Cardiac ryanodine receptor is absent in type I slow skeletal muscle fibers: immunochemical and ryanodine binding studies.
Cardiac ryanodine receptor was purified from canine ventricle as a single polypeptide of Mr 400,000 by a stepwise sucrose density gradient centrifugation and heparin-Sepharose CL-4B column chromatography. The [3H]ryanodine binding capacity (Bmax) was 60-fold enriched from cardiac microsomes without a change in affinity for [3H]ryanodine. The purity of the final preparation was determined to be greater than 95% by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Using this purified preparation as an antigen, we produced six monoclonal antibodies which immunoprecipitated the cardiac ryanodine receptor. Three of these antibodies recognized the cardiac receptor on immunoblot analysis. In contrast, no protein in the microsomes isolated from Type I (slow) or Type II (fast) skeletal muscles was recognized by these antibodies. The [3H]ryanodine binding to cardiac and skeletal muscle microsomes was dependent on free Ca2+ concentration. In skeletal muscle microsomes, the [3H]ryanodine binding was remarkably enhanced by the addition of ATP or KCl and inhibited by high free Ca2+, whereas it was less sensitive to these agents in cardiac microsomes. All of these results clearly demonstrate that the cardiac ryanodine receptor is different from the skeletal muscle receptors and is not present even in Type I (slow) skeletal muscle fibers, in which cardiac isoforms of some of the muscle proteins are constitutively expressed.
Imagawa
T
T
Department of Molecular Physiology, National Cardiovascular Center Research Institute, Osaka.
Takasago
T
T
Shigekawa
M
M
D000470:Alkaloids / Q000378:metabolism*; D000818:Animals; D000911:Antibodies, Monoclonal; D006224:Cricetinae; D004285:Dogs; D006168:Guinea Pigs; D015151:Immunoblotting; D007120:Immunochemistry; D008566:Membranes / Q000378:metabolism; D008861:Microsomes / Q000378:metabolism; D009132:Muscles / Q000378:metabolism*; D009206:Myocardium / Q000378:metabolism* / Q000648:ultrastructure; D011233:Precipitin Tests; D011817:Rabbits; D051381:Rats; D011950:Receptors, Cholinergic / Q000032:analysis / Q000378:metabolism*; D012433:Ryanodine / Q000378:metabolism*; D019837:Ryanodine Receptor Calcium Release Channel; D013045:Species Specificity
D000470:Alkaloids; D000911:Antibodies, Monoclonal; D011950:Receptors, Cholinergic; D019837:Ryanodine Receptor Calcium Release Channel; D012433:Ryanodine