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

• Published in: Biochemical journal 2004-09, Vol.382 (Pt 2), p.557-564
• Main Authors: Du, Guo Guang, Khanna, Vijay K, Guo, Xinghua, MacLennan, David H
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphate - metabolism; Arginine - genetics; Calcium - chemistry; Calcium - metabolism; Calcium Isotopes - metabolism; Calcium-Transporting ATPases - biosynthesis; Cell Line; Glutamic Acid - genetics; Glycine - genetics; Humans; Isoleucine - metabolism; Kidney - chemistry; Kidney - cytology; Kidney - embryology; Kidney - metabolism; Microsomes - chemistry; Microsomes - metabolism; Mutagenesis, Site-Directed; Mutation, Missense - genetics; Myopathy, Central Core - genetics; Oxalates - metabolism; Photometry - methods; Protein Isoforms - genetics; Ryanodine Receptor Calcium Release Channel - biosynthesis; Ryanodine Receptor Calcium Release Channel - genetics; Ryanodine Receptor Calcium Release Channel - metabolism; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Threonine - genetics; Tryptophan - genetics
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj20040580

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.