FGF2 induces ERK phosphorylation through Grb2 and PKC during quiescent myogenic cell activation.
Satellite cells are muscle-resident stem cells, which are located beneath the basement membrane of myofibers. Because the number of satellite cells is normally constant, there must be a tight regulation of satellite cell activation and self-renewal. However, the molecular mechanisms involved in satellite cell maintenance are largely unknown, and thus have become the subject of extensive study these days. Although RNA interference with a small interfering RNA has been widely used to investigate the role of specific gene products, inefficient knockdown of Grb2 expression occurred in quiescent reserve cells, a model for quiescent satellite cells, by ordinary transfection protocol. In this study we report that pretreatment with trypsin greatly enhanced siRNA delivery into quiescent reserve cells, resulting in efficient silencing of Grb2 expression. By applying a combination of Grb2-silencing and protein kinase C inhibitors, we demonstrated that extracellular signal-regulated kinase (ERK) phosphorylation induced with fibroblast growth factor 2 (FGF2) was dependent on both Grb2 and protein kinase C (PKC) with different kinetics. We concluded that the PKC-mediated pathway contributes to rapid initiation and termination of ERK phosphorylation, while the Grb2-mediated pathway contributes to delayed and sustained ERK phosphorylation.
Nagata
Yosuke
Y
The University of Tokyo, Japan.
Honda
Yusuke
Y
Matsuda
Ryoichi
R
D000818:Animals; D045744:Cell Line, Tumor; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology*; D051380:GRB2 Adaptor Protein / Q000235:genetics / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D051379:Mice; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D032496:Satellite Cells, Skeletal Muscle / Q000378:metabolism*; D015398:Signal Transduction
D051380:GRB2 Adaptor Protein; D034741:RNA, Small Interfering; D016222:Fibroblast Growth Factor 2; D011493:Protein Kinase C; D048049:Extracellular Signal-Regulated MAP Kinases