Vitamin A uptake from retinol-binding protein in a cell-free system from pigment epithelial cells of bovine retina. Retinol transfer from plasma retinol-binding protein to cytoplasmic retinol-binding protein with retinyl-ester formation as the intermediate step.
We have investigated the steps by which retinol, released from plasma retinol-binding protein (RBP), enters the cells and is accumulated for the most part as a retinyl-ester, only a small fraction of it being present as a complex with cytoplasmic retinol-binding protein (CRBP). For this purpose, we have developed a cell-free system composed of plasma membrane-enriched fractions from bovine retinal pigment epithelium which selectively incorporates exogenous vitamin A when presented as a retinol-RBP complex. Upon incubation in the presence of [3H]retinol-RBP, isolated plasma membrane fractions take up and esterify retinol. A 4-fold reduction of total vitamin A incorporation is observed in conditions which specifically inhibit retinyl-ester formation, thus indicating that the two processes of retinol uptake and esterification are functionally coupled. Evidence is presented that retinol bound to a plasma membrane receptor sharing functional and structural similarities with CRBP is the actual substrate for esterification. Vitamin A accumulation seems to require retinol esterification to allow the recycling of a limited number of free, plasma membrane-associated, retinol receptors. Mobilization of retinol stored as a membrane-bound retinyl-ester is mediated by a membrane-associated hydrolase activity selectively controlled by the level of apo-CRBP which acts as a carrier for the released retinol. Up to 90% of membrane-bound vitamin A is released upon incubation in the presence of apo-CRBP (11 microM) with concomitant formation of retinol-CRBP. The overall process, in which retinol never needs to leave its binding proteins, allows the accumulation of vitamin A in the form of a membrane-bound retinyl-ester and its regulated mobilization as a retinol-CRBP complex.
D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002474:Cell-Free System; D003593:Cytoplasm / Q000378:metabolism; D004951:Esterification; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D012177:Retinol-Binding Proteins / Q000378:metabolism*; D054840:Retinol-Binding Proteins, Cellular; D054839:Retinol-Binding Proteins, Plasma; D014801:Vitamin A / Q000378:metabolism*
D012177:Retinol-Binding Proteins; D054840:Retinol-Binding Proteins, Cellular; D054839:Retinol-Binding Proteins, Plasma; D014801:Vitamin A