p2y5 is an orphan G protein-coupled receptor that is closely related to the fourth lysophosphatidic acid (LPA) receptor, LPA 4 . Here we report that p2y5 is a novel LPA receptor coupling to the G 13 -Rho signaling pathway. "LPA receptor-null" RH7777 and B103 cells exogenously expressing p2y5 showed [ 3 H]LPA binding, LPA-induced [35 S]guanosine 5-3-O-(thio)triphosphate binding, Rho-dependent alternation of cellular morphology, and G s/13 chimeric protein-mediated cAMP accumulation. LPA-induced contraction of human umbilical vein endothelial cells was suppressed by small interfering RNA knockdown of endogenously expressed p2y5. We also found that 2-acyl-LPA had higher activity to p2y5 than 1-acyl-LPA. A recent study has suggested that p2y5 is an LPA receptor essential for human hair growth. We confirmed that p2y5 is a functional LPA receptor and propose to designate this receptor LPA 6 .Lysophosphatidic acid (LPA 3 ; 1-or 2-acyl-sn-glycero-3-phosphate) is a naturally occurring lipid mediator with diverse biological activities (1, 2). LPA plays important roles in many biological processes, such as the nervous system (3), tumor metastasis (4), wound healing (5), cardiovascular functions (6), and reproduction (7), through its specific G protein-coupled receptors (GPCRs). At least five subtypes of LPA receptors have been identified. Three receptors (LPA 1 (8), LPA 2 (9), and LPA 3 (10, 11)) share about 50% amino acid sequence identities, and form the Edg (endothelial differentiation gene) family together with the GPCRs for sphingosine 1-phosphate. Two additional LPA receptors, p2y9/LPA 4 (12) and GPR92/LPA 5 (13, 14), which show small similarities with the Edg family GPCRs, were recently identified. These LPA receptors, by coupling with different sets of G proteins, transduce various responses in many cell types. Depending on the functional coupling of a given LPA receptor to the G proteins, LPA activates diverse signaling cascades involving phosphoinositide 3-kinase, phospholipase C, mitogen-activated protein kinase, Rho-family GTPase, and adenylyl cyclase (2).The fact that p2y5 shares the highest sequence homology with p2y9/LPA 4 among all GPCRs (12) strongly suggested that LPA is a ligand for p2y5. However, we could not detect LPAinduced Ca 2ϩ mobilization or cAMP level changes in p2y5-overexpressing cells at the time of the identification of p2y9/ LPA 4 as the fourth LPA receptor in our laboratory (12). In the course of the further analysis of p2y5-overexpressing cells, we found that p2y5 actually responded to LPA with activation of the G 13 -Rho signaling pathway. Our results confirm the identification of p2y5 as an LPA receptor and extend the knowledge of the functional roles of LPA. EXPERIMENTAL PROCEDURESLipids-1-Oleoyl-LPA, 1-palmitoyl-LPA, 1-stearoyl-LPA, 1-myristoyl-LPA, and 1-arachidonoyl-LPA were purchased from Avanti Polar Lipids (Alabaster, AL). 1-Linoleoyl-LPA was from Echelon Biosciences (Salt Lake City, UT). These lipids were stored at Ϫ30°C (10 mM stock in 50% ethanol). Alkyl-OMPT (10 mM stock in d...

Polyunsaturated fatty acids (PUFAs) in phospholipids affect the physical properties of membranes, but it is unclear which biological processes are influenced by their regulation. For example, the functions of membrane arachidonate that are independent of a precursor role for eicosanoid synthesis remain largely unknown. Here, we show that the lack of lysophosphatidylcholine acyltransferase 3 (LPCAT3) leads to drastic reductions in membrane arachidonate levels, and that LPCAT3-deficient mice are neonatally lethal due to an extensive triacylglycerol (TG) accumulation and dysfunction in enterocytes. We found that high levels of PUFAs in membranes enable TGs to locally cluster in high density, and that this clustering promotes efficient TG transfer. We propose a model of local arachidonate enrichment by LPCAT3 to generate a distinct pool of TG in membranes, which is required for normal directionality of TG transfer and lipoprotein assembly in the liver and enterocytes.DOI: http://dx.doi.org/10.7554/eLife.06328.001

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Lysophosphatidic acid (LPA) is a potent lipid mediator that evokes a variety of biological responses in many cell types via its specific G protein-coupled receptors. In particular, LPA affects cell morphology, cell survival, and cell cycle progression in neuronal cells. Recently, we identified p2y 9 /GPR23 as a novel fourth

Tumors often are associated with a low extracellular pH, which induces a variety of cellular events. However, the mechanisms by which tumor cells recognize and react to the acidic environment have not been fully elucidated. T-cell death-associated gene 8 (TDAG8) is an extracellular pH-sensing G protein-coupled receptor that is overexpressed in various tumors and tumor cell lines. In this report, we show that TDAG8 on the surface of tumor cells facilitates tumor development by sensing the acidic environment. Overexpression of TDAG8 in mouse Lewis lung carcinoma (LLC) cells enhanced tumor development in animal models and rendered LLC cells resistant to acidic culture conditions by increasing activation of protein kinase A and extracellular signal-regulated kinase in vitro. Moreover, shRNA-mediated knockdown of endogenous TDAG8 in NCI-H460 human non-small cell lung cancer cells reduced cell survival in an acidic environment in vitro as well as tumor development in vivo. Microarray analyses of tumorcontaining lung tissues of mice injected with TDAG8-expressing LLC cells revealed up-regulation of genes related to cell growth and glycolysis. These results support the hypothesis that TDAG8 enhances tumor development by promoting adaptation to the acidic environment to enhance cell survival/proliferation. TDAG8 may represent a therapeutic target for arresting tumor growth.Lewis lung carcinoma cells | orphan receptor | tumor-induced acidity | tumorigenesis