A recombinant tail-less integrin beta 4 subunit disrupts hemidesmosomes, but does not suppress alpha 6 beta 4-mediated cell adhesion to laminins.
To examine the function of the alpha 6 beta 4 integrin we have determined its ligand-binding ability and overexpressed two potentially dominant negative mutant beta 4 subunits, lacking either the cytoplasmic or extracellular domain, in bladder epithelial 804G cells. The results of cell adhesion and radioligand-binding assays showed that alpha 6 beta 4 is a receptor for several laminin isoforms, including laminin 1, 2, 4, and 5. Overexpression of the tail-less or head-less mutant beta 4 subunit did not suppress alpha 6 beta 4-mediated adhesion to laminins, as both types of transfectants adhered to these ligands in the presence of blocking anti-beta 1 antibodies as well as the controls. However, immunofluorescence experiments indicated that the endogenous alpha 6 beta 4 integrin and other hemidesmosomal markers were not concentrated in hemidesmosomes in cells overexpressing tail-less beta 4, while the distribution of these molecules was not altered in cells overexpressing the head-less subunit. Electron microscopic studies confirmed that cells overexpressing tail-less beta 4 had a drastically reduced number of hemidesmosomes, while cells expressing the head-less subunit had a normal number of these structures. Thus, expression of a tail-less, but not a head-less mutant beta 4 subunit leads to a dominant negative effect on hemidesmosome assembly without suppressing initial adhesion to laminins. We conclude that the alpha 6 beta 4 integrin binds to several laminins and plays an essential role in the assembly and/or stability of hemidesmosomes, that alpha 6 beta 4-mediated adhesion and hemidesmosome assembly have distinct requirements, and that it is possible to use a dominant negative approach to selectively interfere with a specific function of an integrin.
Spinardi
L
L
Department of Pathology, New York University School of Medicine, New York 10016, USA.
Einheber
S
S
Cullen
T
T
Milner
T A
TA
Giancotti
F G
FG
D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D001324:Autoantigens / Q000032:analysis; D014408:Biomarkers, Tumor / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D002352:Carrier Proteins; D002448:Cell Adhesion / Q000502:physiology*; D003094:Collagen; D003598:Cytoskeletal Proteins; D003896:Desmosomes / Q000378:metabolism* / Q000648:ultrastructure; D000071821:Dystonin; D004847:Epithelial Cells; D004848:Epithelium / Q000737:chemistry; D006801:Humans; D039161:Integrin alpha6beta4; D039663:Integrin beta4; D016023:Integrins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D007797:Laminin / Q000378:metabolism*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D010391:Pemphigoid, Bullous; D011869:Radioligand Assay; D051381:Rats; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000378:metabolism; D017384:Sequence Deletion / Q000502:physiology; D014407:Tumor Cells, Cultured; D001743:Urinary Bladder; D000097805:Collagen Type XVII
D000954:Antigens, Surface; D001324:Autoantigens; D014408:Biomarkers, Tumor; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D039161:Integrin alpha6beta4; D039663:Integrin beta4; D016023:Integrins; D007797:Laminin; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D011994:Recombinant Proteins; D000097805:Collagen Type XVII; C486268:DST protein, human