Plakophilin 1 interferes with plakoglobin binding to desmoplakin, yet together with plakoglobin promotes clustering of desmosomal plaque complexes atcell-cell borders

Desmosomes are adhesive junctions that link intermediate filament networks to sites of strong intercellular adhesion. These junctions play an importan t role in providing strength to tissues that experience mechanical stress s uch as heart and epidermis. The basic structural elements of desmosomes are similar to those of the better-characterized adherens junctions, which anc hor actin-containing microfilaments to cadherins at the plasma membrane. Th is linkage of actin to classic cadherins is thought to occur through an ind irect mechanism requiring the associated proteins, alpha- and beta -catenin . In the case of desmosomes, both linear and lateral interactions have been proposed as playing an important role in formation of the plaque and linka ge to the cytoskeleton. However, the precise nature of these interactions a nd how they cooperate in desmosome assembly are poorly understood. Here we employ a reconstitution system to examine the assembly of macromole cular complexes from components found in desmosomes of the differentiated l ayers of complex tissues. We demonstrate the existence of a Triton-soluble complex of proteins containing full length desmoplakin (DP), the arm protei n plakoglobin, and the cytoplasmic domain of the desmosomal cadherin, desmo glein 1 (Dsg1). In addition, full length DP, but not an N-terminal plakoglo bin binding domain of DP, coimmunoprecipitated with the Dsg1 tail in the ab sence of plakoglobin in HT1080 cells. The relative roles of the arm protein s plakoglobin and plakophilin 1 (PKP1) were also investigated. Our results suggest that, in the Triton soluble pool, PKP1 interferes with binding of p lakoglobin to full length DP when these proteins are co-expressed. Neverthe less, both plakoglobin and PKP1 are required for the formation of clustered structures containing DP and the Dsg1 tail that ultrastructurally appear s imilar to desmosomal plaques found in the epidermis. These findings suggest that more than one armadillo family member is required for normal assembly and clustering of the desmosomal plaque in the upper layers of the epiderm is.