Sphingosylphosphorylcholine activation of mitogen-activated protein kinase in Swiss 3T3 cells requires protein kinase C and a pertussis toxin-sensitive G protein

J Biol Chem. 1995 Oct 13;270(41):24334-42. doi: 10.1074/jbc.270.41.24334.


Sphingosylphosphorylcholine (SPC) is a potent mitogen for Swiss 3T3 cells, but the signaling mechanisms involved are poorly characterized. Here, we report that addition of SPC induces a rapid and transient activation of p42 mitogen-activated protein kinase (p42MAPK) in these cells. SPC-induced p42MAPK activation peaked at 5 min and was undetectable after 30 min of incubation with SPC. The effect of SPC on p42MAPK activation was comparable to that induced by bombesin and platelet-derived growth factor. As SPC strongly induced phosphorylation of the major protein kinase C (PKC) substrate 80K/MARCKS in either intact or permeabilized cells, we examined whether PKC could be involved in SPC-induced p42MAPK activation. Here, we demonstrate that p42MAPK activation by SPC was dependent on PKC activity as shown by inhibition of PKC with the bisindolymaleimide GF 109203X or down-regulation of PKC by prolonged treatment of Swiss 3T3 cells with phorbol esters. Activation of both PKC and p42MAPK by SPC was markedly inhibited by treatment with pertussis toxin, implicating a G proteins(s) of the Gi/G(o) subfamily in the action of SPC. SPC-induced rapid activation of a downstream target of p42MAPK, p90 ribosomal S6 kinase (p90rsk), also required PKC and a pertussis toxin-sensitive G protein. In addition, SPC-induced mitogenesis was dependent on a Gi protein in Swiss 3T3 cells. SPC also induced p42MAPK activation and DNA synthesis in secondary cultures of mouse embryo fibroblasts through a pertussis toxin-sensitive pathway. As G proteins link many cell surface receptors to effector proteins, we hypothesize, therefore, that SPC could bind to a receptor that mediates at least some of its biological effects in Swiss 3T3 cells and mouse embryo fibroblasts.

MeSH terms

  • 3T3 Cells
  • Amino Acid Sequence
  • Animals
  • Antibodies
  • Blotting, Western
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
  • Cell Division / drug effects
  • Cytochalasin D / pharmacology
  • DNA / biosynthesis
  • DNA Replication / drug effects
  • Egtazic Acid / pharmacology
  • Enzyme Activation
  • GTP-Binding Proteins / metabolism*
  • Intracellular Signaling Peptides and Proteins*
  • Kinetics
  • Membrane Proteins*
  • Mice
  • Molecular Sequence Data
  • Myristoylated Alanine-Rich C Kinase Substrate
  • Peptide Fragments / chemical synthesis
  • Peptide Fragments / chemistry
  • Peptide Fragments / immunology
  • Pertussis Toxin*
  • Phorbol 12,13-Dibutyrate / pharmacology
  • Phosphorylation
  • Phosphorylcholine / analogs & derivatives*
  • Phosphorylcholine / pharmacology
  • Protein Kinase C / metabolism*
  • Proteins / isolation & purification
  • Proteins / metabolism*
  • Sphingosine / analogs & derivatives*
  • Sphingosine / pharmacology
  • Thymidine / metabolism
  • Virulence Factors, Bordetella / pharmacology*


  • Antibodies
  • Intracellular Signaling Peptides and Proteins
  • Marcks protein, mouse
  • Membrane Proteins
  • Peptide Fragments
  • Proteins
  • Virulence Factors, Bordetella
  • sphingosine phosphorylcholine
  • Phosphorylcholine
  • Myristoylated Alanine-Rich C Kinase Substrate
  • Cytochalasin D
  • Phorbol 12,13-Dibutyrate
  • Egtazic Acid
  • DNA
  • Pertussis Toxin
  • Protein Kinase C
  • Calcium-Calmodulin-Dependent Protein Kinases
  • GTP-Binding Proteins
  • Sphingosine
  • Thymidine