CONSTITUTIVE SECRETION OF BETA-TRACE PROTEIN BY CULTIVATED PORCINE CHOROID-PLEXUS EPITHELIAL-CELLS - ELUCIDATION OF ITS COMPLETE AMINO-ACIDAND CDNA SEQUENCES

Primary porcine choroid plexus epithelial cells cultivated in chemical ly defined medium maintain their epithelial characteristics and form c onfluent monolayers. They produce a fluid the composition of which res embles cerebrospinal fluid. The present study demonstrates constitutiv e secretion of large amounts of beta-trace protein. This intrathecally synthesized protein is a prominent polypeptide constituent of natural cerebrospinal fluid. According to the identity of amino acid sequence s it has previously been tentatively identified as a prostaglandin-D s ynthase and as a member of the lipocalin protein family. beta-Trace wa s purified from cell culture supernatants and was subjected to tryptic digestion and amino acid sequencing of the resulting peptides. The co mplete primary structure of the protein was obtained by additional iso lation of the cDNA from cultured epithelial cells. The porcine 163-ami no acid polypeptide showed 69% identity with the human beta-trace and contained two N-glycosylation sites occupied by complex-type oligosacc harides as is the case for the human protein. The amino acid sequences around the N-glycosylation sites of mammalian beta-trace proteins (po rcine, human, murine, and rat) were highly conserved. The nucleotide s equence was found to be less conserved; the porcine cDNA had a strikin gly high GC-content (67%). The constitutive secretion of beta-trace pr otein from the in vitro cultivated porcine choroid plexus epithelial c ells demonstrates that the cells have retained their major in vivo phy siological properties: secretion of cerebrospinal fluid proteins. Ther efore, this in vitro culture system may be used as a versatile tool fo r studying the regulation of the formation of cerebrospinal fluid. (C) 1996 Wiley-Liss, Inc.