Disruption of focal adhesion kinase slows transendothelial migration of AU-565 breast cancer cells.

Transendothelial migration of cancer cells from the vasculature into tissue stroma is a final step in the metastatic cascade, prior to formation of secondary tumors. Due to its role in 2-dimensional migration of cells on extracellular matrix proteins, we hypothesized that focal adhesion kinase (FAK) promotes transendothelial migration of cancer cells. AU-565 cells are weakly invasive metastatic breast adenocarcinoma cells that migrate through bovine lung microvessel endothelial cell monolayers. Electric cell-substrate impedance sensing detects a significant decrease in monolayer resistance upon addition of AU-565 cells. Immunofluorescence microscopy and filter-based migration assays demonstrate that this drop in resistance correlates with transendothelial migration. Transfection of AU-565 cells with FAK siRNA results in significantly diminished transendothelial migration of AU-565 cells within 15h. Expression of the dominant negative FAK inhibitor FAK-related non-kinase (FRNK) also results in delayed AU-565 transendothelial migration, whereas over-expression of wildtype FAK does not impact transendothelial migration substantially. These results demonstrate that FAK affects the rate of a key step in the metastatic cascade.

[1]  D. Schlaepfer,et al.  FRNK blocks v‐Src‐stimulated invasion and experimental metastases without effects on cell motility or growth , 2002, The EMBO journal.

[2]  J. Guan,et al.  Stimulation of cell migration by overexpression of focal adhesion kinase and its association with Src and Fyn. , 1996, Journal of cell science.

[3]  W. Cance,et al.  Expression of focal adhesion kinase gene and invasive cancer , 1993, The Lancet.

[4]  R. Hynes,et al.  An MBoC Favorite: Fibronectin/integrin interaction induces tyrosine phosphorylation of a 120-kDa protein , 1991, Molecular biology of the cell.

[5]  Donna J. Webb,et al.  FAK–Src signalling through paxillin, ERK and MLCK regulates adhesion disassembly , 2004, Nature Cell Biology.

[6]  Kenneth M. Yamada,et al.  Taking Cell-Matrix Adhesions to the Third Dimension , 2001, Science.

[7]  D. Schlaepfer,et al.  Required role of focal adhesion kinase (FAK) for integrin-stimulated cell migration. , 1999, Journal of cell science.

[8]  I. Giaever,et al.  Micromotion of mammalian cells measured electrically. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[9]  J. Parsons,et al.  Autophosphorylation of the focal adhesion kinase, pp125FAK, directs SH2-dependent binding of pp60src , 1994, Molecular and cellular biology.

[10]  S. Kanner,et al.  A transmembrane-anchored chimeric focal adhesion kinase is constitutively activated and phosphorylated at tyrosine residues identical to pp125FAK. , 1994, The Journal of biological chemistry.

[11]  R. Elble,et al.  Focal Adhesion Kinase Activated by β4 Integrin Ligation to mCLCA1 Mediates Early Metastatic Growth* , 2002, The Journal of Biological Chemistry.

[12]  D. Schlaepfer,et al.  Control of motile and invasive cell phenotypes by focal adhesion kinase. , 2004, Biochimica et biophysica acta.

[13]  S. Aizawa,et al.  Reduced cell motility and enhanced focal adhesion contact formation in cells from FAK-deficient mice , 1995, Nature.

[14]  E. Parkinson,et al.  Increased dosage and amplification of the focal adhesion kinase gene in human cancer cells , 1999, Oncogene.

[15]  S. Hanks,et al.  Focal adhesion protein-tyrosine kinase phosphorylated in response to cell attachment to fibronectin. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[16]  J. Parsons,et al.  pp125FAK a structurally distinctive protein-tyrosine kinase associated with focal adhesions. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[17]  F. Luscinskas,et al.  The role of endothelial cell lateral junctions during leukocyte trafficking , 2002, Immunological reviews.

[18]  W. Cance,et al.  Overexpression of the focal adhesion kinase (p125FAK) in invasive human tumors. , 1995, Cancer research.