TGFβ-induced Early Activation of the Small GTPase RhoA Is Smad2/3-independent and Involves Src and the Guanine Nucleotide Exchange Factor Vav2

Cell Physiol Biochem. 2011;28(2):229-38. doi: 10.1159/000331734. Epub 2011 Aug 16.

Abstract

TGFβ has been shown to induce short- and long-term actin reorganization controlled by Rho-GTPase signaling. A number of direct Smad target genes, rapidly activated by TGFβ, have been previously reported to control the long-term Rho activation and actin reorganization. However, the molecular mechanisms that regulate the prompt stimulation of Rho GTPases by TGFβ remain unknown. In the present study we report that TGFβ rapidly stimulated RhoA and RhoB activation in JEG3 choriocarcinoma cells that lack endogenous Smad3. Inhibition of Smad2 expression via siRNA-mediated silencing or by blocking its phosphorylation using the TβRI inhibitor SB431542 did not prevent the early RhoA/B activation by TGFβ indicating that this effect is Smad2/3-independent. Pre-treatment of the cells with the general tyrosine kinase inhibitor Genistein blocked the TGFβ-induced early RhoA activation. In line with this finding, TGFβ-stimulation resulted in a quick activation of the non-receptor tyrosine kinase Src, followed by activation of the guanine nucleotide exchange factor (GEF) Vav2. Inhibition of Src kinase by the selective inhibitor of the Src family tyrosine kinases PP2 totally blocked the early TGFβ-induced RhoA activation. Similarly, Vav2 silencing via siRNA reduced the TGFβ-induced RhoA activation implying that the rapid Src/Vav2 stimulation was effective in regulating RhoA activation. Our present findings provide for the first time a clear evidence for the role of Src and Vav2-GEF in the early Smad2/3-independent Rho activation by TGFβ.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Benzamides / pharmacology
  • Cell Line, Tumor
  • Dioxoles / pharmacology
  • Enzyme Activation / drug effects
  • Genistein / pharmacology
  • Humans
  • Protein Kinase Inhibitors / pharmacology
  • Proto-Oncogene Proteins c-vav / antagonists & inhibitors
  • Proto-Oncogene Proteins c-vav / genetics
  • Proto-Oncogene Proteins c-vav / metabolism*
  • RNA Interference
  • RNA, Small Interfering / metabolism
  • Receptors, Transforming Growth Factor beta / antagonists & inhibitors
  • Receptors, Transforming Growth Factor beta / metabolism
  • Smad2 Protein / antagonists & inhibitors
  • Smad2 Protein / genetics
  • Smad2 Protein / metabolism*
  • Smad3 Protein / metabolism*
  • Transforming Growth Factor beta / pharmacology*
  • rhoA GTP-Binding Protein / metabolism*
  • rhoB GTP-Binding Protein / metabolism
  • src-Family Kinases / metabolism*

Substances

  • 4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide
  • Benzamides
  • Dioxoles
  • Protein Kinase Inhibitors
  • Proto-Oncogene Proteins c-vav
  • RNA, Small Interfering
  • Receptors, Transforming Growth Factor beta
  • Smad2 Protein
  • Smad3 Protein
  • Transforming Growth Factor beta
  • Genistein
  • src-Family Kinases
  • rhoA GTP-Binding Protein
  • rhoB GTP-Binding Protein