RACK1 Regulates VEGF/Flt1-mediated Cell Migration via Activation of a PI3K/Akt Pathway

Feng Wang,Mai Yamauchi,Masashi Muramatsu,Tsuyoshi Osawa,Rika Tsuchida,Masabumi Shibuya

doi:10.1074/jbc.m110.165605

Abstract

Vascular endothelial growth factor (VEGF) is vital to physiological as well as pathological angiogenesis, and regulates a variety of cellular functions, largely by activating its 2 receptors, fms-like tyrosine kinase (Flt1) and kinase domain receptor (KDR). KDR plays a critical role in the proliferation of endothelial cells by controlling VEGF-induced phospholipase Cγ-protein kinase C (PLCγ-PKC) signaling. The function of Flt1, however, remains to be clarified. Recent evidence has indicated that Flt1 regulates the VEGF-triggered migration of endothelial cells and macrophages. Here, we show that RACK1, a ubiquitously expressed scaffolding protein, functions as an important regulator of this process. We found that RACK1 (receptor for activated protein kinase C 1) binds to Flt1 in vitro. When the endogenous expression of RACK1 was attenuated by RNA interference, the VEGF-driven migration was remarkably suppressed whereas the proliferation was unaffected in a stable Flt1-expressing cell line, AG1-G1-Flt1. Further, we demonstrated that the VEGF/Flt-mediated migration of AG1-G1-Flt1 cells occurred mainly via the activation of the PI3 kinase (PI3K)/Akt and Rac1 pathways, and that RACK1 plays a crucial regulatory role in promoting PI3K/Akt-Rac1 activation.

Highlights

Angiogenesis is a critical and complicated process in embryonic vascular development, wound healing, and organ regeneration as well as in multiple pathological conditions, such as rheumatoid arthritis and tumor growth [1]
The phosphorylation of Tyr(Y)-1175 on kinase domain receptor (KDR) leads to the activation of phospholipase C (PLC)␥, which in turn promotes the intracellular mobilization of calcium and activates a crucial protein kinase C-Raf-mitogen-activated protein kinase (PKC-Raf-MAPK) cascade, the latter regulating endothelial cell proliferation (14 –16)
When the endogenous expression of RACK1 was attenuated by RNA interference (RNAi) in a stable Flt1-expressing cell line, the vascular endothelial growth factor (VEGF)-induced migration was remarkably suppressed whereas the proliferation was not affected

Summary

Introduction

Angiogenesis is a critical and complicated process in embryonic vascular development, wound healing, and organ regeneration as well as in multiple pathological conditions, such as rheumatoid arthritis and tumor growth [1]. KDR plays a critical role in the proliferation of endothelial cells by controlling VEGF-induced phospholipase C␥-protein kinase C (PLC␥-PKC) signaling. We demonstrated that the VEGF/Flt-mediated migration of AG1-G1-Flt1 cells occurred mainly via the activation of the PI3 kinase (PI3K)/ Akt and Rac1 pathways, and that RACK1 plays a crucial regulatory role in promoting PI3K/Akt-Rac1 activation.

Results

Conclusion