Two Domains in Dihydropyridine Receptor Activate the Skeletal Muscle Ca 2+ Release Channel

Abstract

The II-III cytoplasmic loop of the skeletal muscle dihydropyridine receptor (DHPR) α 1-subunit is essential for skeletal-type excitation-contraction coupling. Single channel and [ 3H]ryanodine binding studies with a full-length recombinant peptide (p 666–791) confirmed that this region specifically activates skeletal muscle Ca 2+ release channels (CRCs). However, attempts to identify shorter domains of the II-III loop specific for skeletal CRC activation have yielded contradictory results. We assessed the specificity of the interaction of five truncated II-III loop peptides by comparing their effects on skeletal and cardiac CRCs in lipid bilayer experiments; p 671–680 and p 720–765 specifically activated the submaximally Ca 2+-activated skeletal CRC in experiments using both mono and divalent ions as current carriers. A third peptide, p 671–690, showed a bimodal activation/inactivation behavior indicating a high-affinity activating and low-affinity inactivating binding site. Two other peptides (p 681–690 and p 681–685) that contained an RKRRK-motif and have previously been suggested in in vitro studies to be important for skeletal-type E-C coupling, failed to specifically stimulate skeletal CRCs. Noteworthy, p 671–690, p 681–690, and p 681–685 induced similar subconductances and long-lasting channel closings in skeletal and cardiac CRCs, indicating that these peptides interact in an isoform-independent manner with the CRCs.