Hergarden Anne C; Patriarchi Tommaso; Coleman Andrea M; Lillya Mark W; Montagut Bordas Carlota; Lee Boram; Hell Johannes W^*; Horne Mary C^*

来源：Biochemistry, 2017, 56(28): 3669-3681.

DOI：10.1021/acs.biochem.7b00359

The voltage-gated L-type Ca2+ channel Ca(V)1.2 is crucial for initiating heartbeat and control of a number of neuronal functions such as neuronal excitability and long-term potentiation. Mutations of Ca(V)1.2 subunits result in serious health problems, including arrhythmia, autism spectrum disorders, immunodeficiency, and hypoglycemia. Thus, precise control of Ca(V)1.2 surface expression and localization is essential. We previously reported that a-actinin associates and colocalizes with neuronal Ca(V)1.2 channels and that shRNA-mediated depletion of a-actinin significantly reduces localization of endogenous Ca(V)1.2 in dendritic spines in hippocampal neurons. Here we investigated the hypothesis that direct binding of alpha-actinin to Ca(V)1.2 supports its surface expression. Using two-hybrid screens and pull-down assays, we identified three point mutations (K1647A, Y1649A, and I1654A) in the central, pore-forming alpha(1)1.2 subunit of Ca(V)1.2 that individually impaired a-actinin binding. Surface biotinylation and flow cytometry assays revealed that Ca(V)1.2 channels composed of the corresponding a-actinin-binding-deficient mutants result in a 35-40% reduction in surface expression compared to that of wild-type channels. Moreover, the mutant Ca(V)1.2 channels expressed' in HEK293 cells exhibit a 60-75% decrease in current density. The larger decrease in current density as compared to surface expression imparted by these alpha(1)1.2 subunit mutations hints at the possibility that alpha-actinin not only stabilizes surface localization of Ca(V)1.2 but also augments its ion conducting activity.

• 出版日期2017-7-18
• 单位NIH