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

EMBO J. 1993 Sep;12(9):3475-85.

Abstract

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.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Blotting, Western
  • Calcium-Calmodulin-Dependent Protein Kinases / antagonists & inhibitors
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
  • Cell Line
  • Enzyme Activation
  • Epidermal Growth Factor / pharmacology*
  • Fibroblasts / drug effects
  • Fibroblasts / metabolism
  • GTP-Binding Proteins / metabolism*
  • Genes, ras*
  • Kinetics
  • Lysophospholipids / pharmacology*
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3
  • Mitogen-Activated Protein Kinases*
  • Models, Biological
  • Molecular Sequence Data
  • Peptides / chemical synthesis
  • Peptides / immunology
  • Proto-Oncogene Proteins / metabolism
  • Rats
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / metabolism
  • Signal Transduction
  • rap GTP-Binding Proteins

Substances

  • Lysophospholipids
  • Peptides
  • Proto-Oncogene Proteins
  • Recombinant Proteins
  • Epidermal Growth Factor
  • Calcium-Calmodulin-Dependent Protein Kinases
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3
  • Mitogen-Activated Protein Kinases
  • GTP-Binding Proteins
  • rap GTP-Binding Proteins
  • lysophosphatidic acid