Characterization of a calcium-regulation domain of the skeletal-muscle ryanodine receptor.
A negatively charged region of the N-terminal portion of the skeletal ryanodine receptor (RyR), located between residues 1872-1923, is involved in Ca (2+)-dependent regulation of the Ca(2+)-release channel. This region is divergent between the skeletal (RyR1) and cardiac (RyR2) isoforms of the channel, and is known as D3. Ca(2+) exerts important regulatory functions on the RyR, being involved in both activation and inactivation functions of the channel, i.e. the effects occurring at micromolar and millimolar Ca(2+) concentrations respectively. To characterize the role of D3 in the Ca(2+)-dependent regulation of the Ca(2+)-release channel, we studied the functional consequences of deleting the D3 region from RyR1 (DeltaD3-RyR1) using a heterologous expression system, [(3)H]ryanodine binding assays and single-channel recordings in lipid bilayers. Deletion of the D3 region selectively affected Ca(2+)-dependent regulation of RyR1, but did not alter [(3)H]ryanodine binding or the effect of other modulators on the RyR. Compared with full-length RyR1 (wt-RyR1), the Ca(2+)-dependence curve of DeltaD3-RyR1 is broader, reflecting increased sensitivity to Ca(2+) activation and decreased sensitivity to Ca(2+) inactivation. In addition, DeltaD3-RyR1 was more resistant to inhibition by Mg(2+). Comparison of the effect of caffeine on wt-RyR1 and DeltaD3-RyR1 suggested that D3 is an important region of RyR that participates in Ca(2+)-dependent activation and inactivation of the Ca(2+)-release channel.
Hayek
S M
SM
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, 10900 Euclid Ave, Cleveland, OH 44106, USA.
Zhu
X
X
Bhat
M B
MB
Zhao
J
J
Takeshima
H
H
Valdivia
H H
HH
Ma
J
J
D000818:Animals; D016466:CHO Cells; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology*; D015220:Calcium Channels / Q000378:metabolism; D006224:Cricetinae; D015640:Ion Channel Gating / Q000187:drug effects; D008051:Lipid Bilayers / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects; D018482:Muscle, Skeletal / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D012433:Ryanodine / Q000378:metabolism; D019837:Ryanodine Receptor Calcium Release Channel / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017384:Sequence Deletion / Q000235:genetics
D015220:Calcium Channels; D008051:Lipid Bilayers; D019837:Ryanodine Receptor Calcium Release Channel; D012433:Ryanodine; D002110:Caffeine; D008274:Magnesium; D002118:Calcium