Rac affects invasion of human renal cell carcinomas by up-regulating tissue inhibitor of metalloproteinases (TIMP)-1 and TIMP-2 expression.

Helmut E. Gabbert,Frits Michiels,John G. Collard,Rainer Engers,Erik Springer

doi:10.1074/jbc.m105049200

Abstract

Rho-like GTPases, including Cdc42, Rac1, and RhoA, regulate distinct actin cytoskeleton changes required for adhesion, migration, and invasion of cells. Tiam1 specifically activates Rac, and earlier studies have demonstrated that Tiam1-Rac signaling affects migration and invasion in a cell type- and cell substrate-specific manner. In the present study, we examined the role of Tiam1-Rac signaling in migration and invasion of human renal cell carcinomas. Stable overexpression of Tiam1 or constitutively active V12-Rac1 in a human renal cell carcinoma cell line (clearCa-28) strongly inhibited cell migration by promoting E-cadherin-mediated cell-cell adhesion. Blocking E-cadherin-mediated adhesion by E-cadherin-specific HAV peptides allowed cells to migrate, but was not sufficient to antagonize Tiam1- and V12-Rac1-induced inhibition of Matrigel invasion, suggesting that Rac may influence invasion also through other mechanisms. Indeed, Tiam1-mediated Rac activation induced transcriptional up-regulation of tissue inhibitor of metalloproteinases-1 (TIMP-1) and post-transcriptional up-regulation of TIMP-2, whereas secretion and activity levels of their counterparts, matrix metalloproteinase-9 and matrix metalloproteinase-2, respectively, were not affected. Application of recombinant TIMP-1 and TIMP-2 proteins significantly inhibited invasion of mock-transfected clearCa-28 cells, supporting a role of TIMPs in Rac-mediated inhibition of invasion. To our knowledge, this is the first evidence that increased Rac signaling may inhibit invasion of epithelial tumor cells by up-regulation of TIMP-1 and TIMP-2.

Highlights

Tumor invasion is a complex biological process, during which tumor cells detach from the primary tumor and infiltrate the surrounding tissue
We have shown that sustained activation of Rac strongly inhibits cell migration, cell dissociation, and the more complex process of Matrigel invasion of human renal cell carcinoma (RCC) in vitro
We found, that in addition to promoting E-cadherin-mediated cell-cell adhesion, selective up-regulation of tissue inhibitor of metalloproteinases-1 (TIMP-1) and tissue inhibitors of metalloproteinases (TIMPs)-2 is another mechanism through which Rac may inhibit invasion of epithelial cells

Summary

Introduction

Tumor invasion is a complex biological process, during which tumor cells detach from the primary tumor and infiltrate the surrounding tissue This process requires loss of cell contacts between tumor cells, active cell migration, adhesion to the extracellular matrix (ECM) and proteolytic degradation of the ECM [1, 2]. Members of the Rho-like GTPases, including Cdc, Rac, and RhoA, mediate distinct actin cytoskeleton changes required for cell adhesion, migration, and invasion [3,4,5]. Their effects both on migration and invasion of epithelial cells seem to be at least in part cell type- and cell substrate-specific. The functional relevance of this up-regulation was substantiated by demonstrating that TIMP-1 and TIMP-2 inhibit Matrigel invasion of mock-transfected control cells

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Results

Conclusion