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PMID:1400343

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Citation

Kooy, J, Toh, BH, Pettitt, JM, Erlich, R and Gleeson, PA (1992) Human autoantibodies as reagents to conserved Golgi components. Characterization of a peripheral, 230-kDa compartment-specific Golgi protein. J. Biol. Chem. 267:20255-63

Abstract

We have used a serum from a patient with Sjögren's syndrome containing high titer (100,000) anti-Golgi autoantibodies and lower titer (20,000) anti-nuclear autoantibodies to characterize the Golgi complex. The Sjögren's syndrome serum immunoprecipitated a number of components of molecular mass 35-230 kDa from detergent extracts of [35S]methionine-labeled HeLa cells; at high dilution, the serum precipitated one major 230-kDa component. Using the Sjögren's syndrome serum, cDNA clones encoding the Golgi autoantigen were isolated from a lambda gt11 HeLa cell cDNA library. Autoantibodies from the Sjögren's syndrome serum, affinity purified from a recombinant bacterial fusion protein generated from one of the cDNA clones, showed Golgi staining of human, mouse, and chicken cells by immunofluorescence. The purified autoantibodies immunoprecipitated and immunoblotted a 230-kDa component. A rabbit antiserum raised to the recombinant fusion protein specifically stained the Golgi complex by immunofluorescence and reacted with a 230-kDa protein by immunoprecipitation and immunoblotting. The 230-kDa protein was recovered in both the 100,000 x g sedimentable and soluble fractions in cell lysates and in the aqueous phase of Triton X-114 extracts. The 230-kDa autoantigen was dissociated from the Golgi complex by 15-min brefeldin A treatment, dissociation kinetics similar to that of mannosidase II. However, unlike mannosidase II, autoantigen staining was markedly reduced after drug treatment. Removal of brefeldin A resulted in reassociation of the autoantigen with the Golgi complex. The epitopes recognized by the affinity purified human and rabbit antibodies were ultrastructurally localized to the cytosolic face of one side of the Golgi stack, probably the trans-face. Taken together, the 230-kDa protein is a conserved, peripheral membrane component specifically associated with one Golgi compartment. We suggest that this peripheral Golgi protein may have a role in the compartment-specific structural organization of the Golgi or in sorting and transport of proteins.

Links

PubMed

Keywords

3T3 Cells; Animals; Autoantibodies/immunology; Autoantigens/analysis; Autoantigens/immunology; Brefeldin A; Cyclopentanes/pharmacology; Fluorescent Antibody Technique; Golgi Apparatus/chemistry; Golgi Apparatus/immunology; HeLa Cells; Humans; Immunoenzyme Techniques; Membrane Proteins/analysis; Membrane Proteins/immunology; Mice; Precipitin Tests; Sjogren's Syndrome/blood; Sjogren's Syndrome/immunology

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

HUMAN:GOGA4

located_in

GO:0005802: trans-Golgi network

ECO:0000304: author statement supported by traceable reference used in manual assertion

C

Seeded From UniProt

complete

HUMAN:GOGA4

part_of

GO:0005802: trans-Golgi network

ECO:0000304: author statement supported by traceable reference used in manual assertion

C

Seeded From UniProt

complete

Notes

See also

References

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