Selectively suppressed Ca2+-induced Ca2+ release activity of α-ryanodine receptor (α-RyR) in frog skeletal muscle sarcoplasmic reticulum -: Potential distinct modes in Ca2+ release between α- and β-RyR

We reported earlier that the two ryanodine receptor (RyR) isoforms (alpha- and beta -RyR) purified from frog skeletal muscle were equipotent in the Ca2+-induced Ca2+ release (CICR) activity (Murayama, T., Kurebayashi, N., and Ogawa, Y. (2000) Biophys. J. 78, 1810-1824), Whether this is also the case with the native Ca2+ release channel in the sarcoplasmic reticulum (SR), however, remains to be determined. Taking advantage of the facts that [SH]ryanodine binds only to the open form of the channels and that it is practically irreversible at 4 degreesC, we devised a method to separate the total binding to contributions of alpha- and beta -RyR, using immunoprecipitation with an alpha -RyR-specific monoclonal antibody. Surprisingly, the binding of alpha -RyR was strongly suppressed to as low as similar to4% that of beta -RyR in the SR vesicles. The two isoforms, however, showed no difference in sensitivity to Ca2+, adenine nucleotides, or caffeine. This reduced binding of alpha -RyR was ascribed to the low affinity for [3H]ryanodine, with no change in the maximal binding sites. Solubilization of SR with 3-[(3-cholamidopropyl) dimethylammonio]-1-propanesulfonic acid partly remedied this nonequivalence, whereas 1 M NaCl was ineffective. 12-kDa FK506-binding protein (FKBP12), however, could not be responsible for it, because FK506 treatment did not eliminate the suppression, in contrast to marked removal of 12-kDa FK506-binding protein from alpha -RyR. These results suggest that alpha -RyR in the SR may serve Ca2+ release in a mode other than CICR, being selectively suppressed in CICR.