Oxygen-coupled redox regulation of the skeletal muscle ryanodine receptor/Ca2+ release channel (RyR1): sites and nature of oxidative modification.


Journal Article

In mammalian skeletal muscle, Ca(2+) release from the sarcoplasmic reticulum (SR) through the ryanodine receptor/Ca(2+)-release channel RyR1 can be enhanced by S-oxidation or S-nitrosylation of separate Cys residues, which are allosterically linked. S-Oxidation of RyR1 is coupled to muscle oxygen tension (pO2) through O2-dependent production of hydrogen peroxide by SR-resident NADPH oxidase 4. In isolated SR (SR vesicles), an average of six to eight Cys thiols/RyR1 monomer are reversibly oxidized at high (21% O2) versus low pO2 (1% O2), but their identity among the 100 Cys residues/RyR1 monomer is unknown. Here we use isotope-coded affinity tag labeling and mass spectrometry (yielding 93% coverage of RyR1 Cys residues) to identify 13 Cys residues subject to pO2-coupled S-oxidation in SR vesicles. Eight additional Cys residues are oxidized at high versus low pO2 only when NADPH levels are supplemented to enhance NADPH oxidase 4 activity. pO2-sensitive Cys residues were largely non-overlapping with those identified previously as hyperreactive by administration of exogenous reagents (three of 21) or as S-nitrosylated. Cys residues subject to pO2-coupled oxidation are distributed widely within the cytoplasmic domain of RyR1 in multiple functional domains implicated in RyR1 activity-regulating interactions with the L-type Ca(2+) channel (dihydropyridine receptor) and FK506-binding protein 12 as well as in "hot spot" regions containing sites of mutation implicated in malignant hyperthermia and central core disease. pO2-coupled disulfide formation was identified, whereas neither S-glutathionylated nor sulfenamide-modified Cys residues were observed. Thus, physiological redox regulation of RyR1 by endogenously generated hydrogen peroxide is exerted through dynamic disulfide formation involving multiple Cys residues.

Full Text

Duke Authors

Cited Authors

  • Sun, Q-A; Wang, B; Miyagi, M; Hess, DT; Stamler, JS

Published Date

  • August 9, 2013

Published In

Volume / Issue

  • 288 / 32

Start / End Page

  • 22961 - 22971

PubMed ID

  • 23798702

Pubmed Central ID

  • 23798702

Electronic International Standard Serial Number (EISSN)

  • 1083-351X

Digital Object Identifier (DOI)

  • 10.1074/jbc.M113.480228


  • eng

Conference Location

  • United States