The proximal C-terminus of α_1C subunits is necessary for junctional membrane targeting of cardiac L-type calcium channels

In cardiac myocytes, LTCCs (L-type calcium channels) form a functional signalling complex with ryanodine receptors at the JM (junctional membrane). Although the specific localization of LTCCs to the JM is critical for excitation–contraction coupling, their targeting mechanism is unclear. Transient transfection of GFP (green fluorescent protein)–α_1S or GFP–α_1C, but not P/Q-type calcium channel α_1A, in dysgenic (α_1S-null) GLT myotubes results in correct targeting of these LTCCs to the JMs and restoration of action-potential-induced Ca²⁺ transients. To identify the sequences of α_1C responsible for JM targeting, we generated a range of α_1C–α_1A chimaeras, deletion mutants and alanine substitution mutants and studied their targeting properties in GLT myotubes. The results revealed that amino acids L¹⁶⁸¹QAGLRTL¹⁶⁸⁸ and P¹⁶⁹³EIRRAIS¹⁷⁰⁰, predicted to form two adjacent α-helices in the proximal C-terminus, are necessary for the JM targeting of α_1C. The efficiency of restoration of action-potential-induced Ca²⁺ transients in GLT myotubes was significantly decreased by mutations in the targeting motif. JM targeting was not disrupted by the distal C-terminus of α_1C which binds to the second α-helix. Therefore we have identified a new structural motif in the C-terminus of α_1C that mediates the targeting of cardiac LTCCs to JMs independently of the interaction between proximal and distal C-termini of α_1C.