Dynamic phospholipid interaction of ?2e subunit regulates the gating of voltage-gated Ca2+ channels.

ABSTRACT: High voltage-activated Ca2+ (Ca(V)) channels are protein complexes containing pore-forming ?1 and auxiliary ? and ?2? subunits. The subcellular localization and membrane interactions of the ? subunits play a crucial role in regulating Ca(V) channel inactivation and its lipid sensitivity. Here, we investigated the effects of membrane phosphoinositide (PI) turnover on Ca(V)2.2 channel function. The ?2 isoform ?2e associates with the membrane through electrostatic and hydrophobic interactions. Using chimeric ? subunits and liposome-binding assays, we determined that interaction between the N-terminal 23 amino acids of ?2e and anionic phospholipids was sufficient for ?2e membrane targeting. Binding of the ?2e subunit N terminus to liposomes was significantly increased by inclusion of 1% phosphatidylinositol 4,5-bisphosphate (PIP2) in the liposomes, suggesting that, in addition to phosphatidylserine, PIs are responsible for ?2e targeting to the plasma membrane. Membrane binding of the ?2e subunit slowed Ca(V)2.2 current inactivation. When membrane phosphatidylinositol 4-phosphate and PIP2 were depleted by rapamycin-induced translocation of pseudojanin to the membrane, however, channel opening was decreased and fast inactivation of Ca(V)2.2(?2e) currents was enhanced. Activation of the M1 muscarinic receptor elicited transient and reversible translocation of ?2e subunits from membrane to cytosol, but not that of ?2a or ?3, resulting in fast inactivation of Ca(V)2.2 channels with ?2e. These results suggest that membrane targeting of the ?2e subunit, which is mediated by nonspecific electrostatic insertion, is dynamically regulated by receptor stimulation, and that the reversible association of ?2e with membrane PIs results in functional changes in Ca(V) channel gating. The phospholipid-protein interaction observed here provides structural insight into mechanisms of membrane-protein association and the role of phospholipids in ion channel regulation.

SUBMITTER: Kim DI 

PROVIDER: S-EPMC4442786 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature