Identification and characterization of a 230-kDa Golgi-associated protein recognized by autoantibodies from a patient with HBV hepatitis

A serum from a patient with HBV hepatitis was found to contain autoantibodies reacting with various mammalian cells, Immunofluorescence staining of cultured cells with the autoantibodies revealed that the antigen was localized at perinuclear regions, where the Golgi markers alpha-mannosidase II and beta-COP were colocalized, The autoantigen disappeared from the perinuclear regions upon incubation with the fungal metabolite brefeldin A, and the immunostainable structures were fragmented into vesicles by treatment with nocodazole. These results strongly indicate that the antigen is localized at the Golgi complex. Immunoblots of cell lysates showed that the autoantibodies recognized a single protein with a molecular mass of 230 kDa in a variety of cell lines, indicating that the 230-kDa antigen is a conserved protein among mammalian species. We designated this protein GCP230 (Golgi complex-associated protein with a molecular mass of 230 kDa). When a post-nuclear fraction was prepared and centrifuged, GCP230 was recovered in both cytosol and membrane fractions. Peripheral interaction of GCP230 with membranes was confirmed by phase separation in Triton X-114 solution and by extraction with sodium carbonate. Taken together, these results indicate that GCP230 is a peripheral membrane protein of the Golgi derived from the cytosol, although its function is not known at present.