IGF-I Receptor-induced Cell-Cell Adhesion of MCF-7 Breast Cancer Cells Requires the Expression of Junction Protein ZO-1

Loredana Mauro,Monica Bartucci,Catia Morelli,Sebastiano Ando',Eva Surmacz

doi:10.1074/jbc.m106673200

Abstract

Hyperactivation of the insulin-like growth factor I receptor (IGF-IR) contributes to primary breast cancer development, but the role of the IGF-IR in tumor metastasis is unclear. Here we studied the effects of the IGF-IR on intercellular connections mediated by the major epithelial adhesion protein, E-cadherin (E-cad). We found that IGF-IR overexpression markedly stimulated aggregation in E-cad-positive MCF-7 breast cancer cells, but not in E-cad-negative MDA-MB-231 cells. However, when the IGF-IR and E-cad were co-expressed in MDA-MB-231 cells, cell-cell adhesion was substantially increased. The IGF-IR-dependent cell-cell adhesion of MCF-7 cells was not related to altered expression of E-cad or alpha-, beta-, or gamma-catenins but coincided with the up-regulation of another element of the E-cad complex, zonula occludens-1 (ZO-1). ZO-1 expression (mRNA and protein) was induced by IGF-I and was blocked in MCF-7 cells with a tyrosine kinase-defective IGF-IR mutant. By co-immunoprecipitation, we found that ZO-1 associates with the E-cad complex and the IGF-IR. High levels of ZO-1 coincided with an increased IGF-IR/alpha-catenin/ZO-1-binding and improved ZO-1/actin association, whereas down-regulation of ZO-1 by the expression of an anti-ZO-1 RNA inhibited IGF-IR-dependent cell-cell adhesion. The results suggested that one of the mechanisms by which the activated IGF-IR regulates E-cad-mediated cell-cell adhesion is overexpression of ZO-1 and the resulting stronger connections between the E-cad complex and the actin cytoskeleton. We hypothesize that in E-cad-positive cells, the IGF-IR may produce antimetastatic effects.

Highlights

The insulin-like growth factor I (IGF-I)1 receptor (IGF-IR) is a ubiquitous tyrosine kinase capable of regulating different growth-related and -unrelated processes [1,2,3]
1 The abbreviations used are: IGF-I, insulin-like growth factor I; insulin-like growth factor I receptor (IGF-IR), IGF-I receptor; E-cad, E-cadherin; ZO-1, zonula occludens-1 junction protein; Y3F, Tyr-1131, Tyr-1135, and Tyr-1136 replaced with Phe; WB, Western blotting or Western blotted; pAb, polyclonal antibody; mAb, monoclonal antibody; ERK1, extracellular signal-related kinase 1; IRS-1, insulin receptor substrate 1; MAPK, mitogen-activated protein kinase
Coexpression of the IGF-IR and E-cad resulted in robust cell-cell adhesion of MDA-MB-231 cells, whereas the expression of Ecad alone was less efficient in inducing intercellular contacts (Fig. 1, C and D) (Table I)

Summary

Introduction

The insulin-like growth factor I (IGF-I) receptor (IGF-IR) is a ubiquitous tyrosine kinase capable of regulating different growth-related and -unrelated processes [1,2,3]. The intrinsic ligand-independent tyrosine kinase activity of the IGF-IR has been found to be substantially up-regulated (ϳ2– 4-fold) in breast cancer cells [4]. Recent clinical and experimental data indicate that up-regulation of IGF-IR signaling in estrogen receptor-positive breast cancer cells is associated with autonomous cell proliferation, estrogen-independence, and increased resistance to various antitumor treatments [1]. The experimental data suggest that the IGF-IR has a function in cell spreading by effectively stimulating the motility of different metastatic breast cancer cell lines lacking the expression of a major adhesion protein, E-cadherin (E-cad) [1, 5, 6]. In this study we assessed the effects of the IGF-IR on the elements of the E-cad adhesion complex, i.e. E-cad; ␤-, ␥-, and ␣-catenins; and ␣-catenin-associated proteins (see Fig. 6). Representative three-dimensional cultures are shown in Figs. 1 and 7

Methods

Results

Conclusion