Association of calcium channel α1S and β1a subunits is required for the targeting of β1a but not of α1S into skeletal muscle triads

The skeletal muscle L-type Ca2+ channel is a complex of five subunits that is specifically localized in the triad, Its primary function is the rapid activation of Ca2+ release from cytoplasmic stores in a process called excitation-contraction coupling. To study the role of alpha(1S)-beta(1a) interactions in the incorporation of the functional channel complex into the triad, alpha(1S) and beta(1a) [or a beta(1a)-green fluorescent protein (GFP) fusion protein] were expressed alone and in combination in myotubes of the dysgenic cell line GLT, beta GFP expressed in dysgenic myotubes that lack the skeletal muscle alpha(1S) subunit was diffusely distributed in the cytoplasm. On coexpression with the alpha(1S) subunit beta GFP distribution became clustered and colocalized with alpha(1S) immunofluorescence. Based on the colocalization of beta GFP and alpha(1S) with the ryanodine receptor the clusters were identified as T-tubule/sarcoplasmic reticulum junctions. Expression of alpha(1S) with and without beta(1a) restored Ca2+ currents and depolarization-induced Ca2+ release. The translocation of beta GFP from the cytoplasm into the junctions failed when beta GFP was coexpressed with alpha(1S) mutants in which the beta interaction domain had been altered (alpha(1S)-Y366S) or deleted (alpha(1S)-Delta 351-380). Although alpha(1S)-Y366S did not associate with beta GFP it was incorporated into the junctions, and it restored Ca2+ currents and depolarization-induced Ca2+ release. Thus, beta(1a) requires the association with the beta interaction domain in the I-II cytoplasmic loop of alpha(1S) for its own incorporation into triad junctions, but stable alpha(1S)-beta(1a) association is not necessary for the targeting of alpha(1S) into the triads or for its normal function in Ca2+ conductance and excitation-contraction coupling.