EWI-2 regulates alpha3beta1 integrin-dependent cell functions on laminin-5.
EWI-2, a cell surface immunoglobulin SF protein of unknown function, associates with tetraspanins CD9 and CD81 with high stoichiometry. Overexpression of EWI-2 in A431 epidermoid carcinoma cells did not alter cell adhesion or spreading on laminin-5, and had no effect on reaggregation of cells plated on collagen I (alpha2beta1 integrin ligand). However, on laminin-5 (alpha3beta1 integrin ligand), A431 cell reaggregation and motility functions were markedly impaired. Immunodepletion and reexpression experiments revealed that tetraspanins CD9 and CD81 physically link EWI-2 to alpha3beta1 integrin, but not to other integrins. CD81 also controlled EWI-2 maturation and cell surface localization. EWI-2 overexpression not only suppressed cell migration, but also redirected CD81 to cell filopodia and enhanced alpha3beta1-CD81 complex formation. In contrast, an EWI-2 chimeric mutant failed to suppress cell migration, redirect CD81 to filopodia, or enhance alpha3beta1-CD81 complex formation. These results show how laterally associated EWI-2 might regulate alpha3beta1 function in disease and development, and demonstrate how tetraspanin proteins can assemble multiple nontetraspanin proteins into functional complexes.
Stipp
Christopher S
CS
Dana-Farber Cancer Institute, Department of Pathology, Harvard Medical School, Boston, MA 02115, USA.
Kolesnikova
Tatiana V
TV
Hemler
Martin E
ME
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D018801:CD2 Antigens / Q000378:metabolism; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000502:physiology; D006801:Humans; D007136:Immunoglobulins / Q000378:metabolism*; D039201:Integrin alpha3beta1 / Q000378:metabolism*; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D060225:Tetraspanin 28; D060245:Tetraspanin 29; D000097794:Kalinin
D015703:Antigens, CD; D018801:CD2 Antigens; D015815:Cell Adhesion Molecules; D007136:Immunoglobulins; D039201:Integrin alpha3beta1; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D060225:Tetraspanin 28; D060245:Tetraspanin 29; D000097794:Kalinin; C554798:CD81 protein, human; C554869:CD9 protein, human; C440028:IGSF8 protein, human