Isacke, Clare M

Jørgensen, Claus

Lim, Lindsay

Locard-Paulet, Marie

McMahon, Kelly

Sinclair, John

van Weverwijk, Antoinette

Veluscek, Giulia

Worboys, Jonathan D

Yuan, Yinyin

EExit of metastasizing tumor cells from the vascular system is regulated by their dynamic interactions with the endothelial cells that line the internal surface of vessels. While extravasation is a key event within the metastatic cascade, the signals controlling tumor cell adhesion to the endothelium and their subsequent transendothelial migration are poorly understood. By combining Stable Isotope Labeling by Amino acids in Cell culture (SILAC) and phosphoproteomic analysis, we mapped cell-specific changes in protein phosphorylation that were triggered by contact between breast cancer cells and endothelial cells. From the 2,669 unique phosphorylation sites identified, 77 and 43 were differentially phosphorylated in the tumor cells and endothelial cells, respectively. Among the phospho-regulated proteins within the tumor cells, the receptor tyrosine kinase Ephrin type-A receptor 2 (EPHA2) was found to inhibit transendothelial migration of breast cancer cells. We demonstrated that this was dependent on EPHA2-Tyr772 phosphorylation, which was decreased in the breast cancer cells upon endothelial contact. Importantly, comparison of isogenic breast cancer cell lines with different metastatic capacities revealed that EPHA2-Tyr772 dephosphorylation was specifically associated with higher lung metastatic potential. Altogether, our cell-specific phosphoproteomic analysis provides the first bidirectional map of contact-initiated signaling between tumor and endothelial cells, which has led to the characterization of a novel EPHA2-based regulatory mechanism of transendothelial migration

The University of Manchester - Institutional Repository

Phosphoproteomic analysis of tumor-endothelial signaling identifies EPHA2 as a negative regulator of transendothelial migration

Phosphoproteomic analysis of tumor-endothelial signaling identifies EPHA2 as a negative regulator of transendothelial migration

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The University of Manchester - Institutional Repository