RyR1/RyR3 chimeras reveal that multiple domains of RyR1 are involved in skeletal-type E-C coupling.
Skeletal-type E-C coupling is thought to require a direct interaction between RyR1 and the alpha(1S)-DHPR. Most available evidence suggests that the cytoplasmic II-III loop of the dihydropyridine receptor (DHPR) is the primary source of the orthograde signal. However, identification of the region(s) of RyR1 involved in bidirectional signaling with the alpha(1S)-DHPR remains elusive. To identify these regions we have designed a series of chimeric RyR cDNAs in which different segments of RyR1 were inserted into the corresponding region of RyR3 and expressed in dyspedic 1B5 myotubes. RyR3 provides a preferable background than RyR2 for defining domains essential for E-C coupling because it possesses less sequence homology to RyR1 than the RyR2 backbone used in previous studies. Our data show that two regions of RyR1 (chimera Ch-10 aa 1681-2641 and Ch-9 aa 2642-3770), were independently able to restore skeletal-type E-C coupling to RyR3. These two regions were further mapped and the critical RyR1 residues were 1924-2446 (Ch-21) and 2644-3223 (Ch-19). These results both support and refine the previous hypothesis that multiple domains of RyR1 combine to functionally interact with the DHPR during E-C coupling.
Perez
Claudio F
CF
Department of Anesthesiology, Perioperative and Pain Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA. cperez@zeus.bwh.harvard.edu
Voss
Andrew
A
Pessah
Isaac N
IN
Allen
Paul D
PD
D000200:Action Potentials / Q000502:physiology; D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D006801:Humans; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D018485:Muscle Fibers, Skeletal / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology*; D018482:Muscle, Skeletal / Q000737:chemistry / Q000187:drug effects / Q000502:physiology; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000145:classification / Q000172:deficiency / Q000502:physiology; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry* / Q000145:classification / Q000172:deficiency / Q000502:physiology*
D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D019837:Ryanodine Receptor Calcium Release Channel; D002110:Caffeine; D002118:Calcium