Structure of ginseng major latex-like protein 151 and its proposed lysophosphatidic acid-binding mechanism

Lysophosphatidic acid (LPA) is a phospholipid growth factor with myriad effects on biological systems. LPA is usually present bound to animal plasma proteins such as albumin or gelsolin. When LPA complexes with plasma proteins, it binds to its cognate receptors with higher affinity than when it is free. Recently, gintonin from ginseng was found to bind to LPA and to activate mammalian LPA receptors. Gintonin contains two components: ginseng major latex-like protein 151 (GLP) and ginseng ribonuclease-like storage protein. Here, the crystal structure of GLP is reported, which belongs to the plant Bet v 1 superfamily, and a model is proposed for how GLP binds LPA. Amino-acid residues of GLP recognizing LPA were identified using site-directed mutagenesis and isothermal titration calorimetry. The resulting GLP mutants were used to study the activation of LPA receptor-dependent signalling pathways. In contrast to wild-type GLP, the H147A mutant did not bind LPA, elicit intracellular Ca2+transients in neuronal cells or activate Ca2+-dependent Cl−channels inXenopusoocytes. Based on these results, a mechanism by which GLP recognizes LPA and its requirement to activate G protein-coupled LPA receptors to elicit diverse biological responses were proposed.

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Unlabeled lysophosphatidic acid receptor binding in free solution as determined by a compensated interferometric reader

The Journal of Lipid Research ◽

10.1194/jlr.d120000880 ◽

2020 ◽

Vol 61 (8) ◽

pp. 1244-1251 ◽

Cited By ~ 2

Author(s):

Manisha Ray ◽

Kazufumi Nagai ◽

Yasuyuki Kihara ◽

Amanda Kussrow ◽

Michael N. Kammer ◽

...

Keyword(s):

Receptor Binding ◽

Lysophosphatidic Acid ◽

Specific Binding ◽

Test Case ◽

Free Solution ◽

Label Free ◽

Adhesive Properties ◽

Lpa Receptor ◽

System A ◽

G Protein Coupled

Native interactions between lysophospholipids (LPs) and their cognate LP receptors are difficult to measure because of lipophilicity and/or the adhesive properties of lipids, which contribute to high levels of nonspecific binding in cell membrane preparations. Here, we report development of a free-solution assay (FSA) where label-free LPs bind to their cognate G protein-coupled receptors (GPCRs), combined with a recently reported compensated interferometric reader (CIR) to quantify native binding interactions between receptors and ligands. As a test case, the binding parameters between lysophosphatidic acid (LPA) receptor 1 (LPA1; one of six cognate LPA GPCRs) and LPA were determined. FSA-CIR detected specific binding through the simultaneous real-time comparison of bound versus unbound species by measuring the change in the solution dipole moment produced by binding-induced conformational and/or hydration changes. FSA-CIR identified KD values for chemically distinct LPA species binding to human LPA1 and required only a few nanograms of protein: 1-oleoyl (18:1; KD = 2.08 ± 1.32 nM), 1-linoleoyl (18:2; KD = 2.83 ± 1.64 nM), 1-arachidonoyl (20:4; KD = 2.59 ± 0.481 nM), and 1-palmitoyl (16:0; KD = 1.69 ± 0.1 nM) LPA. These KD values compared favorably to those obtained using the previous generation back-scattering interferometry system, a chip-based technique with low-throughput and temperature sensitivity. In conclusion, FSA-CIR offers a new increased-throughput approach to assess quantitatively label-free lipid ligand-receptor binding, including nonactivating antagonist binding, under near-native conditions.

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Lysophosphatidic Acid Upregulates Recepteur D’origine Nantais Expression and Cell Invasion via Egr-1, AP-1, and NF-κB Signaling in Bladder Carcinoma Cells

International Journal of Molecular Sciences ◽

10.3390/ijms21010304 ◽

2020 ◽

Vol 21 (1) ◽

pp. 304

Author(s):

Pham Ngoc Khoi ◽

Shinan Li ◽

Ung Trong Thuan ◽

Dhiraj Kumar Sah ◽

Taek Won Kang ◽

...

Keyword(s):

Bladder Cancer ◽

Lysophosphatidic Acid ◽

Cell Invasion ◽

Bladder Carcinoma ◽

Carcinoma Cells ◽

Lpa Receptor ◽

Carcinoma Metastasis ◽

Muscle Invasive ◽

G Protein Coupled ◽

Bladder Carcinoma Cells

Muscle invasive bladder carcinoma is a highly malignant cancer with a high mortality rate, due to its tendency to metastasize. The tyrosine kinase recepteur d’origine nantais (RON) promotes bladder carcinoma metastasis. Lysophosphatidic acid (LPA) is a phospholipid derivative, which acts as a signaling molecule to activate three high affinity G-protein coupled receptors, LPA1, LPA2, and LPA3. This in turn leads to cell proliferation and contributes to oncogenesis. However, little is known about the effects of LPA on invasive bladder cancer (IBC). In this study, we discovered that LPA upregulated RON expression, which in turn promoted cell invasion in bladder cancer T24 cells. As expected, we found that the LPA receptor was essential for the LPA induced increase in RON expression. More interestingly, we discovered that LPA induced RON expression via the MAPK (ERK1/2, JNK1/2), Egr-1, AP-1, and NF-κB signaling axes. These results provide experimental evidence and novel insights regarding bladder malignancy metastasis, which could be helpful for developing new therapeutic strategies for IBC treatment.

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Lipid kinase and protein kinase activities of G-protein-coupled phosphoinositide 3-kinase γ: structure–activity analysis and interactions with wortmannin

Biochemical Journal ◽

10.1042/bj3240489 ◽

1997 ◽

Vol 324 (2) ◽

pp. 489-495 ◽

Cited By ~ 80

Author(s):

Stefka STOYANOVA ◽

Ginette BULGARELLI-LEVA ◽

Cornelia KIRSCH ◽

Theodor HANCK ◽

Reinhard KLINGER ◽

...

Keyword(s):

Protein Kinase ◽

G Protein ◽

Catalytic Domain ◽

Transmembrane Helix ◽

Site Directed Mutagenesis ◽

Amino Acid Residues ◽

Lipid Kinase ◽

Cell Responses ◽

Phosphoinositide 3 Kinase ◽

G Protein Coupled

Signalling via seven transmembrane helix receptors can lead to a massive increase in cellular PtdIns(3,4,5)P3, which is critical for the induction of various cell responses and is likely to be produced by a trimeric G-protein-sensitive phosphoinositide 3-kinase (PI3Kγ). We show here that PI3Kγ is a bifunctional lipid kinase and protein kinase, and that both activities are inhibited by wortmannin at concentrations equal to those affecting the p85/p110α heterodimeric PI3K (IC50 approx. 2 nM). The binding of wortmannin to PI3Kγ, as detected by anti-wortmannin antisera, closely followed the inhibition of the kinase activities. Truncation of more than the 98 N-terminal amino acid residues from PI3Kγ produced proteins that were inactive in wortmannin binding and kinase assays. This suggests that regions apart from the core catalytic domain are important in catalysis and inhibitor interaction. The covalent reaction of wortmannin with PI3Kγ was prevented by preincubation with phosphoinositides, ATP and its analogues adenine and 5′-(4-fluorosulphonylbenzoyl)adenine. Proteolytic analysis of wortmannin-prelabelled PI3Kγ revealed candidate wortmannin-binding peptides around Lys-799. Replacement of Lys-799 by Arg through site-directed mutagenesis aborted the covalent reaction with wortmannin and the lipid kinase and protein kinase activities completely. The above illustrates that Lys-799 is crucial to the phosphate transfer reaction and wortmannin reactivity. Parallel inhibition of the PI3Kγ-associated protein kinase and lipid kinase by wortmannin and by the Lys-799 → Arg mutation reveals that both activities are inherent in the PI3Kγ polypeptide.

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Transcriptional regulation of lysophosphatidic acid receptors 2 and 3 regulates myeloid commitment of hematopoietic stem cells

AJP Cell Physiology ◽

10.1152/ajpcell.00506.2020 ◽

2021 ◽

Author(s):

Kuan-Hung Lin ◽

Jui-Chung Chiang ◽

Wei-Min Chen ◽

Ya-Hsuan Ho ◽

Chao-Ling Yao ◽

...

Keyword(s):

Lysophosphatidic Acid ◽

Stem Cell Differentiation ◽

Receptor Subtypes ◽

Physical Interaction ◽

Hematopoietic Stem ◽

Lpa Receptor ◽

Gata Factors ◽

Lpa Receptors ◽

Myeloid Progenitors

Lysophosphatidic acid (LPA) is one of the lipids identified to be involved in stem cell differentiation. It exerts various functions through activation of G protein-coupled LPA receptors (LPARs). In previous studies, we have demonstrated that activation of LPA receptor 3 (LPA3) promotes erythropoiesis in human HSCs and zebrafish using molecular andpharmacological approaches. Our results show that treatment of LPA2 agonist suppressed erythropoiesis, whereas activation of LPA3 by 2S-OMPT promoted it, both in vitro and in vivo. Furthermore, we have demonstrated the inhibitory role of LPA3 during megakaryopoiesis. However, the mechanism underlying these observation remains elusive. In the present study, we suggest that the expression pattern of LPARs may be correlated with the transcriptional factors GATA-1 and GATA-2 at different stages of myeloid progenitors. We determined that manipulation of GATA factors affected the expression levels of LPA2 and LPA3. Using luciferase assays, we demonstrate that the promoter regions of LPAR2 and LPAR3 were regulated by these GATA factors. Mutation of GATA binding sites in these regions abrogated luciferase activity, suggesting that LPA2 and LPA3 are regulated by GATA factors. Moreover, physical interaction between GATA factors and the promoter region of LPA receptors was verified using chromatin immunoprecipitation (ChIP) studies. Taken together, our results suggest that balance between LPA2 and LPA3, which may be determined by GATA factors, is a regulatory switch for lineage commitment in myeloid progenitors. The expression-level balance of LPA receptor subtypes represents a novel mechanism regulating erythropoiesis and megakaryopoiesis.

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Lysophosphatidic acid receptor 1 modulates lipopolysaccharide-induced inflammation in alveolar epithelial cells and murine lungs

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00058.2011 ◽

2011 ◽

Vol 301 (4) ◽

pp. L547-L556 ◽

Cited By ~ 42

Author(s):

Jing Zhao ◽

Donghong He ◽

Yanlin Su ◽

Evgeny Berdyshev ◽

Jerold Chun ◽

...

Keyword(s):

Epithelial Cells ◽

Lysophosphatidic Acid ◽

Lung Inflammation ◽

Lung Epithelial Cells ◽

Prostaglandin E ◽

Dependent Manner ◽

Wild Type ◽

Lpa Receptor ◽

Knock Down ◽

Lpa Receptors

Lysophosphatidic acid (LPA), a bioactive phospholipid, plays an important role in lung inflammation by inducing the release of chemokines and lipid mediators. Our previous studies have shown that LPA induces the secretion of interleukin-8 and prostaglandin E2 in lung epithelial cells. Here, we demonstrate that LPA receptors contribute to lipopolysaccharide (LPS)-induced inflammation. Pretreatment with LPA receptor antagonist Ki16425 or downregulation of LPA receptor 1 (LPA1) by small-interfering RNA (siRNA) attenuated LPS-induced phosphorylation of p38 MAPK, I-κB kinase, and I-κB in MLE12 epithelial cells. In addition, the blocking of LPA1 also suppressed LPS-induced IL-6 production. Furthermore, LPS treatment promoted interaction between LPA1 and CD14, a LPS coreceptor, in a time- and dose-dependent manner. Disruption of lipid rafts attenuated the interaction between LPA1 and CD14. Mice challenged with LPS increased plasma LPA levels and enhanced expression of LPA receptors in lung tissues. To further investigate the role of LPA receptors in LPS-induced inflammation, wild-type, or LPA1-deficient mice, or wild-type mice pretreated with Ki16425 were intratracheally challenged with LPS for 24 h. Knock down or inhibition of LPA1 decreased LPS-induced IL-6 release in bronchoalveolar lavage (BAL) fluids and infiltration of cells into alveolar space compared with wild-type mice. However, no significant differences in total protein concentration in BAL fluids were observed. These results showed that knock down or inhibition of LPA1 offered significant protection against LPS-induced lung inflammation but not against pulmonary leak as observed in the murine model for lung injury.

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A l-Lysine Transporter of High Stereoselectivity of the Amino Acid-Polyamine-Organocation (APC) Superfamily

Journal of Biological Chemistry ◽

10.1074/jbc.m113.510743 ◽

2013 ◽

Vol 289 (3) ◽

pp. 1377-1387 ◽

Cited By ~ 8

Author(s):

Jagdeep Kaur ◽

Elena Olkhova ◽

Viveka Nand Malviya ◽

Ernst Grell ◽

Hartmut Michel

Keyword(s):

Amino Acids ◽

Amino Acid ◽

Ligand Binding ◽

Isothermal Titration Calorimetry ◽

Functional Characterization ◽

Homology Model ◽

Docking Studies ◽

Site Directed Mutagenesis ◽

Titration Calorimetry ◽

Amino Acid Residues

Membrane proteins of the amino acid-polyamine-organocation (APC) superfamily transport amino acids and amines across membranes and play an important role in the regulation of cellular processes. We report the heterologous production of the LysP-related transporter STM2200 from Salmonella typhimurium in Escherichia coli, its purification, and functional characterization. STM2200 is assumed to be a proton-dependent APC transporter of l-lysine. The functional interaction between basic amino acids and STM2200 was investigated by thermoanalytical methods, i.e. differential scanning and isothermal titration calorimetry. Binding of l-lysine to STM2200 in its solubilized monomer form is entropy-driven. It is characterized by a dissociation constant of 40 μm at pH 5.9 and is highly selective; no evidence was found for the binding of l-arginine, l-ornithine, l-2,4-diaminobutyric acid, and l-alanine. d-Lysine is bound 45 times more weakly than its l-chiral form. We thus postulate that STM2200 functions as a specific transport protein. Based on the crystal structure of ApcT (Shaffer, P. L., Goehring, A., Shankaranarayanan, A., and Gouaux, E. (2009) Science 325, 1010–1014), a proton-dependent amino acid transporter of the APC superfamily, a homology model of STM2200 was created. Docking studies allowed identification of possible ligand binding sites. The resulting predictions indicated that Glu-222 and Arg-395 of STM2200 are markedly involved in ligand binding, whereas Lys-163 is suggested to be of structural and functional relevance. Selected variants of STM2200 where these three amino acid residues were substituted using single site-directed mutagenesis showed no evidence for l-lysine binding by isothermal titration calorimetry, which confirmed the predictions. Molecular aspects of the observed ligand specificity are discussed.

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Lysophosphatidic acid: mitogen and motility factor

Biochemical Society Transactions ◽

10.1042/bst0311209 ◽

2003 ◽

Vol 31 (6) ◽

pp. 1209-1212 ◽

Cited By ~ 54

Author(s):

F.N. van Leeuwen ◽

B.N.G. Giepmans ◽

L.A. van Meeteren ◽

W.H. Moolenaar

Keyword(s):

G Proteins ◽

Lysophosphatidic Acid ◽

Recent Progress ◽

Small Gtpases ◽

G Protein Coupled Receptors ◽

Signal Transduction Pathways ◽

Potent Inducer ◽

Motility Factor ◽

Lpa Receptors ◽

G Protein Coupled

LPA (lysophosphatidic acid), the simplest of al glycerophospholipids, is a potent inducer of cell proliferation, migration and survival. It does so by activating its cognate G-protein-coupled receptors, four of which have been identified. LPA receptors couple to at least three distinct G-proteins and thereby activate multiple signal transduction pathways, particularly those initiated by the small GTPases Ras, Rho and Rac. Our recent work has shown that LPA signals Rac activation via the Tiam1 GDP/GTP exchange factor and thereby stimulates cell migration. Here we discuss recent progress in our understanding of LPA action.

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Epidermal growth factor increases lysophosphatidic acid production in human ovarian cancer cells: roles for phospholipase D2 and receptor transactivation

AJP Cell Physiology ◽

10.1152/ajpcell.00001.2009 ◽

2010 ◽

Vol 298 (1) ◽

pp. C163-C170 ◽

Cited By ~ 26

Author(s):

Ashley J. Snider ◽

Zhihong Zhang ◽

Yuhuan Xie ◽

Kathryn E. Meier

Keyword(s):

Ovarian Cancer ◽

Growth Factor ◽

Epidermal Growth Factor ◽

Tyrosine Kinase ◽

Lysophosphatidic Acid ◽

Cross Talk ◽

Human Ovarian Cancer ◽

Lpa Receptor ◽

Lpa Receptors ◽

Epidermal Growth

Lysophosphatidic acid (LPA), is a lipid mediator that binds to G-protein coupled receptors. Epidermal growth factor (EGF), a polypeptide growth factor, binds to the EGF receptor (EGFR), a receptor tyrosine kinase. Both LPA and EGF induce responses in tumor cells that include proliferation, migration, metastasis, and induction of angiogenesis. LPA has the potential to act as an autocrine/paracrine factor and can transactivate the EGFR. This study explores the role of phospholipase D2 (PLD2) activation in LPA production, as well as cross-talk between EGF and LPA receptors. We demonstrate that EGF and LPA both stimulate production of LPA by OVCAR3 and SKOV3 human ovarian cancer cell lines. PD158780, an EGFR-selective tyrosine kinase inhibitor, blocks LPA production in response to both EGF and LPA in OVCAR3 and SKOV3 cells. Pertussis toxin, an inhibitor of LPA receptor signaling, inhibits LPA production in response to both EGF and LPA. Similar results were observed for the LPA receptor antagonist, Ki16425. Overexpression of PLD2 increases LPA production, while knockdown of PLD2 blocks EGF-induced LPA production. A phospholipase A2 (PLA2) inhibitor also blocks LPA- and EGF-induced LPA production. These results indicate that EGF stimulates LPA production in a manner that requires PLD2, and suggest that cross-talk can occur bidirectionally between EGF and LPA receptors.

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Lysophosphatidic Acid Receptor Antagonists and Cancer: The Current Trends, Clinical Implications, and Trials

Cells ◽

10.3390/cells10071629 ◽

2021 ◽

Vol 10 (7) ◽

pp. 1629

Author(s):

Yu-Hsuan Lin ◽

Yueh-Chien Lin ◽

Chien-Chin Chen

Keyword(s):

Cancer Progression ◽

Lysophosphatidic Acid ◽

Lipid Mediator ◽

Membrane Phospholipids ◽

Receptor Antagonists ◽

Preclinical Trial ◽

Lpa Receptor ◽

Cellular Effects ◽

Lpa Receptors ◽

And Function

Lysophosphatidic acid (LPA) is a bioactive lipid mediator primarily derived from membrane phospholipids. LPA initiates cellular effects upon binding to a family of G protein-coupled receptors, termed LPA receptors (LPAR1 to LPAR6). LPA signaling drives cell migration and proliferation, cytokine production, thrombosis, fibrosis, angiogenesis, and lymphangiogenesis. Since the expression and function of LPA receptors are critical for cellular effects, selective antagonists may represent a potential treatment for a broad range of illnesses, such as cardiovascular diseases, idiopathic pulmonary fibrosis, voiding dysfunctions, and various types of cancers. More new LPA receptor antagonists have shown their therapeutic potentials, although most are still in the preclinical trial stage. This review provided integrative information and summarized preclinical findings and recent clinical trials of different LPA receptor antagonists in cancer progression and resistance. Targeting LPA receptors can have potential applications in clinical patients with various diseases, including cancer.

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Autotaxin-Lysophosphatidic Acid: From Inflammation to Cancer Development

Mediators of Inflammation ◽

10.1155/2017/9173090 ◽

2017 ◽

Vol 2017 ◽

pp. 1-15 ◽

Cited By ~ 27

Author(s):

Silvia Anahi Valdés-Rives ◽

Aliesha González-Arenas

Keyword(s):

Wound Healing ◽

G Protein ◽

Cancer Progression ◽

Lysophosphatidic Acid ◽

Therapeutic Target ◽

G Protein Coupled Receptors ◽

Cancer Development ◽

Cellular Processes ◽

Lpa Receptors ◽

G Protein Coupled

Lysophosphatidic acid (LPA) is a ubiquitous lysophospholipid and one of the main membrane-derived lipid signaling molecules. LPA acts as an autocrine/paracrine messenger through at least six G protein-coupled receptors (GPCRs), known as LPA1–6, to induce various cellular processes including wound healing, differentiation, proliferation, migration, and survival. LPA receptors and autotaxin (ATX), a secreted phosphodiesterase that produces this phospholipid, are overexpressed in many cancers and impact several features of the disease, including cancer-related inflammation, development, and progression. Many ongoing studies aim to understand ATX-LPA axis signaling in cancer and its potential as a therapeutic target. In this review, we discuss the evidence linking LPA signaling to cancer-related inflammation and its impact on cancer progression.

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