Altered stored calcium release in skeletal myotubes deficient of triadin and junctin.
Triadin and junctin are integral sarcoplasmic reticulum membrane proteins that form a macromolecular complex with the skeletal muscle ryanodine receptor (RyR1) but their roles in skeletal muscle calcium homeostasis remain incompletely understood. Here we report that delivery of siRNAs specific for triadin or junctin into C2C12 skeletal myoblasts reduced the expression of triadin and junctin in 8-day-old myotubes by 80 and 100%, respectively. Knocking down either triadin or junctin in these cells reduced Ca2+ release induced by depolarization (10mM KCl) by 20-25%. Unlike triadin knockdown myotubes, junctin knockdown and junctin/triadin double knockdown myotubes also had reduced Ca2+ release induced by 400 microM 4-chloro-m-cresol, 10mM caffeine, 400 microM UTP, or 1 microM thapsigargin. Thus, knocking down junctin compromised the Ca2+ stores in the sarcoplasmic reticulum of these cells. Our subsequent studies showed that in junctin knockdown myotubes at least two sarcoplasmic reticulum proteins (RyR1 and skeletal muscle calsequestrin) were down-regulated while these proteins' mRNA expression was not affected. The results suggest that triadin has a role in facilitating KCl depolarization-induced Ca2+ release in contrast to junctin which has a role in maintaining sarcoplasmic reticulum Ca2+ store size in C2C12 myotubes.
Wang
Ying
Y
Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC 27599-7260, United States.
Li
Xinghai
X
Duan
Hongzhe
H
Fulton
Timothy R
TR
Eu
Jerry P
JP
Meissner
Gerhard
G
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000235:genetics; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002155:Calsequestrin / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D025461:Feedback, Physiological; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D032448:Myoblasts, Skeletal / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009467:Neuromuscular Depolarizing Agents / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism; D014161:Transduction, Genetic
D002135:Calcium-Binding Proteins; D002155:Calsequestrin; D002352:Carrier Proteins; D008565:Membrane Proteins; D009124:Muscle Proteins; D009467:Neuromuscular Depolarizing Agents; D034741:RNA, Small Interfering; D019837:Ryanodine Receptor Calcium Release Channel; C070048:triadin; C498841:Asph protein, mouse; D006899:Mixed Function Oxygenases; D002118:Calcium