INTRACELLULAR-TRANSPORT OF LOW-DENSITY-LIPOPROTEIN DERIVED FREE-CHOLESTEROL BEGINS AT CLATHRIN-COATED PITS AND TERMINATES AT CELL-SURFACE CAVEOLAE

Free cholesterol (FC) is selectively internalized from low-density lip oprotein (LDL) by confluent fibroblast monolayers (Fielding & Fielding (1995) Biochemistry 34, 14237-14244), The kinetics of transport of LD L-derived H-3-FC within the cell were studied by density-gradient ultr acentrifugal fractionation and in terms of the effects of inhibitors o f endocytosis and intracellular transport. By these criteria, the init ial uptake of LDL-FC was mediated by the cell-surface clathrin-coated pits. FC label then appeared in clathrin-coated dense vesicles. Uncoat ing of clathrin from these vesicles led to the appearance of label in a light density fraction and, subsequently, in an intermediate density fraction coincident with protein markers of the trans-Golgi network i n these cells. H-3-FC was finally transported to the plasma membrane v ia a temperature-sensitive, probably microtubule-dependent pathway. Th ese data are consistent with a role for the trans-Golgi network as an intermediate compartment in intracellular FC transport. They provide f urther evidence of a role for cell-surface caveolae in FC efflux.