ChemiScreen™ Human Recombinant EP₄ Prostanoid Receptor Calcium-Optimized Stable Cell Line

ChemiScreen™ Human Recombinant EP₄ Prostanoid Receptor Calcium-Optimized Stable Cell Line

• ChemiScreen
• Chemicon (Millipore)

2 vials

Full-length human PTGER4 cDNA encoding EP₄.

Prostanoids are a series of arachidonic acid metabolites produced by the action of cyclooxygenase and further modified by isomerases and synthases. Cells rapidly secrete prostanoids after synthesis, whereupon the prostanoids bind to a family of 8 GPCRs to exert their biological effects (Narumiya and FitzGerald, 2001). The prostaglandin PGE² causes pain, vasodilation, immunosuppression of T cells, bone remodeling and promotion of carcinogenesis. Four related GPCRs, EP¹, EP², EP³ and EP⁴, each bind to PGE², but the different G protein coupling status of each receptor leads to distinct biological effects. EP⁴ couples primarily to Gs to increase intracellular cAMP levels. During neonatal development, EP⁴ participates in closure of the ductus arteriosus, a process required for switching circulation from the placenta to the lungs (Nguyen et al., 1997). In addition, EP⁴ mediates PGE²-induced bone formation by promoting osteoblastogenesis, and selective EP⁴ agonists are being evaluated as potential treatments for osteoporosis (Yoshida et al., 2002). Chemicon's cloned human EP⁴-expressing cell line is made in the Chem-1 host, which supports high levels of recombinant EP⁴ expression on the cell surface and contains high levels of the promiscuous G protein Gα15 to couple the receptor to the calcium signaling pathway. Thus, the cell line is an ideal tool for screening for agonists and antagonists at EP⁴.

Calcium flux assay, ligand binding assays

Human

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The protein encoded by this gene is a member of the G-protein coupled receptor family. This protein is one of four receptors identified for prostaglandin E2 (PGE2). This receptor can activate T-cell factor signaling. It has been shown to mediate PGE2 induced expression of early growth response 1 (EGR1), regulate the level and stability of cyclooxygenase-2 mRNA, and lead to the phosphorylation of glycogen synthase kinase-3. Knockout studies in mice suggest that this receptor may be involved in the neonatal adaptation of circulatory system, osteoporosis, as well as initiation of skin immune responses.

FUNCTION: SwissProt: P35408 # Receptor for prostaglandin E2 (PGE2). The activity of this receptor is mediated by G(s) proteins that stimulate adenylate cyclase. Has a relaxing effect on smooth muscle. May play an important role in regulating renal hemodynamics, intestinal epithelial transport, adrenal aldosterone secretion, and uterine function.
SIZE: 488 amino acids; 53119 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
TISSUE SPECIFICITY: High in intestine and in peripheral blood mononuclear cells; low in lung, kidney, thymus, uterus, vasculature and brain. Not found in liver, heart, retina oe skeletal muscle.
SIMILARITY: SwissProt: P35408 ## Belongs to the G-protein coupled receptor 1 family.

Human

EC50 for calcium mobilization by Prostaglandin E2 ~ 13 nM

Chem-1

GPCR Cell Lines

Chemicon

Chem-1

Cells are frozen at 2 x 10⁶ cells/mL in DMEM/20% fetal bovine serum/100 U/ml penicillin and streptomycin/10% DMSO. Cell line tests negative for mycoplasma.

EP4

P35408

non-peptide

Prostanoid

Place cells in liquid nitrogen immediately upon receipt. Maintain frozen in liquid nitrogen for up to 5 years. Thaw cells rapidly by removing from liquid nitrogen and immediately immersing in a 37ºC water bath. Immediately after ice has thawed, sterilize the exterior of the vial with 70% ethanol. Transfer contents of the vial to a T75 flask containing 20 mL growth media, and place in a humidified 37°C incubator with 5% CO₂. After 8-24 h, cells will adhere to the plate, at which time the media should be replaced to remove residual DMSO. Cells are passaged by washing with Ca⁺⁺ and Mg⁺⁺-free HBSS (10 mL/T75), incubating with 0.05% trypsin/0.2 g/L EDTA (1 mL/T75) for 5-10 minutes at 37°C, and rapping the side of the flask to dislodge the cells. Neutralize the trypsin by addition of 4 volumes growth media. Cells are typically passaged 1:10 with every 3-4 days, and should be passaged at least once after thawing prior to use in calcium flux assays.

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A

2 x 10⁶ cells/vial

• PTGER4
• MGC126583
• EP4
• EP4R
• PTGER2

GPCRs

ChemiScreen™ Human Recombinant EP₄ Prostanoid Receptor Calcium-Optimized Stable Cell Line

NM_000958.2

DMEM containing 4.5 g/L glucose/10% heat inactivated fetal bovine serum/1x nonessential amino acids/10 mM HEPES/0.25 mg/ml Geneticin (G418)/100 U/ml each penicillin and streptomycin.