Insertion of the full-length calcium channel alpha(1S) subunit into triads of skeletal muscle in vitro.
A full-length and a C-terminally truncated form of the calcium channel alpha(1S) subunit can be isolated from skeletal muscle. Here we studied whether full-length alpha(1S) is functionally incorporated into the skeletal muscle excitation-contraction coupling apparatus. A fusion protein of alpha(1S) with the green fluorescent protein attached to its C-terminus (alpha(1S)-GFP) or alpha(1S) and GFP separately (alpha(1S)+GFP) were expressed in dysgenic myotubes, which lack endogenous alpha(1S). Full-length alpha(1S)-GFP was targeted into triad junctions and restored calcium currents and excitation-contraction coupling. GFP remained colocalized with alpha(1S), indicating that intact alpha(1S)-GFP was inserted into triads and that the C-terminus remained associated with the excitation-contraction coupling apparatus.
Flucher
B E
BE
Department of Physiology and Department of Biochemical Pharmacology, University of Innsbruck, Fritz-Pregl-Str. 3, A-6020, Innsbruck, Austria. bernhard.e.flucher@uibk.ac.at
Kasielke
N
N
Gerster
U
U
Neuhuber
B
B
Grabner
M
M
D020746:Calcium Channels, L-Type / Q000172:deficiency / Q000235:genetics* / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004553:Electric Conductivity; D004558:Electric Stimulation; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D008870:Microtubules / Q000378:metabolism; D009119:Muscle Contraction; D018482:Muscle, Skeletal / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D014162:Transfection
D020746:Calcium Channels, L-Type; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins