title: Functional relevance of urinary-type plasminogen activator receptor-alpha3beta1 integrin association in proteinase regulatory pathways.
abstract: Squamous cell carcinoma of the oral cavity is characterized by persistent, disorganized expression of integrin alpha3beta1 and enhanced production of urinary-type plasminogen activator (uPA) and its receptor (uPAR) relative to normal oral mucosa. Because multivalent aggregation of alpha3beta1 integrin up-regulates uPA and induces a dramatic co-clustering of uPAR, we explored the hypothesis that lateral ligation of alpha3beta1 integrin by uPAR contributes to uPA regulation in oral mucosal cells. To investigate mechanisms by which uPAR/alpha3beta1 binding enhances uPA expression, integrin-dependent signal activation was assessed. Both Src and ERK1/2 were phosphorylated in response to integrin aggregation, and blocking Src kinase activity completely abrogated ERK1/2 activation and uPA induction, whereas inhibition of epidermal growth factor receptor tyrosine kinase activity did not alter uPA expression. Proteinase up-regulation occurred at the transcriptional level and mutation of the AP1 (-1967) site in the uPA promoter blocked the uPAR/integrin-mediated transcriptional activation. Because uPAR is redistributed to clustered alpha3beta1 integrins, the requirement for uPAR/alpha3beta1 interaction in uPA regulation was assessed. Clustering of alpha3beta1 in the presence of a peptide (alpha325) that disrupts uPAR/alpha3beta1 integrin binding prevented uPA induction. Depletion of cell surface uPAR using small interfering RNA also blocked uPA induction following integrin alpha3beta1 clustering. These results were confirmed using a genetic strategy in which alpha3 null epithelial cells reconstituted with wild type alpha3 integrin, but not a mutant alpha3 unable to bind uPAR, induced uPA expression upon integrin clustering, confirming the critical role of uPAR in integrin-regulated proteinase expression. Disruption of uPAR/alpha3beta1 binding using peptide alpha325 or small interfering RNA blocked filopodia formation and matrix invasion, indicating that this interaction stimulates invasive behavior. Together these data support a model wherein matrix-induced clustering ofalpha3beta1 integrin promotes uPAR/alpha3beta1 interaction, thereby potentiating cellular signal transduction pathways culminating in activation of uPA expression and enhanced uPA-dependent invasive behavior.
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University, Chicago, Illinois 60611.
authors.forename: Supurna
authors.initials: S
authors.lastname: Ghosh
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University, Chicago, Illinois 60611.
authors.forename: Jeff J
authors.initials: JJ
authors.lastname: Johnson
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University, Chicago, Illinois 60611.
authors.forename: Ratna
authors.initials: R
authors.lastname: Sen
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University, Chicago, Illinois 60611.
authors.forename: Subhendu
authors.initials: S
authors.lastname: Mukhopadhyay
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University, Chicago, Illinois 60611.
authors.forename: Yueying
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Division of Pulmonary and Critical Care Medicine, University of California, San Francisco, California 94143.
authors.forename: Feng
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Division of Pulmonary and Critical Care Medicine, University of California, San Francisco, California 94143.
authors.forename: Ying
authors.initials: Y
authors.lastname: Wei
authors.affiliation: Division of Pulmonary and Critical Care Medicine, University of California, San Francisco, California 94143.
authors.forename: Harold A
authors.initials: HA
authors.lastname: Chapman
authors.affiliation: Department of Cell and Molecular Biology, Northwestern University, Chicago, Illinois 60611; Robert H. Lurie Comprehensive Cancer Center, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611. Electronic address: mss130@northwestern.edu.
authors.forename: M Sharon
authors.initials: MS
authors.lastname: Stack
mesh_terms: D000818:Animals; D002448:Cell Adhesion; D002460:Cell Line; D015536:Down-Regulation; D005109:Extracellular Matrix / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D039201:Integrin alpha3beta1 / Q000235:genetics / Q000378:metabolism*; D015603:Keratinocytes; D051379:Mice; D011485:Protein Binding; D011494:Protein Kinases / Q000378:metabolism; D011554:Pseudopodia / Q000502:physiology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator; D015398:Signal Transduction; D014568:Urokinase-Type Plasminogen Activator / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism
chemical_list: D039201:Integrin alpha3beta1; C524898:PLAUR protein, human; C524899:Plaur protein, mouse; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D011494:Protein Kinases; D014568:Urokinase-Type Plasminogen Activator