Presence of an immunologically-defined class of peri-Golgi vesicles in cultured mammalian cells.
Immunsera of mice injected with clathrin-depleted coated vesicle membranes, purified from rat liver, revealed a preferential labeling of some perinuclear structures by immunofluorescence microscopy on NRK cells. Subsequent production of 4 monoclonal antibodies was achieved. The antigen was strictly located in the Golgi area of the cell but was not an intrinsic element of the Golgi complex. The restricted location of the structures excluded these were lysosomes which appeared more dispersed in these cells. After nocodazole treatment the material was found dispersed in the cytoplasm. This provided a means of distinguishing the antigen from clathrin-coated structures and Golgi intrinsic elements. Immunolocalization at the electron microscope level confirmed the data obtained at the light level. Some peroxidase reaction product was rarely associated with Golgi elements, but predominantly stained small neighboring Golgi vesicles (50 nm diameter), as well as tubulo-elongated structures and some large (500 nm) irregular-shaped vesicles. A 32 kDa molecular weight antigen was characterized by immunopurification from NRK cells metabolically labeled with 35S-Met. This 32 kDa antigen appeared as part of a higher multimolecular membrane component of 300 kDa. A 170 kDa and a 70 kDa components were immunodetected in a semi-purified membrane fraction from rat liver, demonstrating that the antigen was a minor but very antigenic contaminant of the coated vesicle preparation used as immunogen. In conclusion, the labeled peri-Golgi structures may be part of the newly characterized trans-Golgi network and/or of the reticular/vesicular endosomal, prelysosomal structure recently described.
Mangeat
P
P
Centre CNRS-INSERM de Pharmacologie-Endocrinologie, Montpellier, France.
Devilliers
G
G
Regaya
F
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D000918:Antibody Specificity; D002478:Cells, Cultured; D005455:Fluorescent Antibody Technique; D006056:Golgi Apparatus / Q000187:drug effects / Q000648:ultrastructure*; D008099:Liver / Q000648:ultrastructure; D008985:Monensin / Q000494:pharmacology; D015739:Nocodazole / Q000494:pharmacology; D015388:Organelles / Q000187:drug effects / Q000648:ultrastructure*; D051381:Rats; D011919:Rats, Inbred Strains
D000911:Antibodies, Monoclonal; D008985:Monensin; D015739:Nocodazole