Decidual prolactin-related protein: heterologous expression and characterization.

The prolactin (PRL) family consists of a collection of genes expressed in the uterus, placenta and anterior pituitary. These cytokines/hormones participate in the control of maternal-fetal adaptations to pregnancy. In this report, we establish the presence of three new members of the PRL family. Novel expressed sequence tags (ESTs) with homology to PRL were isolated from embryonic and placental cDNA libraries. The cDNAs were sequenced and compared with those of other members of the PRL family. The three new cDNAs were assigned to the PRL family on the basis of sequence similarities and were referred to as PRL-like protein-J (PLP-J), PRL-like protein-K (PLP-K) and PRL-like protein-M (PLP-M). Both rat and mouse PLP-J cDNAs were identified. Rat PLP-J cDNA encodes for a predicted 211 amino acid protein containing a 29 amino acid signal peptide and two putative N-linked glycosylation sites, whereas the mouse PLP-J cDNA encodes for a 212 amino acid protein containing a 29 amino acid signal peptide with a single N-linked glycosylation site. Rat and mouse PLP-J proteins share approximately 79% and 70% nucleotide and amino acid sequence identity, respectively. A full-length rat PLP-K cDNA and a partial tentative mouse PLP-K cDNA were identified. The rat PLP-K cDNA encodes for a predicted 228 amino acid protein containing a 31 amino acid signal peptide and one putative N-linked glycosylation site; the mouse PLP-M cDNA encodes for a predicted 228 amino acid protein containing a 28 amino acid signal peptide and one putative N-linked glycosylation site. Genes for PLP-J, PLP-K and PLP-M are situated at the Prl family locus on mouse chromosome 13. PLP-J was exclusively expressed in decidual tissue from both the mouse and rat. PLP-K was expressed in trophoblast cells of the chorioallantoic placenta and showed an apparent species difference. In the mouse, virtually all trophoblast lineages expressed PLP-K, whereas in the rat, PLP-K expression was restricted to the labyrinthine trophoblast cells. Mouse PLP-M expression was restricted to the junctional zone of the chorioallantoic placenta. In summary, we have identified three new members of the rodent PRL gene family that are expressed in uterine and placental structures. Future experimentation is needed to determine the specific roles of each of these ligands in the biology of pregnancy.

Section: Plp-jmentioning

confidence: 97%

Three novel paralogs of the rodent prolactin gene family

Dai

Wang²,

Liu³

et al. 2000

“…In rodents, d/tPRP has been reported to be capable of binding heparin-containing molecules (Rasmussen et al 1996). Despite the abundant accumulation of d/tPRP in decidual tissues, d/tPRP does not present in maternal serum at a detectable level.…”

Section: Discussionmentioning

confidence: 99%

“…Binding to extracellular matrix (ECM) components has been reported for several cytokines and growth factors, such as oncostatin M , basic fibroblast growth factor (Faham et al 1998), platelet-derived growth factor (Somasundaram & Schuppan 1996), hepatocyte growth factor (Lyon et al 1994), transforming growth factor-b 1 (Paralkar et al 1991), tumor necrosis factor-a and interleukin-2 (Somasundaram et al 2000), interleukin-7 (Ariel et al 1997), and a member of the rat PRL family, decidual/trophoblast PRP (d/t PRP; Rasmussen et al 1996, Wang et al 2000. The purpose of the present study was to explore the target sites of bPL, bPRP-I, and bPRP-VI in situ and to dissect the targetbinding characteristics of bPRP-I.…”

Section: Introductionmentioning

confidence: 99%

Bovine prolactin-related protein-I is anchored to the extracellular matrix through interactions with type IV collagen

Takahashi¹,

Soares²,

Hashizume³

2007

The bovine placenta produces an array of proteins structurally similar to pituitary prolactin (PRL). At least ten genes of the bovine placental PRL family, including bovine placental lactogen (bPL) and ten bovine PRL-related protein-I to -X (bPRP-I to -X), encode hormones/cytokines predicted to be involved in the establishment and maintenance of pregnancy. Targets and biological roles for most members of the bovine PRL family have yet to be specified. This study focused on three members of bovine PRL family, bPL, bPRP-I, and bPRP-VI. An alkaline phosphatase (AP) tagging strategy was used to monitor interactions of the ligands with their targets. AP-bPRP-I and AP-bPRP-VI specifically bound to tissue sections of the bovine placentome. AP-bPRP-I and AP-bPRP-VI binding within the placentome mimicked the distribution of the extracellular matrix (ECM). Consequently, AP fusion protein binding to individual ECM components (heparin, laminin, fibronectin, type I collagen, and type IV collagen) was evaluated. AP-bPRP-I specifically bound to type IV collagen, but not to the other ECM components. AP-bPRP-VI exhibited weak interactions with ECM components, while AP-bPL and AP did not show significant binding to any of the ECM components. Binding of AP-bPRP-I to type IV collagen was concentration-dependent, influenced by salt concentrations, and specific to the N-terminal cross-linking domain (7S) of type IV collagen but not its triple-helical domain. The interaction of bPRP-I with type IV collagen suggests that bPRP-I accumulates in the ECM where it likely acts on cells traversing the bovine placentome.

“…Conditioned culture medium and stromal cell extracts were tested for dPRP expression by Western blotting (Rasmussen et al 1996). As a positive control, CHO cells were stably transfected with the pMSXND expression vector containing the rat dPRP (Rasmussen et al 1996).…”

Section: Western Blot Analysis Of Endogenous Dprpmentioning

confidence: 99%

“…Fractionated proteins were transferred to a nitrocellulose membrane at 4 C for 18 h. Non-specific binding sites were blocked using standard methods (Jones et al 2000). The proteins were reacted with anti-dPRP antibody (1:500) (Rasmussen et al 1996) for 90 min at 22 C. The blots were washed and reacted with goat anti-rabbit alkaline phosphatase secondary antibody (1:1000) (Jackson ImmunoResearch, West Grove, PA, USA). Bound antibody was detected by incubating the blots with 0·3 mg/ml nitroblue tetrazolium and 0·15 mg/ml 5-bromo-4-chloro-3-indolyl phosphate in 0·1 M NaHCO 3 , 10 mM MgCl 2 , pH 9·8.…”

Section: Western Blot Analysis Of Endogenous Dprpmentioning

confidence: 99%

Stimulation of a rat uterine stromal cell line in culture reveals a molecular switch for endocrine-dependent differentiation

Rider¹,

Potapova²,

Dai³

et al. 2005

Self Cite

Differentiation of uterine stromal cells is critical for the establishment of pregnancy. This study had two purposes: (i) to validate the use of the UIII rat uterine stromal cell model for investigating mechanisms underlying decidual cell differentiation, and (ii) to use this cell model to identify a molecular switch for cellular entry into the decidual cell differentiation pathway. Quiescent rat uterine stromal cells were transfected with a 500 bp segment of the decidual prolactin-related protein (dPRP) promoter ligated to a luciferase reporter gene. Cells were incubated in low-serum medium, or in low-serum medium containing progesterone (1 µM), estradiol 17-(10 nM), cholera toxin (10 ng/ml) and interleukin-11 (10 ng/ml). Protein extracts were collected 48 h later and luciferase was measured in the cellular lysates. Cholera toxin and interleukin-11 stimulated luciferase expression (P<0·05) and addition of sex steroids further increased (P<0·05) dPRP promoter activity. Stromal cells did not proliferate (P>0·05) under differentiation conditions. Deletion analysis of the dPRP promoter revealed maximal luciferase expression between 250 and 500 bp relative to the transcription start site. Comparison of cyclin E/Cdk2 activity between proliferating and differentiating cells showed a 3-fold increase (P<0·05) at 12 h in differentiating cells. The results suggest that cyclin E/Cdk2 serves as a molecular switch for uterine stromal cell entry into the decidual cell differentiation pathway.