Chunhong Gong | PubMed Health

PubMed2005

VEGF treatment induces signaling pathways that regulate both actin polymerization and depolymerization.

The angiogenic growth factor vascular endothelial growth factor (VEGF) enhances endothelial cell migration through the activation of multiple signaling transduction pathways. Actin reorganization is an important component in VEGF-induced migration, yet the signaling pathways mediating this process remain unclear. Actin reorganization involves both actin polymerization and depolymerization, and in this study we demonstrate that VEGF-treatment regulates both of these activities. With respect to actin polymerization, our results indicate that the actin nucleation promoting factors (NPF) neural Wiskott-Aldrich syndrome protein (N-WASP) binds the SH2- plus SH3-domain containing adaptor protein Nck in both control and VEGF-treated cells. We had previously showed that VEGF treatment leads to the recruitment of Nck to activated receptor, and our current results indicate a VEGF-dependent redistribution of N-WASP to the cell surface. A Nck dominant-negative blocked Nck recruitment to receptor, blocked N-WASP cellular redistribution and attenuated actin stress fiber formation. With respect to actin depolymerization, VEGF-treatment led to the rapid phosphorylation of the actin depolymerization factor cofilin, and its upstream regulator, LIM-kinase (LIMK). Unlike what is observed in certain other cell types, the p21-activated kinase (PAK), a Nck binding protein, does not mediate VEGF-induced LIMK phosphorylation, as a PAK dominant-negative had no effect on this activity. The PAK dominant-negative also did not affect VEGF-induced actin reorganization. Pharmacological inhibitors of phosphoinositide-3 kinase (PI3-K) and the rho-activated kinase (ROCK) attenuated VEGF-induced LIMK phosphorylation, indicating a role for (PI3-K) and ROCK in the signaling pathways leading to regulation of LIMK activity.

Actins↗

PubMed2004

Nck and Crk mediate distinct VEGF-induced signaling pathways that serve overlapping functions in focal adhesion turnover and integrin activation.

We have asked whether the Nck and Crk adaptor proteins play important roles in the vascular endothelial growth factor (VEGF)-induced signaling pathways that lead to an enhancement in cell migration. The introduction into human umbilical vein endothelial cells of a dominant-negative inhibitor for either Nck or Crk blocked the recruitment of both endogenous proteins to the KDR VEGF receptor subtype indicating that both proteins are recruited to the same docking site. The Nck and Crk dominant-negatives led to the formation of abnormally large focal adhesion, blocked VEGF-induced integrin activation, and blocked VEGF-induced actin dynamics. The dominant-negatives had no effects on these properties in cells expressing constitutively active Rac1 or RhoA. Since a DN to either Nck or Crk blocks the cellular responses mediated by both proteins, we performed experiments directed at clarifying signaling pathways specifically mediated by each protein. Inhibition of the interaction between Nck with its downstream effector PAK led to abnormally large focal adhesions, but had no effect on integrin activation or cell adhesiveness. Evidence is presented that Crk complexes with C3G in control cells, and VEGF treatment leads to the recruitment of the complex to the cell surface. Inhibition of the C3G downstream effector Rap1 leads to enlarged focal adhesions and blocks VEGF-induced integrin activation. We conclude that Nck and Crk mediate distinct VEGF-induced signaling pathways that serve overlapping functions in cell migration.

Adaptor Proteins, Signal Transducing↗