Crk-associated substrate tyrosine phosphorylation sites are critical for invasion and metastasis of SRC-transformed cells.
Crk-associated substrate (CAS, p130Cas) is a major tyrosine phosphorylated protein in cells transformed by v-crk and v-src oncogenes. We recently reported that reexpression of CAS in CAS-deficient mouse embryo fibroblasts transformed by oncogenic Src promoted an invasive phenotype associated with enhanced cell migration through Matrigel, organization of actin into large podosome ring and belt structures, activation of matrix metalloproteinase-2, and elevated tyrosine phosphorylation of the focal adhesion proteins FAK and paxillin. We have now extended these studies to examine the mechanism by which CAS achieves these changes and to evaluate the potential role for CAS in promoting in vivo tumor growth and metastasis. Whereas the presence or absence of CAS did not alter the primary growth of subcutaneous-injected Src-transformed mouse embryo fibroblasts, CAS expression was required to promote lung metastasis following removal of the primary tumor. The substrate domain YxxP tyrosines, the major sites of CAS phosphorylation by Src that mediate interactions with Crk, were found to be critical for promoting both invasive and metastatic properties of the cells. The ability of CAS to promote Matrigel invasion, formation of large podosome structures, and tyrosine phosphorylation of Src substrates, including FAK, paxillin, and cortactin, was also strictly dependent on the YxxP tyrosines. In contrast, matrix metalloproteinase-2 activation was most dependent on the CAS SH3 domain, whereas the substrate domain YxxP sites also contributed to this property. Thus multiple CAS-mediated signaling events are implicated in promoting invasive and metastatic properties of Src-transformed cells.
Department of Cell and Developmental Biology, Vanderbilt University School of Medicine, Nashville, TN 37212, USA.
Jan
J
Brábek
Sábata S
SS
Constancio
Priscila Fonseca
PF
Siesser
Nah-Young
NY
Shin
Ambra
A
Pozzi
Steven K
SK
Hanks
D000818:Animals; D001665:Binding Sites; D002461:Cell Line, Transformed; D002465:Cell Movement; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004338:Drug Combinations; D004789:Enzyme Activation; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D016391:Genes, src; D007279:Injections, Subcutaneous; D007797:Laminin / Q000378:metabolism; D008175:Lung Neoplasms / Q000473:pathology / Q000556:secondary; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D051379:Mice; D008819:Mice, Nude; D009361:Neoplasm Invasiveness; D009368:Neoplasm Transplantation; D050736:Oncogene Protein v-crk; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011509:Proteoglycans / Q000378:metabolism; D015682:Retroviridae Proteins, Oncogenic / Q000235:genetics / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry* / Q000378:metabolism; D018909:src Homology Domains
D003598:Cytoskeletal Proteins; D004338:Drug Combinations; D007797:Laminin; D050736:Oncogene Protein v-crk; D051419:Paxillin; D010750:Phosphoproteins; D011509:Proteoglycans; C496062:Pxn protein, mouse; D015682:Retroviridae Proteins, Oncogenic; C059485:matrigel; D014443:Tyrosine; D003094:Collagen; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D020778:Matrix Metalloproteinase 2