PARENT SESSION

(W558) IDENTIFICATION OF PREGNANCY-ASSOCIATED GLYCOPROTEINS (PAGs) EXPRESSED BY THE PREIMPLANTATION CONCEPTUS OF DOMESTIC RUMINANTS.

Garbayo, Juana ¹, Serrano, Beatriz¹, Aguilar, Begonia¹, Lopez-Gatius, Fernando², ¹ Centro de Investigacion y Tecnologia Agroalimentaria (CITA), Zaragoza, Spain² University of Lleida, Lleida, Spain

ABSTRACT- PAGs are a multigene family related to aspartic proteinases, expressed in the outer cell layer of the placenta of Artiodactyla. Multiple PAGs have been cloned from ruminants and can be classified into two main groupings. The so-called PAG I group, expressed in the binucleate cells of the trophectoderm, is a rapidly-evolving group, undergoing diversification at a rapid pace, possibly related to adquiring different peptide binding capabilities. The PAG II group is of more ancient origin, closer to active aspartic proteinases and expressed throughout trophectoderm. The aim of this study was to detect onset of PAG expression by RT-PCR in domestic ruminants by using preimplantational conceptuses and to identify first PAG transcripts to be expressed. A general PAG 5′primer and PAG-specific 3′primers were used to check for PAG expression by days 12 and 14 ovine; days 12, 14, 15, 16 and 18 caprine; and days 15 and 18 bovine conceptuses. PCR products were analysed by restriction digest and subcloned. About 10 colonies from each PCR product were selected and partially sequenced. Similar sequences were aligned and the consensus sequence was checked for identity/similarity in the Gene Bank. PAG expression was first detected from day 12 (ovine), day 16 (caprine) and day 15 (bovine) conceptuses. A single PAG transcript, ovPAG2 (NM–001009782), belonging to the PAG II group, was detected at day 14 in ovine. Two different PAG transcripts were identified in both day 16 and 18 caprine conceptuses. One was similar to caPAG2, also from the PAG II group, (AF191327; 99.8% nt; 99.2% aa) while the other (caPAG12; accession number AY911497) was distinct from those described previously. caPAG12 was closer to boPAG15 (90% nt, 83% aa). Phylogenetic analysis as well as a conserved site for propeptide cleavage situated this new PAG into the PAG I group. Two different PAGs transcripts were also expressed by day 18 bovine conceptuses. Twenty out of 21 inserts sequenced corresponded to boPAG2, from the PAG II group, (NM–176614) but one was new (boPAG22, accession number AY911498). boPAG22 was closer to boPAG2 (96.4% nt; 94.4% aa). In summary, a limited number of PAGs, mainly belonging to the PAG II group, are expressed in domestic ruminants before implantation. Also, two previously unknown PAG transcripts have been cloned. Supported by INIA grant RTA 02-094.

KEY WORDS: PAG, conceptus, ruminants