Identification and characterization of a new member of the placental prolactin-like protein-C (PLP-C) subfamily, PLP-Cbeta.

We have isolated a complementary DNA (cDNA) clone that encodes a new member of the PRL-like protein-C (PLP-C) subfamily of the PRL gene family. The clone was amplified from a 13.5-day-old mouse conceptus cDNA library by PCR using primers based on conserved regions of PLP-C sequences. The full-length cDNA encodes a predicted protein of 241 residues, which contains a putative signal sequence and 2 putative N-linked glycosylation sites. The predicted protein shares 55-66% amino acid identity with mouse PLP-Calpha and rat PLP-D, PLP-H, PLP-Cv, and PLP-C and also contains 6 homologously positioned cysteine residues. Thus, we named this protein PLP-Cbeta for consistency. We have also isolated rat PLP-Cbeta from rat placenta cDNA library. Surprisingly, two messenger RNA (mRNA) isoforms of rat PLP-Cbeta were isolated: one mRNA (rPLP-Cbeta) encodes a 241-amino acid product, but another mRNA (rPLP-Cbetadelta39) lacks 39 bases that encode for a region rich in aromatic amino acids. The 39-bp region corresponds to exon 3 of other PLP-C subfamily members, such as PLP-Calpha, PLP-Cv, and d/tPRP. It suggests that the two isoforms are probably generated by an alternative splicing from a single gene. RT-PCR analysis revealed that the rPLP-Cbeta form was dominantly expressed in placenta, although both isoforms are coexpressed during placentation. The mouse PLP-Cbeta mRNA expression, which was specific to the placenta, was first detected by Northern analysis on embryonic day 11.5 (E 11.5) and persisted until birth. However, in situ hybridization analysis revealed mPLP-Cbeta expression on E 10.5 in specific trophoblast subsets, such as giant cells and spongiotrophoblast cells. mPLP-Cbeta mRNA was detected in the labyrinthine zone on E 18.5, suggesting that spongiotrophoblast cells had penetrated the labyrinthotrophoblast zone. Consistent with the observed expression in trophoblast giant cells, PLP-Cbeta expression was also detected in in vitro differentiated Rcho-1 cells, which express the trophoblast giant cell phenotype. In summary, overall high amino acid identity (79%), the locations of cysteine residues, and consensus sites for N-linked glycosylation between mouse and rat PLP-Cbeta clearly indicate that PLP-Cbeta is a bona fide member of the PLP-C subfamily. The conservation between mouse and rat, the presence of alternative isoforms, and the pattern of expression during gestation suggest the biological significance of PLP-Cbeta during pregnancy.