A ROLE FOR RETROSOMES IN INTRACELLULAR CHOLESTEROL TRANSPORT FROM ENDOSOMES TO THE PLASMA-MEMBRANE

The recycling component (retrosome) of the endocytic pathway was evalu ated as a potential vehicle for the recycling of lipoprotein-derived c holesterol and the maintenance of a high concentration of free cholest erol in plasma membranes. Receptor-to-ligand ratios were established i n three distinct endosomal compartments using a recycling receptor (ap olipoprotein B/E) to confirm isolated retrosomes as recycling vesicles . Compositional studies showed that retrosomes have twice the free cho lesterol in their limiting membranes as do the endosomal compartments from which they derive. Furthermore, of the three isolated endosomal f ractions, retrosomes showed the highest ratio of free to esterified ch olesterol derived from injected very low density lipoprotein as well a s the highest free-to-esterified cholesterol mass ratio overall, confi rming endosomal cholesteryl ester hydrolysis and sorting. Endosomal ne utral cholesterol esterase was identified by immunoblot, whereas elect ron microscopy employing membrane cholesterol-specific filipin reveale d a high concentration of cholesterol in appendages that appear to be the formative stage of ertrosomal biogenesis.