Insulin-like growth factor-induced transcriptional activity of the skeletal alpha-actin gene is regulated by signaling mechanisms linked to voltage-gated calcium channels during myoblast differentiation.
IGF-I activates signaling pathways that increase the expression of muscle-specific genes in differentiating myoblasts. Induction of skeletal alpha-actin expression occurs during differentiation through unknown mechanisms. The purpose of this investigation was to examine the mechanisms that IGF-I uses to induce skeletal alpha-actin gene expression in C2C12 myoblasts. IGF-I increased skeletal alpha-actin promoter activity by 107% compared with the control condition. Ni(+) [T-type voltage-gated Ca(2+) channel (VGCC) inhibitor] reduced basal-induced activation of the skeletal alpha-actin promoter by approximately 84%, and nifedipine (L-type VGCC inhibitor) inhibited IGF-I-induced activation of the skeletal alpha-actin promoter by 29-48%. IGF-I failed to increase skeletal alpha-actin promoter activity in differentiating dysgenic (lack functional L-type VGCC) myoblasts; 30 mm K(+) and 30 mm K(+)+IGF-I increased skeletal alpha-actin promoter activity by 162% and 76% compared with non-IGF-I or IGF-I-only conditions, respectively. IGF-I increased calcineurin activity, which was inhibited by cyclosporine A. Further, cyclosporine A inhibited K(+)+IGF-I-induced activation of the skeletal alpha-actin promoter. Constitutively active calcineurin increased skeletal alpha-actin promoter activity by 154% and rescued the nifedipine-induced inhibition of L-type VGCC but failed to rescue the Ni(+)-inhibition of T-type VGCC. IGF-I-induced nuclear factor of activated T-cells transcriptional activity was not inhibited by nifedipine or Ni(+). IGF-I failed to increase serum response factor transcriptional activity; however, serum response factor activity was reduced in the presence of Ni(+). These data suggest that IGF-I-induced activation of the skeletal alpha-actin promoter is regulated by the L-type VGCC and calcineurin but independent of nuclear factor of activated T-cell transcriptional activity as C2C12 myoblasts differentiate into myotubes.
Spangenburg
Espen E
EE
Department of Biomedical Sciences, University of Missouri, Columbia 65211, USA. eespangenburg@ucdavis.edu
Bowles
Douglas K
DK
Booth
Frank W
FW
D000199:Actins / Q000235:genetics*; D000818:Animals; D001842:Bone and Bones / Q000378:metabolism*; D019703:Calcineurin / Q000502:physiology; D015220:Calcium Channels / Q000378:metabolism* / Q000502:physiology; D002454:Cell Differentiation; D002460:Cell Line; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology / Q000502:physiology*; D051379:Mice; D032446:Myoblasts / Q000166:cytology*; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000502:physiology; D011994:Recombinant Proteins / Q000494:pharmacology; D026362:Serum Response Factor / Q000235:genetics; D015398:Signal Transduction / Q000502:physiology*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology*
D000199:Actins; D015220:Calcium Channels; D011994:Recombinant Proteins; D026362:Serum Response Factor; D007334:Insulin-Like Growth Factor I; D019703:Calcineurin