Central core disease mutations R4892W, I4897T and G4898E in the ryanodine receptor isoform 1 reduce the Ca2+ sensitivity and amplitude of Ca2+-dependent Ca2+ release.
Three CCD (central core disease) mutants, R4892W (Arg4892-->), I4897T and G4898E, in the pore region of the skeletal-muscle Ca2+-release channel RyR1 (ryanodine receptor 1) were characterized using a newly developed assay that monitored Ca2+ release in the presence of Ca2+ uptake in microsomes isolated from HEK-293 cells (human embryonic kidney 293 cells), co-expressing each of the three mutants together with SERCA1a (sarcoplasmic/endoplasmic-reticulum Ca2+-ATPase 1a). Both Ca2+ sensitivity and peak amplitude of Ca2+ release were either absent from or sharply decreased in homotetrameric mutants. Co-expression of wild-type RyR1 with mutant RyR1 (heterotetrameric mutants) restored Ca2+ sensitivity partially, in the ratio 1:2, or fully, in the ratio 1:1. Peak amplitude was restored only partially in the ratio 1:2 or 1:1. Reduced amplitude was not correlated with maximum Ca2+ loading or the amount of expressed RyR1 protein. High-affinity [3H]ryanodine binding and caffeine-induced Ca2+ release were also absent from the three homotetrameric mutants. These results indicate that decreased Ca2+ sensitivity is one of the serious defects in these three excitation-contraction uncoupling CCD mutations. In CCD skeletal muscles, where a mixture of wild-type and mutant RyR1 is expressed, these defects are expected to decrease Ca2+-induced Ca2+ release, as well as orthograde Ca2+ release, in response to transverse tubular membrane depolarization.
Du
Guo Guang
GG
Banting and Best Department of Medical Research, Charles H. Best Institute, University of Toronto, 112 College St., Toronto, ON, Canada M5G 1L6.
Khanna
Vijay K
VK
Guo
Xinghua
X
MacLennan
David H
DH
D000255:Adenosine Triphosphate / Q000378:metabolism; D001120:Arginine / Q000235:genetics; D002118:Calcium / Q000737:chemistry / Q000378:metabolism; D002127:Calcium Isotopes / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000096:biosynthesis; D002460:Cell Line; D018698:Glutamic Acid / Q000235:genetics; D005998:Glycine / Q000235:genetics; D006801:Humans; D007532:Isoleucine / Q000378:metabolism; D007668:Kidney / Q000737:chemistry / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D008861:Microsomes / Q000737:chemistry / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense / Q000235:genetics*; D020512:Myopathy, Central Core / Q000235:genetics*; D010070:Oxalates / Q000378:metabolism; D010783:Photometry / Q000379:methods; D020033:Protein Isoforms / Q000235:genetics; D019837:Ryanodine Receptor Calcium Release Channel / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D013912:Threonine / Q000235:genetics; D014364:Tryptophan / Q000235:genetics
D002127:Calcium Isotopes; D010070:Oxalates; D020033:Protein Isoforms; D019837:Ryanodine Receptor Calcium Release Channel; D007532:Isoleucine; D013912:Threonine; D018698:Glutamic Acid; D014364:Tryptophan; D000255:Adenosine Triphosphate; D001120:Arginine; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium; D005998:Glycine