Specific uptake of retinol-binding protein by variant F9 cell lines.
Serum retinol-binding protein (RBP) specifically binds to and is internalized by F9 embryonal carcinoma cells. Monolayers of F9 cells were differentiated into a primitive endoderm stage by addition of retinoic acid. Fluorescein-derivatized or radiolabeled RBP associated with F9 cell monolayers at 37 degrees C in a time- and concentration-dependent manner. Competition by simultaneous incubation with excess unlabeled RBP indicated that this association was specific and saturable; the apparent dissociation constant was 200-300 nM using either tracer. At 37 degrees C, over 80% of the cell-associated RBP was internalized, and only a small fraction was bound to the cell surface; fluorescence microscopy indicated that internalized RBP was in small vesicles within the cytoplasm. Internalized RBP was subsequently degraded and released from the cell in an acid-soluble form. Parental F9 cells were heterogeneous in their ability to associate with RBP. Random subcloning identified natural variant F9 cell lines which did, or did not, express this biological activity upon retinoic acid-induced differentiation. The clonal nature of the capacity for RBP uptake suggests that this specific internalization is a heritable trait. Together, these observations provide strong evidence that RBP uptake occurs by a receptor-mediated process in F9 cells. The cycle of RBP internalization and degradation by F9 cells bearing specific RBP receptors may provide a regulable mechanism for the cellular accumulation of serum retinol.
Matarese
V
V
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142.
Lodish
H F
HF
D000818:Animals; D001667:Binding, Competitive; D002454:Cell Differentiation / Q000187:drug effects; D054278:Embryonal Carcinoma Stem Cells; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D014644:Genetic Variation; D051379:Mice; D008856:Microscopy, Fluorescence; D014411:Neoplastic Stem Cells / Q000378:metabolism*; D012177:Retinol-Binding Proteins / Q000378:metabolism*; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D005456:Fluorescent Dyes; D012177:Retinol-Binding Proteins; D014212:Tretinoin; D016650:Fluorescein-5-isothiocyanate