Dissection of the bovine epidermal desmosome into cytoplasmic protein and membrane glycoprotein domains.

Epidermal desmosomes contain two main regions. The core consists of a pair of membranes, one on either side of a cross-striated intercellular space bisected by a denser midline. The cytoplasmic compartment comprises a dense plaque deposited on the cytoplasmic surface of each membrane and a diffuse layer occupying the zone between the plaque and attached alpha-keratin filaments. Analysis of isolated desmosomes by SDS-PAGE has shown the presence of four major protein (dpl-4) and three major glycoprotein (dgl-3) bands, which have been allocated to the cytoplasmic and core compartments, respectively. In the present paper, we report the use of urea to fractionate this complex structure, both in situ and following isolation with citrate buffer, pH2.6. Extraction of the living layers of bovine epidermis with 9M-urea, pH7.5, resulted in rapid removal of the dense desmosomal plaques, followed by separation and vesiculation of desmosomal membranes. The resistance of the plaque to urea increased abruptly at the transition between living epidermis and dead, dehydrated horny layer. A similar sequence of morphological changes accompanied the extraction of isolated desmosomes with urea. Analysis of residues and extracts of isolated desmosomes by SDS-PAGE confirmed the selectivity of 9 M-urea, pH7.5, for the cytoplasmic compartment. The four major desmosomal proteins, dpl-4 (Mr240, 215, 90 and 83 (X 10(3)), respectively) predominated in the extracts. Desmosomal membranes, both paired and vesiculated, consisted almost entirely of the three desmosomal glycoproteins dgl-3 (Mr150, 120 and 110 (X 10(3)), respectively). These results provide evidence that all three desmosomal glycoproteins are integral membrane proteins. The separation of desmosomal membranes by urea, which is not accompanied by additional loss of proteins, further suggests that desmosomal adhesion is based on interactions between membrane components with no separate extracellular molecules being involved. The dissection of the desmosome by urea into two topographically and biochemically distinct domains should facilitate further studies on the molecular basis of desmosomal adhesion and alpha-keratin filament binding.