CLEAVAGE OF BP180, A 180-KDA BULLOUS PEMPHIGOID ANTIGEN, YIELDS A 120-KDA COLLAGENOUS EXTRACELLULAR POLYPEPTIDE

The hemidesmosome (HD) is a cell-to-substrate adhesion apparatus found in stratified and complex epithelia. One of the putative cell-matrix adhesion molecules present in the HD is the 18O-kDa bullous pemphigoid antigen (BP180), also termed type XVII collagen. In our previous stud y, using a monoclonal antibody (mAb) 1337, we have detected a 120-kDa collagenase-sensitive polypeptide in the HD fraction (Uematsu, J. and Owaribe, H. (1993) Cell Struct. Funct. 18, 588 (abstr.)). The present study was undertaken to assess the relation of the 120-kDa polypeptide to this BP180. Immunofluorescence microscopy of bovine skin revealed the basement membrane zone of skin to be stained clearly with mAb 1337 , whereas the lateral surfaces of basal cells, which were decorated by typical antibodies against BP180, were not. The antibody did not dete ct HDs in cultured cells but rather in the culture medium. These resul ts indicate a localization of mAb 1337 antigen distinct from BP180. Ho wever, the same polypeptide was also recognized by monoclonal antibodi es to the extracellular but not the cytoplasmic part of BP180, and fou nd to react with a polyclonal antibody against the non-collagenous 16A domain of BP180. Therefore, the polypeptide was identified as an extr acellular fragment of BP180. mAb 1337 immunoprecipitated the 120-kDa f ragment from the medium, but not the 180-kDa molecule of BP180 extract ed from cultured cells, indicating that the antibody specifically reco gnizes the fragment. The mAb 1337 apparently recognizes a unique epito pe that is exposed or formed by the cleavage. Hence, the staining patt ern observed for bovine skin demonstrated the presence of the 120-kDa extracellular fragment. Rotary shadow electron microscopy of affinity- purified 120-kDa fragments demonstrated that they have the unique mole cular shape consisting of a central rod and a flexible tail, without t he globular head that is present in the BP180 molecule. From these res ults, we conclude that mAb 1337 shows unique epitope specificity, reco gnizing only the 120-kDa extracellular fragment of BP180, which is con stitutively cleaved on the cell surface as a 120-kDa fragment both in in vivo and in vitro.