Synthesis and secretion of retinol-binding protein by cultured rat Sertoli cells

We report here that retinol-binding protein (RBP) is synthesized and secreted by rat Sertoli cells in culture. This was demonstrated in four ways. First, transthyretin (TTR) bound to Sepharose 4B retained a labeled protein from media collected from Sertoli cells provided with 35S-methionine, under the same conditions as authentic RBP is bound. The protein was co-eluted with authentic RBP by pure water. Second, this same radioactive protein co-eluted with pure RBP upon gel filtration. Third, when subjected to SDS-PAGE, the protein again migrated with pure RBP, as shown by radioautography. Finally, Sertoli cells were incubated with 35S-cysteine and the conditioned medium was put over a TTR-Sepharose column to isolate the radioactive protein, as characterized above. Cysteine residues were oxidized to cysteic acid residues, and the protein was submitted for sequencing through the first ten residues. Radioactivity was located only in the fourth residue, where a cysteine residue is found in rat RBP. This indicates that RBP is secreted by the Sertoli cell and may serve as the carrier of retinol to the developing germ cells, which are known to be dependent upon vitamin A.