Focal Adhesion Kinase Activated by β4 Integrin Ligation to mCLCA1 Mediates Early Metastatic Growth

Abstract

Hematogenous metastases originate from tumor cells arrested in the vasculature of select target organs. This arrest is tumor-and tissue-specific and is mediated at least in part by distinct tumor cell/endothelial cell adhesion ligand/receptor pairs (reviewed in Refs. 1 and 2). Studies in our laboratory have shown that lung metastatic human breast cancer cells colonized the lungs following adhesion to hCLCA2, a Ca 2ϩ -sensitive chloride channel protein that is expressed on the endothelial cell luminal surface of human pulmonary arteries, arterioles, and interlobular venules (3). Similar to the unique adhesion functions of other channel proteins (reviewed in Ref. 4), CLCA proteins mediate adhesion via the ␤ 4 integrin tumor cell ligand, which for the first time has been associated with a cell-cell adhesion function (3, 4). This novel adhesion interaction between members of CLCA family of proteins (e.g. bCLCA2 (Lu-ECAM-1), hCLCA2) and the ␤ 4 integrin has been scrutinized by a variety of stringently controlled biochemical and functional assay procedures (3, 5, 6). These assays included (i) the co-immunoprecipitation of the ␤ 4 -hCLCA2 complex from extracts of lung metastatic MDA-MB-231 breast cancer cells bound to monolayers of hCLCA2-transfected human embryonic kidney (HEK) 1 293 cells (3); (ii) the selective binding of immunopurified, recombinant hCLCA2 to membrane-immobilized, reconstituted ␤ 4 integrin in Far Westerns (3); (iii) the increased expression of the ␤ 4 integrin in breast cancer cell lines selected in vivo for enhanced lung colonization and the concomitant increased adhesion of the selected cells to hCLCA2 (3); (iv) the loss of hCLCA2 adhesion of breast cancer cells subjected to selective cleavage of the ␤ 4 integrin with the metalloproteinase matrilysin (3, 7) or tumor cells transfected with mutant ␤ 4 integrin (tail-less) (3); (v) the inhibition of pulmonary metastases by ␤ 4 -hCLCA2 adhesion-blocking antibodies directed against either of the interacting adhesion molecules (3, 5, 6); and (vi) the increased lung metastatic performance of tumor cells overexpressing the ␤ 4 integrin (3). These data are in agreement with previous reports showing that overexpression of the ␤ 4 integrin is associated with an aggressive/metastatic cancer phenotype in several malignancies (reviewed in Ref. 8) and, more recently, with the finding that selection for enhanced lung colonization concurs with prominent overexpression of the ␤ 4 integrin gene in a murine metastasis model analyzed by cDNA microarray (9).Here, B16-F10 melanoma cells, characterized by strong surface expression of the ␤ 4 integrin (10, 11) and consistently high lung colonization potential (12), were employed to explore whether the ␤ 4 integrin interacts with a murine CLCA family member to promote metastasis of mouse lungs. Although the existence of a murine counterpart of hCLCA2 has been suspected by the positive immunohistochemical staining of endothelia of mouse pulmonary blood vessels with anti-bCLCA2 mAb6D3 and by the anti-metastatic effect of ...

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“…FAK is also important in integrin-mediated cell cycle progression through the mitogen activating protein (MAP) kinase pathway [74,75]. Additionally, FAK and Pyk2 activation are implicated in the growth and metastasis of transformed cells [76][77][78]. Together, these data imply that FAK is an important intersection point for cell motility and growth transformation.…”

Molecular and Functional Analyses of Two New Calcium-activated Chloride Channel Family Members from Mouse Eye and Intestine

“…The importance of cloning mCLCA5 and mCLCA6 is, thus, apparent in terms of the interesting roles of their human homologues. mCLCA5 is highly homologous to hCLCA2, a protein involved in the vascular arrest of lung metastatically competent cancer cells (22,25). Both hCLCA2 and mCLCA5 have ␤ 4 integrin binding motifs (26).…”

“…␤ 4 integrin expressed on the surface of these melanoma cells can adhere to mCLCA1 in lung endothelium. This leads to focal adhesion kinase complexing, activation, and downstream signaling to extracellular signal-regulated kinase, which promotes intravascular metastatic growth (25). Human breast cancer cells can colonize the lung by expression of ␣ 6 ␤ 4 integrin and adhesion to hCLCA2 (22).…”

Synergistic Effect of Focal Adhesion Kinase Overexpression and Hepatocyte Growth Factor Stimulation on Cell Transformation

“…Alternatively, HGF may cause the tyrosine phosphorylation of certain integrins such as ␣ 6 ␤ 4 (49), which then induces FAK to undergo autophosphorylation. Recently, FAK has been shown to be activated by integrin ␤ 4 (50).…”