Dominant negative farnesyltransferase alpha-subunit inhibits insulin mitogenic effects.
Farnesylation of p21Ras is required for translocation to the plasma membrane and subsequent activation by growth factors. Previously we demonstrated that insulin stimulates the phosphorylation of farnesyltransferase (FTase) and its activity, whereby the amount of farnesylated p21Ras anchored at the plasma membrane is increased. Herein we report that substitution of alanine for two serine residues (S60A)(S62A) of the alpha-subunit of FTase creates a dominant negative (DN) mutant. VSMC expressing the FTase alpha-subunit (S60A)(S62A) clone showed a 30% decreased basal FTase activity concurrent with a 15% decrease in the amount of farnesylated p21Ras compared to controls. Expression of alpha-subunit (S60A,S62A) blunted FTase phosphorylation and activity in the presence of hyperinsulinemia, and inhibited insulin-stimulated increases in farnesylated p21Ras. Insulin-stimulated VSMC expressing the FTase alpha-subunit (S60A,S62A) showed decreased (i) phosphorylation of FTase, (ii) FTase activity, (iii) amounts of farnesylated p21Ras, (iv) DNA synthesis, and (v) migration. Thus, down-regulation of FTase activity appears to mitigate the potentially detrimental mitogenic effects of hyperinsulinemia on VSMC.
Solomon
C S
CS
Research Service, Denver VA Medical Center, 1055 Clermont Street, Denver, CO 80220, USA.
Goalstone
M L
ML
D019883:Alkyl and Aryl Transferases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D002455:Cell Division / Q000187:drug effects*; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003001:Cloning, Molecular; D051231:Farnesyltranstransferase; D006946:Hyperinsulinism; D007328:Insulin / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D010710:Phosphates / Q000378:metabolism; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D011106:Polyisoprenyl Phosphates / Q000378:metabolism; D017368:Protein Prenylation; D021122:Protein Subunits; D016283:Proto-Oncogene Proteins p21(ras) / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012717:Sesquiterpenes; D014162:Transfection
D007328:Insulin; D010710:Phosphates; D010982:Platelet-Derived Growth Factor; D011106:Polyisoprenyl Phosphates; D021122:Protein Subunits; D011994:Recombinant Proteins; D012717:Sesquiterpenes; C004808:farnesyl pyrophosphate; D019883:Alkyl and Aryl Transferases; D051231:Farnesyltranstransferase; D016283:Proto-Oncogene Proteins p21(ras)