Luminal Ca2+ regulation of single cardiac ryanodine receptors: insights provided by calsequestrin and its mutants.
The luminal Ca2+ regulation of cardiac ryanodine receptor (RyR2) was explored at the single channel level. The luminal Ca2+ and Mg2+ sensitivity of single CSQ2-stripped and CSQ2-associated RyR2 channels was defined. Action of wild-type CSQ2 and of two mutant CSQ2s (R33Q and L167H) was also compared. Two luminal Ca2+ regulatory mechanism(s) were identified. One is a RyR2-resident mechanism that is CSQ2 independent and does not distinguish between luminal Ca2+ and Mg2+. This mechanism modulates the maximal efficacy of cytosolic Ca2+ activation. The second luminal Ca2+ regulatory mechanism is CSQ2 dependent and distinguishes between luminal Ca2+ and Mg2+. It does not depend on CSQ2 oligomerization or CSQ2 monomer Ca2+ binding affinity. The key Ca2+-sensitive step in this mechanism may be the Ca2+-dependent CSQ2 interaction with triadin. The CSQ2-dependent mechanism alters the cytosolic Ca2+ sensitivity of the channel. The R33Q CSQ2 mutant can participate in luminal RyR2 Ca2+ regulation but less effectively than wild-type (WT) CSQ2. CSQ2-L167H does not participate in luminal RyR2 Ca2+ regulation. The disparate actions of these two catecholaminergic polymorphic ventricular tachycardia (CPVT)-linked mutants implies that either alteration or elimination of CSQ2-dependent luminal RyR2 regulation can generate the CPVT phenotype. We propose that the RyR2-resident, CSQ2-independent luminal Ca2+ mechanism may assure that all channels respond robustly to large (>5 muM) local cytosolic Ca2+ stimuli, whereas the CSQ2-dependent mechanism may help close RyR2 channels after luminal Ca2+ falls below approximately 0.5 mM.
Qin
Jia
J
Department of Molecular Physiology and Biophysics, Rush University Medical Center, Chicago, IL 60612, USA.
Valle
Giorgia
G
Nani
Alma
A
Nori
Alessandra
A
Rizzi
Nicoletta
N
Priori
Silvia G
SG
Volpe
Pompeo
P
Fill
Michael
M
D019943:Amino Acid Substitution; D000818:Animals; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D020013:Calcium Signaling; D002155:Calsequestrin / Q000737:chemistry / Q000378:metabolism*; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D019281:Dimerization; D004553:Electric Conductivity; D015640:Ion Channel Gating / Q000502:physiology; D008051:Lipid Bilayers; D008274:Magnesium / Q000737:chemistry / Q000378:metabolism; D008861:Microsomes; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D018408:Patch-Clamp Techniques; D051381:Rats; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry* / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D013329:Structure-Activity Relationship; D017180:Tachycardia, Ventricular / Q000209:etiology / Q000503:physiopathology
D002155:Calsequestrin; D008051:Lipid Bilayers; D050505:Mutant Proteins; D019837:Ryanodine Receptor Calcium Release Channel; D008274:Magnesium; D002118:Calcium