Selective cloning of cDNA for secretory proteins of early embryos. Identification of a transiently expressed kunitz domain protein from preimplantation sheep trophoblast.

Abstract

The preimplantation ovine conceptus transiently secretes several proteins, including a type I interferon (IFN-tau), that are likely involved in establishment of pregnancy. A method has been developed to identify proteins produced simultaneously with IFN-tau. An antiserum against total ovine conceptus secretory proteins, from which IFN-tau and proteins common to maternal uterine tract secretions had first been removed, was used to immunoscreen cDNA libraries created from mRNA of days 13 and 15 ovine conceptuses. This approach has allowed several unique cDNA to be identified, including one particularly abundant transcript for a novel member of the Kunitz family of serine protease inhibitors. This cDNA encodes a 265-amino acid protein with a 20-amino acid signal sequence. A 64-amino acid Kunitz domain occupies the carboxyl terminus. It is preceded by two similar repeats of 84 residues that bear no obvious similarity to any sequences present in the protein data banks. The protein present in conceptus secretions (M(r) of approximately 14,000) represents only the carboxyl terminus of the molecule. The mRNA for this putative proteinase inhibitor was confined to trophectoderm and was highly expressed for only a few days (approximately 13-18) of development. A similar transcript was detected during the days 17-21 period in cattle embryos. Despite their high expression, no proteinase-inhibitory activity can so far be ascribed to either the ovine or bovine proteins. The P1 residue, an asparagine, is not represented in any other known Kunitz inhibitors.