Biochemical and functional characterization of three types of coated vesicles in bovine adrenocortical cells: implication in the intracellular traffic.
Three populations of pure coated vesicles from adrenocortical cells, differing in their density, i.e., 1.125-1.155, 1.155-1.175, and 1.175-1.210 g/cm3, are obtained after separation on two successive sucrose-2H2O gradients. They are involved in LDL internalization and in the receptor cycle as confirmed by the presence, in each population, of the LDL receptor. Electron micrographs confirm the existence of three homogeneous populations exhibiting the typical polygonal structure of the clathrin coat. They differ in their size distribution (small, congruent to 70-nm diameter; medium, congruent to 90-nm diameter; large, congruent to 110-nm diameter) and in the organization of clathrin and of the coat proteins as evidenced on electrophoreses carried out under nondenaturing and denaturing conditions. Activity measurements of marker enzymes, phosphodiesterase and galactosyltransferase, suggest that medium coated vesicles might originate from plasma membranes and small ones from the Golgi complex. Large coated vesicles exhibit phosphokinase enzyme and substrate polypeptides different from those of the two other populations, tubulins being the preferred kinase substrates for the small and medium coated vesicles. These kinases are autophosphorylating enzymes and are revealed, by nondenaturing electrophoreses, as different high molecular mass complexes in the three populations. Clathrin and coat proteins are not part of these complexes.
Bomsel
M
M
Equipe de Recherche CNRS 64, UER Biomédicale des Saints Pères, Paris, France.
de Paillerets
C
C
Weintraub
H
H
Alfsen
A
A
D000302:Adrenal Cortex / Q000378:metabolism* / Q000648:ultrastructure; D000818:Animals; D015415:Biomarkers; D002417:Cattle; D002458:Cell Fractionation; D002966:Clathrin / Q000378:metabolism; D003034:Coated Pits, Cell-Membrane / Q000378:metabolism* / Q000648:ultrastructure; D011992:Endosomes / Q000378:metabolism*; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D011973:Receptors, LDL / Q000378:metabolism
D015415:Biomarkers; D002966:Clathrin; D008565:Membrane Proteins; D011973:Receptors, LDL; D011494:Protein Kinases; D010727:Phosphoric Diester Hydrolases