Functional loss of DHRS7C induces intracellular Ca2+ overload and myotube enlargement in C2C12 cells via calpain activation.
Dehydrogenase/reductase member 7C (DHRS7C) is a newly identified NAD/NADH-dependent dehydrogenase that is expressed in cardiac and skeletal muscle and localized in the endoplasmic/sarcoplasmic reticulum (ER/SR). However, its functional role in muscle cells remains to be fully elucidated. Here, we investigated the role of DHRS7C by analyzing mouse C2C12 myoblasts deficient in DHRS7C (DHRS7C-KO cells), overexpressing wild-type DHRS7C (DHRS7C-WT cells), or expressing mutant DHRS7C [DHRS7C-Y191F or DHRS7C-K195Q cells, harboring point mutations in the NAD/NADH-dependent dehydrogenase catalytic core domain (YXXXK)]. DHRS7C expression was induced as C2C12 myoblasts differentiated into mature myotubes, whereas DHRS7C-KO myotubes exhibited enlarged cellular morphology after differentiation. Notably, both DHRS7C-Y191F and DHRS7C-K195Q cells also showed similar enlarged cellular morphology, suggesting that the NAD/NADH-dependent dehydrogenase catalytic core domain is pivotal for DHRS7C function. In DHRS7C-KO, DHRS7C-Y191F, and DHRS7C-K195Q cells, the resting level of cytosolic Ca2+and total amount of Ca2+storage in the ER/SR were significantly higher than those in control C2C12 and DHRS7C-WT cells after differentiation. Additionally, Ca2+release from the ER/SR induced by thapsigargin and 4-chloro-m-cresol was augmented in these cells and calpain, a calcium-dependent protease, was significantly activated in DHRS7C-KO, DHRS7C-Y191F, and DHRS7C-K195Q myotubes, consistent with the higher resting level of cytosolic Ca2+concentration and enlarged morphology after differentiation. Furthermore, treatment with a calpain inhibitor abolished the enlarged cellular morphology. Taken together, our findings suggested that DHRS7C maintains intracellular Ca2+homeostasis involving the ER/SR and that functional loss of DHRS7C leads to Ca2+overload in the cytosol and ER/SR, resulting in enlarged cellular morphology via calpain activation.
Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Shinobu
S
Arai
Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Masataka
M
Ikeda
Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan; tomomi_i@cardiol.med.kyushu-u.ac.jp.
Tomomi
T
Ide
Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Yuka
Y
Matsuo
Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Takeo
T
Fujino
Department of Cardiovascular Physiology Faculty of Medicine, Kagawa University, Kagawa, Japan; and.
Katsuya
K
Hirano
Department of Therapeutic Regulation of Cardiovascular Homeostasis, Center for Disruptive Cardiovascular Medicine, Kyushu University, Fukuoka, Japan.
Kenji
K
Sunagawa
Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka, Japan.
Hiroyuki
H
Tsutsui
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology; D002154:Calpain / Q000378:metabolism*; D002460:Cell Line; D048429:Cell Size; D051379:Mice; D018485:Muscle Fibers, Skeletal / Q000166:cytology* / Q000378:metabolism*; D010088:Oxidoreductases / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D015854:Up-Regulation / Q000502:physiology*
C000619980:DHRS7C protein, mouse; D010088:Oxidoreductases; D002154:Calpain; D002118:Calcium
calcium homeostasis; calpain; dehydrogenase/reductase member 7C; endoplasmic reticulum; morphogenesis