NIH-3T3 cells transformed by the EJ-ras oncogene exhibit reduced platelet-derived growth factor-mediated Ca2+ mobilization.
NIH-3T3 cells transformed by the EJ-ras oncogene synthesize only 10-15% as much inositol 1,4,5-trisphosphate (InsP3) as control cells after stimulation with platelet-derived growth factor (PDGF). This is despite the fact that the basal (unstimulated) levels of InsP3 synthesized in control and EJ-ras-transformed cells are not significantly different. Using the fluorescent indicator fura-2 and digital-imaging techniques, we have visualized and quantified changes in intracellular Ca2+ concentrations in control and EJ-ras-transformed NIH-3T3 cells in response to PDGF. Within 3 min after exposure of control cells to PDGF, intracellular Ca2+ levels are increased 3- to 9-fold, paralleling the increase in InsP3. In contrast, the majority (greater than 90%) of the EJ-ras-transformed cells show no increase in Ca2+ levels after PDGF exposure and the few that did respond exhibited only a small transient increase. Pronounced differences in the intracellular localization of Ca2+ increases in control and the responding EJ-ras-transformed cells were also observed. Despite the inhibition of InsP3 synthesis and subsequent Ca2+ mobilization, the EJ-ras-transformed cells respond mitogenically to PDGF. These data do not support the hypothesis that the EJ-ras gene product (p21) stimulates a phosphatidylinositol 4,5-bisphosphate-specific phospholipase C in NIH-3T3 cells; instead they suggest that the EJ-ras p21 may uncouple the PDGF receptor from phospholipase C resulting in inhibition of PDGF-stimulated activity of phospholipase C, InsP3 synthesis, and Ca2+ mobilization.
Benjamin
C W
CW
Department of Cell Biology, Upjohn Company, Kalamazoo, MI 49001.
Connor
J A
JA
Tarpley
W G
WG
Gorman
R R
RR
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002461:Cell Line, Transformed / Q000187:drug effects; D002471:Cell Transformation, Neoplastic / Q000378:metabolism*; D002478:Cells, Cultured; D011905:Genes, ras; D007294:Inositol / Q000378:metabolism; D007295:Inositol Phosphates / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D014162:Transfection
D007295:Inositol Phosphates; D010982:Platelet-Derived Growth Factor; D007294:Inositol; D002118:Calcium