THE LOCALIZATION OF BULLOUS PEMPHIGOID ANTIGEN-180 (BP180) IN HEMIDESMOSOMES IS MEDIATED BY ITS CYTOPLASMIC DOMAIN AND SEEMS TO BE REGULATED BY THE BETA-4 INTEGRIN SUBUNIT

Bullous pemphigoid antigen 180 (BP180) is a component of hemidesmosome s, i.e., cell-substrate adhesion complexes. To determine the function of specific sequences of BP180 to its incorporation in hemidesmosomes, we have transfected 804G cells with cDNA-constructs encoding wild-typ e and deletion mutant forms of human BP180, The results show that the cytoplasmic domain of BP180 contains sufficient information for the re cruitment of the protein into hemidesmosomes because removal of the ex tracellular and transmembrane domains does not abolish targeting. Expr ession of chimeric proteins, which consist of the membrane targeting s equence of K-Ras fused to the cytoplasmic domain of BP180 with increas ing internal deletions or lacking the NH2 terminus, indicates that the localization of BP180 in hemidesmosomes is mediated by a segment that spans 265 amino acids. This segment comprises two important regions l ocated within the central part and at the NH2 terminus of the cytoplas mic domain of BP180. To investigate the effect of the alpha 6 beta 4 i ntegrin on the subcellular distribution of BP180, we have transfected COS-7 cells, which lack alpha 6 beta 4 and BP180, with cDNAs for BP180 as well as for human alpha 6A and beta 4. We provide evidence that a mutant form of BP180 lacking the collagenous extracellular domain as w ell as a chimeric protein, which contains the entire cytoplasmic domai n of BP180, are colocalized with alpha 6 beta 4. In contrast, when cel ls were transfected with cDNAs for alpha 6A and mutant forms of beta 4 , either lacking the cytoplasmic COOH-terminal half or carrying phenyl alanine substitutions in the tyrosine activation motif of the cytoplas mic domain, the recombinant BP180 molecules were mostly not colocalize d with alpha 6 beta 4, but remained diffusely distributed at the cell surface, Moreover, in cells transfected with cDNAs for alpha 6A and a beta 4/beta 1 chimera, in which the cytoplasmic domain of beta 4 was r eplaced by that of the beta 1 integrin subunit, BP180 was not colocali zed with the alpha 6 beta 4/beta 1 chimera in focal adhesions, but rem ained again diffusely distributed. These results indicate that sequenc es within the cytoplasmic domain of beta 4 determine the subcellular d istribution of BP180.