Clathrin-coated vesicles purified from a human B lymphocyte cell line contain newly synthesized glycoproteins

Clathrin-coated vesicles were purified from the cultured human B lymphocyte cell line RPMI 1788. These preparations were shown to be substantially free of contaminating membranes by the following criteria: (a) polypeptide composition, as assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, (b) electron microscopy, (c) lack of binding of intact vesicles to concanavalin A-Sepharose, (d) migration of coated vesicles as a single major peak when applied to a Sephacryl S-1000 gel filtration column. An association between HLA-A, -B, and -DR antigens and immunoglobulins with coated vesicle fractions was demonstrated by lentil lectin affinity chromatography and by immunoprecipitation using antisera directed against these glycoproteins. [35S]Methionine pulse-chase experiments revealed that there was a kinetic correlation between the expression of newly synthesized glycoproteins and their presence in coated vesicles. These results indicate that coated vesicles may be involved in the transport of cell surface and secreted eukaryotic glycoproteins through the cell to their final destinations at the plasma membrane or in the extra-cellular medium.