Specific expression patterns and cell distribution of ancient and modern PAG in bovine placenta during pregnancy

Abstract

Pregnancy-associated glycoproteins (PAGs) constitute a multigenic family of aspartic proteinases expressed in the trophoblast of the ruminant placenta. In Bos taurus, this family comprises 21 members segregated into ancient and modern phylogenetic groups. Ancient PAGs have been reported to be synthesized throughout the trophoblastic cell layer whereas modern PAGs are produced by binucleate cells of cotyledons. The aim of this study was to investigate modern and ancient PAGs during gestation in cotyledonary and intercotyledonary tissues. To obtain convincing and innovative results despite the high sequence identity shared between PAGs, we designed specific tools such as amplification primers and antibodies. Using real-time RT-PCR, we described the transcript expression of 16 bovine PAGs. Overall, PAGs are characterized by an increase in their expression during gestation. However, we demonstrated a segregation of modern PAGs in cotyledons and of ancient PAGs in the intercotyledonary chorion, except for the ancient PAG2 expressed in cotyledons. By raising specific antibodies against the modern PAG1 and ancient PAG11 and PAG2, we established the expression kinetics of the proteins using western blotting. Immunohistochemistry showed that PAGs were produced by specific cellular populations: PAG1 by binucleate cells in the whole trophoblastic layer, PAG11 was localized in binucleate cells of the intercotyledonary trophoblast and the chorionic plate of the cotyledon, while PAG2 was produced in mononucleate cells of the internal villi of the cotyledon. These results revealed a highly specific regulation of PAG expression and cell localization as a function of their phylogenetic status, suggesting distinct biological functions within placental tissues.

Placental PAGs: gene origins, expression patterns, and use as markers of pregnancy

“…For those PAGs exhibiting this expression pattern, the secreted PAGs accumulated at the microvillar junction in a manner similar to what was also observed at the pig placenta-uterine interface (Wooding et al 2005). It is worth noting that a recent published work by Touzard et al (2013) also investigated the expression of bovine PAG2 by immunolocalization. They observed mononucleated trophoblast staining, but they did not report immunolocalization of their anti-bovine PAG2 peptide antibody to binucleated trophoblasts.…”

“…These data may suggest that a change in cell-specific expression occurs over the course of placental development for this particular PAG. Finally, the report by Touzard et al (2013) also provided some evidence that binucleated trophoblasts may represent distinct cell populations that are distinguishable based on the PAGs they produce. An immunolocalization experiment was carried out in bovine placenta with antibodies against PAG1 and PAG11.…”

“…As a group, the ancient PAGs are usually detectable earlier in development (authors' unpublished data) and generally at greater abundance compared with the modern PAGs (Touzard et al 2013). The message profiles of individual PAGs can differ substantially from one another; some are elevated early in pregnancy, while others do not become abundant until later (Patel et al 2004b, Telugu et al 2009, Touzard et al 2013. A further level of complexity arises from the fact that ruminant PAGs vary in their cell-specific expression (mononucleated vs binucleated trophoblast giant cells) and in the location(s) in which they accumulate after secretion.…”

“…In contrast, bovine PAG10 message was maximal in the day 280 cotyledonary samples. A recent paper from Touzard et al (2013) sought to further characterize bovine PAG message and protein abundance in both cotyledonary and intercotyledonary chorion. These authors observed that most, but not all, of the modern PAGs were two to four times more abundant in cotyledons compared with intercotyledonary chorion.…”

Morphological Characterization of Basally Located Uninucleate Trophoblast Cells as Precursors of Bovine Binucleate Trophoblast Giant Cells

“…Even if the number of animals used in this study is relatively low, we consider the collected information as representative to answer the question posed in this study. This appears to be justified, first by the very de- tailed information obtained by the 2 serial-sectioning methods used in this study and, second by the fact that there are no indications about major, gestational stagedependent morphological variations within each trophoblast cell type from gd 45 until term [Björkmann, 1969], even though protein expression has been shown to vary within gestation [Green et al, 2000;Touzard et al, 2013]. A lack of contact to the BM is generally considered to be a distinctive feature of ruminant TGCs [Hoffman and Wooding, 1993;Wooding and Burton, 2008].…”