Identification of GPR99 Protein as a Potential Third Cysteinyl Leukotriene Receptor with a Preference for Leukotriene E4 Ligand

Abstract

Background: A most stable cysteinyl leukotriene, LTE 4 , mediates vascular permeability in mice lacking the two known receptors. Results: GPR99 deficiency abolishes LTE 4 -induced vascular permeability in mice also lacking the two known receptors. Conclusion: GPR99 is a potential third cysteinyl leukotriene receptor. Significance: GPR99 may be a therapeutic target for inflammatory diseases.The cysteinyl leukotrienes (cys-LTs), leukotriene C 4 (LTC 4 ), a conjugation product of glutathione and eicosatetraenoic acid, and its metabolites, LTD 4 and LTE 4 , are lipid mediators of smooth muscle constriction and inflammation in asthma. LTD 4 is the most potent ligand for the type 1 cys-LT receptor (CysLT 1 R), and LTC 4 and LTD 4 have similar lesser potency for CysLT 2 R, whereas LTE 4 has little potency for either receptor. Cysltr1/Cysltr2 ؊/؊ mice, lacking the two defined receptors, exhibited a comparable dose-dependent vascular leak to intradermal injection of LTC 4 or LTD 4 and an augmented response to LTE 4 as compared with WT mice. As LTE 4 retains a cysteine residue and might provide recognition via a dicarboxylic acid structure, we screened cDNAs within the P2Y nucleotide receptor family containing CysLTRs and dicarboxylic acid receptors with trans-activator reporter gene assays. GPR99, previously described as an oxoglutarate receptor (Oxgr1), showed both a functional and a binding response to LTE 4 in these transfectants. We generated Gpr99 ؊/؊ and Gpr99/Cysltr1/Cysltr2 ؊/؊ mice for comparison with WT and Cysltr1/Cysltr2 ؊/؊ mice. Strikingly, GPR99 deficiency in the Cysltr1/Cysltr2 ؊/؊ mice virtually eliminated the vascular leak in response to the cys-LT ligands, indicating GPR99 as a potential CysLT 3 R active in the Cysltr1/Cysltr2 ؊/؊ mice. Importantly, the Gpr99 ؊/؊ mice showed a dose-dependent loss of LTE 4 -mediated vascular permeability, but not to LTC 4 or LTD 4 , revealing a preference of GPR99 for LTE 4 even when CysLT 1 R is present. As LTE 4 is the predominant cys-LT species in inflamed tissues, GPR99 may provide a new therapeutic target.Leukotriene C 4 (LTC 4 ), 2 LTD 4 , and LTE 4 , collectively called cysteinyl leukotrienes (cys-LTs), are proinflammatory mediators derived from arachidonic acid through the 5-lipoxygenase/ LTC 4 synthase pathway (1, 2). Intracellularly synthesized LTC 4 is exported via multidrug resistance-associated proteins and is rapidly metabolized in the extracellular space by cleavage removal of glutamic acid and then glycine to LTD 4 and LTE 4 , respectively. The type 1 cys-LT receptor (CysLT 1 R) is the high affinity receptor for LTD 4 , whereas the type 2 receptor (CysLT 2 R) can bind both LTC 4 and LTD 4 with one-log less affinity than that of CysLT 1 R for LTD 4 in transfected cells (3, 4). The cys-LTs are mediators of bronchial asthma on the basis of their potent bronchoconstrictive activity via the CysLT 1 R in pharmacologic studies and the clinical effectiveness of CysLT 1 R antagonists (5, 6).LTE 4 , the most abundant cys-LT at sites of inflammation due to its stab...

Leukotriene E4elicits respiratory epithelial cell mucin release through the G-protein–coupled receptor, GPR99

“…As GPR99 belongs to the same nucleotide P2Y receptor family as CysLT 1 R and CysLT 2 R and recognizes dicarboxylic acids, it was identified as a candidate LTE 4 receptor and subsequently demonstrated to mediate cutaneous swelling in response to low doses of LTE 4 (25). Our findings for GPR99 control of mucin secretion and EpC homeostasis indicate a central role for this CysLTR at low doses of LTE 4 .…”

“…LTE 4 , the stable cysLT (15)(16)(17)(18), is a weak agonist for CysLT 1 R and CysLT 2 R in transfected cells (5,19), but elicits airflow obstruction in patients with asthma (20)(21)(22). Moreover, LTE 4 has comparable activity to LTC 4 and LTD 4 in eliciting a wheal and flare response in human skin (23), and LTE 4 elicits cutaneous vascular permeability in mice lacking both CysLT 1 R and CysLT 2 R, suggesting the existence of a high-affinity receptor for LTE 4 , which was recently identified as GPR99 (24,25). However, the mechanism by which LTE 4 induces lung pathobiology and the role of GPR99 remain poorly understood.…”

Prostaglandin E2deficiency causes a phenotype of aspirin sensitivity that depends on platelets and cysteinyl leukotrienes

“…Oral administration of zileuton to subjects with AERD blocked the release of tryptase into the nasal lavage fluid following intranasal Lys-ASA challenge (50). MCs express all of the known CysLTRs (14)(15)(16)51), and blockade of CysLT 1 R with montelukast mirrored the effect of zileuton on the release of MC activation products in our study. Thus, AERD involves a prominent autocrine and/or paracrine signaling loop in which cysLTs promote MC activation.…”

“…After export, LTC 4 is converted sequentially to LTD 4 and LTE 4 . CysLTs induce bronchoconstriction (10,11), tissue eosinophilia (12), and remodeling (13) through G-protein-coupled receptors (GPCRs) expressed by structural and hematopoietic cells (14)(15)(16). Individuals with AERD display higher urinary levels of LTE 4 than do aspirin-tolerant asthmatic (ATA) control subjects (17).…”

Phenotype-based Discovery of 2-[(E)-2-(Quinolin-2-yl)vinyl]phenol as a Novel Regulator of Ocular Angiogenesis

“…Finally, the chemical structure of quininib (molecular weight, 283.75 g/mol) extensively overlaps with a portion of the much larger, clinically approved CysLT 1 antagonist montelukast (Merck, MW 586.18g/mol) (32). Recently montelukast has been reported to reduce neuroinflammation in an aged rat model, acting via inhibition of a separate cysteinyl leukotriene receptor, GPR17 (56), and a fourth cysteinyl leukotriene receptor, GPR99, has been proposed (26). In genetic loss-of function models, cysteinyl leukotriene receptors demonstrate cognate compensatory up-regulation (37,39,57).…”