Ligand-dependent secretion of rat retinol-binding protein expressed in HeLa cells.
A minigene encoding rat retinol-binding protein (RBP) was transfected into HeLa cells, which do not express endogenous RBP, transthyretin, or cellular retinol-binding protein. The HeLa cells manufactured and secreted the transfected gene product, demonstrating that RBP-transthyretin assembly is not a requirement for the secretion of RBP. When HeLa cells were grown under vitamin A-deficient conditions, RBP accumulated in the endoplasmic reticulum. Both serum and retinol stimulated secretion of RBP in a concentration-dependent manner. The retinol-regulated secretion occurred also after protein synthesis had been blocked by cycloheximide. Addition of holo-RBP or retinal, but not retinoic acid, stimulated secretion of RBP. Thus, an in vitro model system that resembles the rat hepatocyte in vivo with regard to the known regulation of RBP secretion has been established in a human cell line of extrahepatic origin. It can be concluded that cellular retinol-binding protein is not required for the transfer of retinol to RBP and that the mechanism whereby retinol controls the intracellular transport of RBP is neither specific for tissues synthesizing RBP nor species-specific. To investigate the structural properties responsible for the endoplasmic reticulum retention of RBP in the absence of its ligand, a cDNA encoding chicken purpurin, a protein that is 50% identical to RBP and that binds retinol, was expressed in HeLa cells. In contrast to RBP, purpurin was not retained in vitamin A-deficient HeLa cells.
Melhus
H
H
Department of Cell Research, Uppsala University, Sweden.
Laurent
B
B
Rask
L
L
Peterson
P A
PA
D000818:Animals; D000880:Anthraquinones; D002645:Chickens; D003513:Cycloheximide / Q000494:pharmacology; D004247:DNA / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D037102:Lectins / Q000235:genetics / Q000378:metabolism; D008024:Ligands; D008856:Microscopy, Fluorescence; D011228:Prealbumin / Q000235:genetics / Q000378:metabolism; D051381:Rats; D012177:Retinol-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D054840:Retinol-Binding Proteins, Cellular; D014162:Transfection; D014801:Vitamin A / Q000378:metabolism*; D001613:beta 2-Microglobulin / Q000378:metabolism
D000880:Anthraquinones; D037102:Lectins; D008024:Ligands; D011228:Prealbumin; D012177:Retinol-Binding Proteins; D054840:Retinol-Binding Proteins, Cellular; D001613:beta 2-Microglobulin; D014801:Vitamin A; D004247:DNA; D003513:Cycloheximide; C511975:purpurin anthraquinone