Editors' ChoiceCancer

Promoting Single-Cell Invasion

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Sci. Signal.  07 May 2013:
Vol. 6, Issue 274, pp. ec103
DOI: 10.1126/scisignal.2004305

Ephrins are membrane ligands for the Eph family of receptor tyrosine kinases, and this large family of receptors and ligands are regulators of cell adhesion and migration. Tumor cells have co-opted these essential mediators of development, as well as matrix proteases, to promote metastasis. Sugiyama et al. studied the relationship between EphA2 and membrane type-1 matrix metalloproteinase (MT1-MMP) in the migratory behavior and metastatic potential of breast cancer cell lines. Cell lines with the greatest invasiveness into a collagen matrix and that formed the largest colonies in a three-dimensional (3D) matrix were positive for both MT1-MMP and EphA2, and migration or growth into either substrate was impaired by knockdown of either protein. In the 3D culture system, the double-positive cell lines exhibited distinct growth phenotypes that correlated with the presence of an MT1-MMP–dependent EphA2 cleavage product. Cells with EphA2 cleavage exhibited invasive single cells that were separate from the main colony, whereas cells without this cleavage event formed multicellular outgrowths, typical of collective cell invasion. The localization of EphA2 was also different: The cells exhibiting the single-cell invasion phenotype had abundant EphA2 in intracellular perinuclear structures and at the cell surface, whereas the collectively migrating cells had EphA2 only at the cell surface. The site of cleavage in EphA2 was identified by mass spectrometry, enabling the engineering of a mutant resistant to cleavage (G391R) and a mutant with increased processing (D359I). Expression of the D359I mutant in MT1-MMP–positive cells (cis), but not when the two proteins were expressed in separate cells (trans), resulted in enhanced cleavage, increased intracellular localization of EphA2, and cell rounding. Examination of cells expressing the D359I mutant with or without MT1-MMP knockdown by video microscopy suggested that EphA2 cleavage contributed to contact-mediated cellular repulsion. In xenograft studies, compared with MT1-MMP–positive cells overexpressing wild-type EphA2, expression of the D359I mutant promoted single-cell invasion into the stroma and reduced cell-cell contacts, although overexpression of either protein was sufficient to increase metastasis compared with tumors derived from the control cell line. In human invasive breast cancer samples, both EphA2 abundance and MT1-MMP abundance were higher in primary tumors and metastases than in noncancerous tissues, and cancer cells separated from the main tumor mass that had increased MT1-MMP abundance also tended to have intracellular EphA2, whereas those within the main tumor had surface EphA2. Thus, MT1-MMP–mediated cleavage of EphA2 appears to contribute to the conversion of cells within a solid tumor to those with a single-cell invasive phenotype.

N. Sugiyama, E. Gucciardo, O. Tatti, M. Varjosalo, M. Hyytiäinen, M. Gstaiger, K. Lehti, EphA2 cleavage by MT1-MMP triggers single cancer cell invasion via homotypic cell repulsion. J. Cell Biol. 201, 467–484 (2013). [Abstract] [Full Text]

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