Excitation-contraction coupling is unaffected by drastic alteration of thesequence surrounding residues L720-L764 of the alpha(1S) II-III loop

The II-III loop of the skeletal muscle dihydropyridine receptor (DHPR) alph a (15) subunit is responsible for bidirectional-signaling interactions with the ryanodine receptor (RyR1): transmitting an orthograde, excitation-cont raction (EC) coupling signal to RyR1 and receiving a retrograde, current-en hancing signal from RyR1. Previously, several reports argued for the import ance of two distinct regions of the skeletal II-III loop (residues R681-L69 0 and residues L720-Q765, respectively), claiming for each a key function i n DHPR-RyR1 communication. To address whether residues 720-765 of the II-II I loop are sufficient to enable skeletal-type (Ca2+ entry-independent) EC c oupling and retrograde interaction with RyR1, we constructed a green fluore scent protein (GFP)-tagged chimera (GFP-SkLM) having rabbit skeletal (Sk) D HPR sequence except for a II-III loop (L) from the DHPR of the house fly, M usca domestica (M). The Musca II-III loop (75% dissimilarity to alpha (15)) has no similarity to alpha (15) in the regions R681-L690 and L720-Q765. GF P-SkLM expressed in dysgenic myotubes (which lack endogenous cuts subunits) was unable to restore EC coupling and displayed strongly reduced Ca2+ curr ent densities despite normal surface expression levels and correct triad ta rgeting (colocalization with RyR1), Introducing rabbit alpha (15) residues L720-L764 into the Musca II-III loop of GFP-SkLM (substitution for Musca DH PR residues E724-T755) completely restored bidirectional coupling, indicati ng its dependence on cuts loop residues 720-764 but its independence from o ther regions of the loop. Thus, 45 alpha (15)-residues embedded in a very d issimilar background are sufficient to restore bidirectional coupling, indi cating that these residues may be a site of a protein-protein interaction r equired for bidirectional coupling.