RapV12 antagonizes Ras‐dependent activation of ERK1 and ERK2 by LPA and EGF in Rat‐1 fibroblasts.

Rap1 is a small Ras‐related GTPase which when over‐expressed is able to revert transformation by Ki‐Ras. We have investigated the role of Rap1 in regulating ‘normal’ Ras function by studying the activation of the mitogen‐activated protein (MAP) kinases ERK1 and ERK2 by two fundamentally different growth factors, epidermal growth factor (EGF) and 1‐oleoyl‐lyso‐phosphatidic acid (LPA). Conditional expression of RasN17 (a dominant‐negative mutant) in Rat‐1 cells inhibited activation of MAP kinases by EGF and also LPA, the first time a defined G‐protein‐coupled receptor mitogen has been shown to require Ras to exert its effects. Conditional or constitutive expression of even low levels of RapV12 (a mutant insensitive to Rap‐GAP) attenuated activation of MAP kinases by EGF and LPA, but did not interfere with growth factor‐stimulated increases in Ras‐GTP, indicating that signalling from receptors to Ras was not impaired. Inhibition of Ras‐mediated signalling with either RasN17 or RapV12 attenuated DNA synthesis by EGF and LPA. We conclude that receptor tyrosine kinases and G‐protein‐coupled receptors use Ras as a common step in signalling to MAP kinases and that Rap‐GTP (RapV12) at physiological levels interferes with downstream signalling from Ras to MAP kinases in vivo.