The cDNA cloning of conglutinin and identification of liver as a primary site of synthesis of conglutinin in members of the Bovidae

Abstract

Bovine conglutinin is a collagen-like, C-type, plasma lectin which belongs to the group of proteins called 'collectins'. Two inosine-containing oligonucleotides were synthesized, based on the published protein sequence for bovine conglutinin [Lee, Leiby, Allar, Paris, Lerch and Okarma (1991) J. Biol. Chem. 266, 2715-2723], and PCR on target DNA from a bovine liver lambda gt 11 cDNA library yielded a product of the expected size of 210 bp. Screening of the library with this cDNA fragment identified a single positive clone, with an insert of 0.9 kb, coding for bovine conglutinin from residue 70 to the C-terminus. The 5' cDNA sequence, encompassing 150 bp of the 5' non-translated sequence plus the sequence encoding the leader peptide and the N-terminal residues 1-70, was completed by the use of PCR techniques. The cDNA sequence of bovine conglutinin showed 86% identity with that of bovine lung surfactant protein D (SP-D), and the derived amino acid sequence of bovine conglutinin showed 78% identity with that of bovine SP-D, which included complete identity of the leader-peptide sequences. The amino acid sequence derived from the cDNA sequence differs from the published protein sequence at four positions. Northern-blot analysis on total RNA, purified from various tissues from cattle, sheep, humans, rats and mice, showed that a strong signal of approx. 1.8 kb is present in bovine liver RNA. A weak signal of similar size was also observed in sheep liver, but not in human, rat and mouse livers. A weak signal, also of 1.8 kb, is present in the lung RNAs of all the species tested. The signals from the lung tissues are likely to be due to the cross-hybridization of the bovine conglutinin cDNA to the SP-D mRNAs of the respective species. The finding of significant signals in only the bovine and sheep liver RNA samples is indicative that serum conglutinin may be present in significant amounts only in members of the Bovidae (the family encompassing cattle, antelopes, sheep and goats) and closely related species.