RAS IS REQUIRED FOR EPIDERMAL GROWTH FACTOR-STIMULATED ARACHIDONIC-ACID RELEASE IN RAT-1 FIBROBLASTS

Previous studies have provided suggestive evidence for an interaction between ras activation and signalling pathways involved in agonist-sti mulated arachidonic acid release in a variety of cell systems. In orde r to clarify this interaction, we have measured epidermal growth facto r (EGF)-stimulated arachidonic acid release in rat-1 fibroblasts trans fected with the N-17 dominant negative mutation of ras. Cells transfec ted with the N-17 ras mutant, display a markedly attenuated arachidoni c acid-release response to EGF, compared to sham-transfected and non-t ransfected cells. In contrast, the response to phorbol myristate aceta te (PMA) was not attenuated in the N-17-mutant expressing cells. No di fferences were detected between sham-transfected and N-17 mutant expre ssing cells in levels of immunodetectable EGF receptor, cytosolic phos pholipase A(2) or mitogen-activated protein (MAP) kinase. Attenuation of EGF-stimulated arachidonic acid release in the N-17 mutant expressi ng cells, was accompanied by a marked diminution in EGF-stimulated tyr osine phosphorylation of MAP kinase. We conclude that the signalling p athway involved in epidermal growth factor-stimulated arachidonic acid release is similar to the signalling pathway for mitogenic responses to epidermal growth factor and requires ras activation, likely followe d by a downstream cascade of kinases eventuating in MAP kinase activat ion.