Biochemical characterization of the soluble form of the junctional plaque protein, plakoglobin, from different cell types.
A polypeptide of identical molecular mass (Mr 83,000) and charge to desmosomal plakoglobin from bovine snout epidermis was identified in soluble and pelletable fractions from diverse tissues and cells of different mammalian species, including cells and tissues devoid of desmosomes (e.g. endothelial, retinal, lenticular cells, fibroblasts). The protein, however, was not detected in erythrocytes and platelets and in myeloma cells, nor in smooth muscle tissue. In all cells examined, the plakoglobin soluble upon cell lysis in buffers of near-physiological pH and ionic strength (21-31% of the total plakoglobin in the different cell types) was found to exist in a distinct molecular form. On sucrose gradient centrifugation it appeared at about 7 S and gel filtration chromatography revealed a Stokes radius of about 5.0 nm, from which an Mr of about 170,000 was estimated. By using isoelectric focusing under non-denaturing conditions, soluble approximately equal to 7-S plakoglobin had an isoelectric point at about pH 5.3. The plaque-bound and the soluble form of plakoglobin were indistinguishable by electrical charge and molecular mass, regardless of the source, indicating molecular identity. Cross-linking of soluble proteins with cupric 1,10-phenanthroline resulted in the formaton of a cross-linked product of plakoglobin with similar physical properties as the native approximately equal to 7-S particle, which is compatible with the interpretation that the soluble plakoglobin particle is a dimer. While a major proportion of the plakoglobin in the desmosomal plaque was resistant to various extraction procedures, plakoglobin present in the plaques of non-desmosome-containing cells and tissues was readily extractable under low and high salt conditions. This indicates that differences exist in the binding of plakoglobin to desmosomal plaques and the plaques of non-demosomal junctions.
D000818:Animals; D002417:Cattle; D002460:Cell Line; D002499:Centrifugation, Density Gradient; D002850:Chromatography, Gel; D003598:Cytoskeletal Proteins; D051180:Desmoplakins; D003896:Desmosomes / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004727:Endothelium / Q000032:analysis; D004817:Epidermis / Q000032:analysis; D005347:Fibroblasts / Q000032:analysis; D006801:Humans; D007525:Isoelectric Focusing; D008565:Membrane Proteins / Q000302:isolation & purification*; D051381:Rats; D012995:Solubility; D051185:gamma Catenin
D003598:Cytoskeletal Proteins; D051180:Desmoplakins; D008565:Membrane Proteins; D051185:gamma Catenin