A point mutation in the integrin beta 6 subunit abolishes both alpha v beta 6 binding to fibronectin and receptor localization to focal contacts.
The integrin alpha v beta 6 was initially identified from primary cultures of airway epithelial cells. This integrin is expressed in bronchiolar and alveolar epithelium during development and in settings of injury and/or inflammation and mediates attachment of epithelial cells to fibronectin and tenascin. Like other integrins, this receptor localizes to structures called focal contacts in cells plated on appropriate ligands. In the present study, we produced a mutant beta 6 cDNA (beta 6m) containing a single substitution of Asp140 with Ala and transfected mutant (or wild-type) beta 6 cDNA into the human colon carcinoma cell line SW480. In parallel, we used cDNAs truncated just proximal to the transmembrane domain to generate secreted forms of mutant alpha v beta 6 in Chinese hamster ovary (CHO) cells. The mutant beta 6, like the wild type, formed heterodimers with human alpha v that were expressed on the cell surface of SW480 cells and secreted by CHO cells. Secreted alpha v beta 6 containing this point mutation did not bind to fibronectin-Sepharose. Furthermore, in contrast to wild-type beta 6, the mutant form did not allow SW480 cells to bind to fibronectin in the presence of beta 1-blocking antibody and did not localize to focal contacts. Our results confirm that the Asp140 of beta 6, like the corresponding residues in beta 1 (Asp130) and beta 3 (Asp119), is critical for interactions of alpha v beta 6 with ligand, and also suggest that ligand binding to alpha v beta 6 is necessary for localization of this receptor to focal contacts.
Huang
X Z
XZ
Lung Biology Center, UCSF 94143, USA.
Chen
A
A
Agrez
M
M
Sheppard
D
D
D000409:Alanine / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000235:genetics; D001483:Base Sequence; D016466:CHO Cells / Q000502:physiology; D002448:Cell Adhesion / Q000235:genetics; D006224:Cricetinae; D004847:Epithelial Cells; D005353:Fibronectins / Q000378:metabolism*; D039641:Integrin beta Chains; D016023:Integrins / Q000235:genetics* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation / Q000502:physiology*; D011485:Protein Binding / Q000502:physiology; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism / Q000648:ultrastructure; D012685:Sepharose; D014162:Transfection; D014407:Tumor Cells, Cultured / Q000166:cytology / Q000502:physiology
D005353:Fibronectins; D039641:Integrin beta Chains; D016023:Integrins; D011956:Receptors, Cell Surface; C072170:integrin beta6; D001216:Asparagine; D012685:Sepharose; D000409:Alanine