Formation of hemidesmosomes in vitro by a transformed rat bladder cell line

Abstract

Two hemidesmosomal plaque components of 230 and 180 kD have recently been characterized using autoantibodies in the serum samples of bullous pemphigoid (BP) patients (Klatte, D. H., M. A. Kurpakus, K. A. Grelling, and J. C. R. Jones. 1989, J. Cell Biol. 109:3377-3390). These BP autoantibodies generate the type of staining patterns that one would predict for formed hemidesmosomes, i.e., a punctate staining pattern towards the substratum; in less than 50% of various primary epithelial and transformed epidermal cell lines even when such cells are maintained in culture for prolonged periods. In contrast, affinity-purified human autoantibodies against the 230-kD hemidesmosomal plaque component generate intense immunofluorescence staining along the region of cell-substratum interaction in the rat bladder tumor cell line 804G maintained on uncoated glass cover-slips. This pattern is distinct from that observed in the 804G cells using an antibody preparation directed against vinculin, a component of adhesion plaques. Ultrastructural analyses of the 804G cells reveals that hemidesmosome-like structures occur along the basal surface of cells where they abut the substratum. These structures are present in 804G cells maintained in culture in reduced levels of Ca2+ and are recognized by autoantibodies directed against the 230-kD hemidesmosomal plaque component as determined by immunogold ultrastructural localization. To study hemidesmosome appearance in this cell line, 804G cells were trypsinized and then allowed to readhere to glass coverslips. In rounded, unattached 804G cells, hemidesmosome-like plaque structures occur along the cell surface. These structures are recognized by the 230-kD autoantibodies. At 1 h after plating, hemidesmosomes are observed along the substratum attached surface of cells. Protein synthesis is not required for the appearance of these hemidesmosomes. Within 4 h of plating, autoantibody staining and hemidesmosomes appear towards the cell periphery. Subsequently, the polypeptide recognized by the BP autoantibodies becomes concentrated in the perinuclear region, where there are numerous hemidesmosomes. We propose that the hemidesmosomes in 804G cells are involved in cell-substratum adhesion. We discuss possible mechanisms of assembly of hemidesmosomes in the 804G cells. Indeed, the 804G cells should prove an invaluable cell line for the biochemical and molecular dissection of hemidesmosome structure, function, and assembly.