Integrin α2β1 Mediates Tyrosine Phosphorylation of Vascular Endothelial Cadherin Induced by Invasive Breast Cancer Cells

Mehran Haidari,Wei Zhang,Amy Caivano,Zhenping Chen,Leila Ganjehei,Ahmadreza Mortazavi,Christopher Stroud,Darren G Woodside,James T Willerson,Richard A.F Dixon

doi:10.1074/jbc.m112.395905

Mehran Haidari, Wei Zhang + Show 8 more

Open Access

https://doi.org/10.1074/jbc.m112.395905

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Abstract

The molecular mechanisms that regulate the endothelial response during transendothelial migration (TEM) of invasive cancer cells remain elusive. Tyrosine phosphorylation of vascular endothelial cadherin (VE-cad) has been implicated in the disruption of endothelial cell adherens junctions and in the diapedesis of metastatic cancer cells. We sought to determine the signaling mechanisms underlying the disruption of endothelial adherens junctions after the attachment of invasive breast cancer cells. Attachment of invasive breast cancer cells (MDA-MB-231) to human umbilical vein endothelial cells induced tyrosine phosphorylation of VE-cad, dissociation of β-catenin from VE-cad, and retraction of endothelial cells. Breast cancer cell-induced tyrosine phosphorylation of VE-cad was mediated by activation of the H-Ras/Raf/MEK/ERK signaling cascade and depended on the phosphorylation of endothelial myosin light chain (MLC). The inhibition of H-Ras or MLC in endothelial cells inhibited TEM of MDA-MB-231 cells. VE-cad tyrosine phosphorylation in endothelial cells induced by the attachment of MDA-MB-231 cells was mediated by MDA-MB-231 α(2)β(1) integrin. Compared with highly invasive MDA-MB-231 breast cancer cells, weakly invasive MCF-7 breast cancer cells expressed lower levels of α(2)β(1) integrin. TEM of MCF-7 as well as induction of VE-cad tyrosine phosphorylation and dissociation of β-catenin from the VE-cad complex by MCF-7 cells were lower than in MDA-MB-231 cells. These processes were restored when MCF-7 cells were treated with β(1)-activating antibody. Moreover, the response of endothelial cells to the attachment of prostatic (PC-3) and ovarian (SKOV3) invasive cancer cells resembled the response to MDA-MB-231 cells. Our study showed that the MDA-MB-231 cell-induced disruption of endothelial adherens junction integrity is triggered by MDA-MB-231 cell α(2)β(1) integrin and is mediated by H-Ras/MLC-induced tyrosine phosphorylation of VE-cad.

Highlights

The disruption of endothelial barrier function by tumor cells was studied
Attachment of Invasive Breast Cancer Cells (MDA-MB-231) to HUVECs Induces Tyr Phosphorylation of vascular endothelial cadherin (VE-cad), Dissociates ␤-Catenin from the VE-cad Complex, and Retracts Endothelial Cells—We recently showed that the interaction of monocytes with endothelial cells leads to Tyr phosphorylation of VE-cad and disruption of endothelial adherens junctions [14]
As indicated by immunofluorescence staining of VE-cad (Fig. 1D), the attachment of MDA-MB-231 cells to a monolayer of HUVECs led to a dramatic retraction in endothelial cells, a process that may precede MDA-MB-231 endothelial transmigration

Summary

Background

The disruption of endothelial barrier function by tumor cells was studied. Results: The attachment of tumor cells to endothelial cells leads to the disorganization of endothelial adherens junction. We sought to determine the signaling mechanisms underlying the disruption of endothelial adherens junctions after the attachment of invasive breast cancer cells. Our study showed that the MDA-MB-231 cell-induced disruption of endothelial adherens junction integrity is triggered by MDA-MB-231 cell ␣2␤1 integrin and is mediated by H-Ras/MLC-induced tyrosine phosphorylation of VE-cad. Most previous studies regarding the molecular mechanisms underlying tumor cell metastasis have focused on the role of cancer cells, whereas the mechanisms that protect endothelial barrier integrity during the diapedesis of metastatic cancer cells remain elusive. We explored the molecular pathways that regulate the disruption of endothelial barrier function after the attachment of invasive breast cancer cells. We determined that the attachment of invasive breast cancer cells (MDA-MB-231 cells) to endothelial cells leads to the disorganization of adherens junction structure and Tyr phosphorylation of VE-cad. We characterized the mechanisms underlying these events by identifying roles for ␣2␤1 integrin, endothelial H-Ras/Raf/MEK/ERK signaling, and myosin light chain (MLC) phosphorylation

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION