Analysis of the Complex between Ca2+ Channel β-Subunit and the Rem GTPase

• Published in: The Journal of biological chemistry Vol. 281; no. 33; p. 23557
• Main Authors: Brian S. Finlin, Robert N. Correll, Chunyan Pang, Shawn M. Crump, Jonathan Satin, Douglas A. Andres
• Format: Journal Article
• Language: English
• Published: American Society for Biochemistry and Molecular Biology 18.08.2006
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M604867200

Voltage-gated calcium channels are multiprotein complexes that regulate calcium influx and are important contributors to cardiac excitability and contractility. The auxiliary β-subunit (Ca V β) binds a conserved domain (the α-interaction domain (AID)) of the pore-forming Ca V α 1 subunit to modulate channel gating properties and promote cell surface trafficking. Recently, members of the RGK family of small GTPases ( R em, R em2, R ad, G em/ K ir) have been identified as novel contributors to the regulation of L-type calcium channel activity. Here, we describe the Rem-association domain within Ca V β 2a . The Rem interaction module is located in a ∼130-residue region within the highly conserved guanylate kinase domain that also directs AID binding. Importantly, Ca V β mutants were identified that lost the ability to bind AID but retained their association with Rem, indicating that the AID and Rem association sites of Ca V β 2a are structurally distinct. In vitro binding studies indicate that the affinity of Rem for Ca V β 2a interaction is lower than that of AID for Ca V β 2a . Furthermore, in vitro binding studies indicate that Rem association does not inhibit the interaction of Ca V β 2a with AID. Instead, Ca V β can simultaneously associate with both Rem and Ca V α 1 -AID. Previous studies had suggested that RGK proteins may regulate Ca 2+ channel activity by blocking the association of Ca V β subunits with Ca V α 1 to inhibit plasma membrane trafficking. However, surface biotinylation studies in HIT-T15 cells indicate that Rem can acutely modulate channel function without decreasing the density of L-type channels at the plasma membrane. Together these data suggest that Rem-dependent Ca 2+ channel modulation involves formation of a Rem·Ca V β·AID regulatory complex without the need to disrupt Ca V α 1 ·Ca V β association or alter Ca V α 1 expression at the plasma membrane.