ChemiSCREEN™ CXCR2 Calcium-Optimized FLIPR Cell Line

ChemiSCREEN™ CXCR2 Calcium-Optimized FLIPR Cell Line

• ChemiScreen
• Chemicon (Millipore)

2 vials

CXCR2 is a 7-TM G-protein coupled receptor that binds to the chemokines GROalpha, GRObeta, GROgamma, IL-8, ENA-78, NAP-2 and GCP-2 (Olson and Ley, 2002). Neutrophils, mast cells and microvascular endothelial cells express CXCR2, and interactions of CXCR2 with its ligands promotes chemotaxis of these cell types (Heidemann et al., 2003; Nilsson et al., 1999; White et al., 1998). Studies with mice lacking CXCR2 indicate that CXCR2 promotes growth of primary tumors and secondary metastases (Keane et al., 2004), and plays an essential role in hyperoxia-induced lung injury (Sue et al., 2004). In addition, cytomegalovirus encodes a CXCR2-binding chemokine, vCXC-1, that promotes neutrophil migration to infected cells (Penfold et al., 1999). Chemicon's cloned human CXCR2-expressing cell line is made in the Chem-1 host, which supports high levels of recombinant CXCR2 expression on the cell surface and contains high levels of the promiscuous G protein Galpha15 to couple the receptor to the calcium signaling pathway. Thus, the cell line is an ideal tool for screening for antagonists of interactions between CXCR2 and its ligands.

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 a receptor for interleukin 8 (IL8). It binds to IL8 with high affinity, and transduces the signal through a G-protein activated second messenger system. This receptor also binds to chemokine (C-X-C motif) ligand 1 (CXCL1/MGSA), a protein with melanoma growth stimulating activity, and has been shown to be a major component required for serum-dependent melanoma cell growth. This receptor mediates neutrophil migration to sites of inflammation. The angiogenic effects of IL8 in intestinal microvascular endothelial cells are found to be mediated by this receptor. Knockout studies in mice suggested that this receptor controls the positioning of oligodendrocyte precursors in developing spinal cord by arresting their migration. This gene, IL8RA, a gene encoding another high affinity IL8 receptor, as well as IL8RBP, a pseudogene of IL8RB, form a gene cluster in a region mapped to chromosome 2q33-q36.

FUNCTION: SwissProt: P25025 # Receptor to interleukin-8, which is a powerful neutrophils chemotactic factor. Binding of IL-8 to the receptor causes activation of neutrophils. This response is mediated via a G-protein that activate a phosphatidylinositol-calcium second messenger system. This receptor binds to IL-8 with a high affinity and to GRO/MGSA and NAP-2 also with a high affinity.
SIZE: 360 amino acids; 40759 Da
SUBCELLULAR LOCATION: Cell membrane; Multi-pass membrane protein.
PTM: Phosphorylated upon ligand binding; which is required for desensitization.
SIMILARITY: SwissProt: P25025 ## Belongs to the G-protein coupled receptor 1 family.

Human

Chemicon

CXCR2 / IL-8b

Cells are frozen at 2 x 10⁶ cells/mL in DMEM/20% fetal bovine serum containing 10% DMSO. Cell line tests negative for mycoplasma.

peptide

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 growth media, and replace media after 8-24 h. Cells are typically passaged 1:10 every 3-4 days, and should be passaged at least once after thawing prior to use in calcium flux assays.

Chemokine

A

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• IL8RB
• CD182
• CMKAR2
• CXCR-2
• IL8RA
• IL8R2
• CXCR2
• CDw128b

GPCRs

ChemiSCREEN™ CXCR2 Calcium-Optimized FLIPR Cell Line

2 x 10⁶ cells/mL

"SPECIFICATIONS: EC50 for calcium mobilization by IL-8: ~ 7.2 nM; GROα: ~5.6 nM
Signal/noise ratio at GROα E_max: 568
Z' = 0.90 with EC50 concentration of GROα

HOST CELLS: Chem-1, an adherent cell line expressing the promiscuous G-protein, Gα15.

TRANSFECTION: Full-length human CXCR2 cDNA (Accession Number: M73969)

GROWTH MEDIA: 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"

Chem-1

GPCR Cell Lines

Chem-1

Cell Line

P25025

2x10⁶ cells/vial

NM_001557.2