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Takehiko Yokomizo

Publications and source records attributed to Takehiko Yokomizo.

14 recordsLinked to original sources
PubMed2026

Lipid-mediated activation of BLT2 promotes membrane repair to prevent cell death.

Various pathogenic microorganisms produce toxins that create pores in cell membranes, causing cell damage and disrupting the host epithelial barrier. Recently, we reported that mice lacking the G protein-coupled receptor leukotriene B4 receptor 2 (BLT2), which is expressed in vascular endothelial and alveolar epithelial cells, are highly susceptible to pneumolysin (PLY), a pneumococci-generated toxin. Although we clarified the protective roles of BLT2 in vascular endothelial cells, those in alveolar epithelial cells have not been elucidated. Here, we report that lipid mediator 12-hydroxyheptadecatrienoic acid (12-HHT), which is produced by membrane-damaged epithelial cells, prevents cell death by promoting membrane repair through BLT2. BLT2 promoted the release of PLY-bound plasma membranes as extracellular vesicles in a sphingomyelinase-dependent manner. Additionally, BLT2 activated Rac1 and subsequent actin polymerization, leading to resistance to cell death. Furthermore, inhibition of 12-HHT production by aspirin and treatment with a BLT2 antagonist abolished the protective effect of BLT2. These findings provide a new therapeutic strategy for bacterial infection.

Receptors, Leukotriene B4↗
PubMed2003

Glucocorticoids up-regulate leukotriene B4 receptor-1 expression during neutrophilic differentiation of HL-60 cells.

Leukotriene B(4) (LTB(4)) is a potent activator of granulocytes and macrophages. The actions of LTB(4) are mediated by the specific G protein-coupled receptors, BLT1 and BLT2. We report up-regulation of BLT1 expression by dexamethasone (Dex), a synthetic glucocorticoid, in a promyelocytic cell line HL-60 during differentiation by retinoic acid (RA) into neutrophilic phenotype. The expression of BLT1 mRNA was also augmented by Dex in DMSO-differentiated neutrophilic HL-60 cells, but not in vitamin D(3)-differentiated monocytic HL-60 cells. Augmented expression of BLT1 by Dex was associated with enhanced functional activities, such as LTB(4)-induced intracellular calcium mobilization and chemotaxis. On the other hand, Dex failed to enhance BLT2 expression in RA-differentiated HL-60 cells, indicating different transcriptional regulations for these two receptors in spite of the fact that their genes are closely located (J. Exp. Med. 192 (2000) 413-420). These results suggest glucocorticoids enhance the functions of neutrophils during differentiation by up-regulating BLT1 expression, thus contributing to host defense.

Blotting, Western↗
PubMed2003

Helix 8 of the leukotriene B4 receptor is required for the conformational change to the low affinity state after G-protein activation.

Recent studies have revealed that G-protein-coupled receptors contain a putative cytoplasmic helical domain, helix 8. Leukotriene B4 (LTB4) receptor 1 derivatives with truncated or mutated helix 8 showed much higher LTB4 binding than wild-type (WT) receptors. Similar to the WT receptor, LTB4 promoted guanosine 5'-3-O-(thio)triphosphate (GTPgammaS) binding in these mutants. Unlike the WT receptor, however, the addition of GTPgammaS did not inhibit LTB4 binding to the mutant receptors. Scatchard analyses revealed that mutants maintained high affinity for LTB4, even in the presence of excess GTPgammaS. Consistently, mutant receptors showed a more prolonged Ca2+ mobilization and cellular metabolic activation than the WT receptor. From mutational studies and three-dimensional modeling based on the structure of bovine rhodopsin, we conclude that the helix 8 of LTB4 receptor 1 plays an important role in the conformational change of the receptor to the low affinity state after G-protein activation, possibly by sensing the status of coupling Galpha subunits as GTP-bound.

Amino Acid Sequence↗
PubMed2003

Cloning of adiponectin receptors that mediate antidiabetic metabolic effects.

Adiponectin (also known as 30-kDa adipocyte complement-related protein; Acrp30) is a hormone secreted by adipocytes that acts as an antidiabetic and anti-atherogenic adipokine. Levels of adiponectin in the blood are decreased under conditions of obesity, insulin resistance and type 2 diabetes. Administration of adiponectin causes glucose-lowering effects and ameliorates insulin resistance in mice. Conversely, adiponectin-deficient mice exhibit insulin resistance and diabetes. This insulin-sensitizing effect of adiponectin seems to be mediated by an increase in fatty-acid oxidation through activation of AMP kinase and PPAR-alpha. Here we report the cloning of complementary DNAs encoding adiponectin receptors 1 and 2 (AdipoR1 and AdipoR2) by expression cloning. AdipoR1 is abundantly expressed in skeletal muscle, whereas AdipoR2 is predominantly expressed in the liver. These two adiponectin receptors are predicted to contain seven transmembrane domains, but to be structurally and functionally distinct from G-protein-coupled receptors. Expression of AdipoR1/R2 or suppression of AdipoR1/R2 expression by small-interfering RNA supports our conclusion that they serve as receptors for globular and full-length adiponectin, and that they mediate increased AMP kinase and PPAR-alpha ligand activities, as well as fatty-acid oxidation and glucose uptake by adiponectin.

Adiponectin↗
PubMed2003

A combinatorial G protein-coupled receptor reconstitution system on budded baculovirus. Evidence for Galpha and Galphao coupling to a human leukotriene B4 receptor.

To investigate the coupling selectivity of G proteins and G protein-coupled receptors (GPCRs), we developed a reconstitution system made up of GPCR and heterotrimeric G proteins on extracellular baculovirus particles (budded virus (BV)). BV released from Sf9 cells infected with a recombinant baculovirus coding for human leukotriene B4 receptor (BLT1) cDNA exhibited a high level of BLT1 expression (27.3 pmol/mg of protein) and specific [3H]leukotriene B4 binding activity (Kd = 3.67 nm). The apparent low affinity of the expressed BLT1 is thought to be due to relative non-availability of the Galphai isoform, which couples to BLT1, in BV. Co-infection of heterotrimeric G protein recombinant viruses led to co-expression of BLT1 and G protein subunits on BV. A guanosine-5'-(beta,gamma-imido)triphosphate-sensitive, high affinity ligand binding was observed in the BLT1 BV co-expressing Galphai1beta1gamma2 (Kd = 0.17 nm). A relatively large amount of high affinity receptor protein was recovered in the co-expressing BV fraction (6.81 pmol/mg of protein). A combination of BLT1 and Galphai1 without Gbeta1gamma2 did not exhibit high affinity ligand binding on BV, indicating the low background environment for the GPCR-G protein coupling in this BV reconstitution system. To test other G proteins for coupling, various Galpha subunits were combinatorially expressed in BV with BLT1 and Gbeta1gamma2. The BLT1 BV co-expressing GalphaoAbeta1gamma2 exhibited a comparably high affinity ligand binding as well as ligand-stimulated guanosine 5'-3-O-(thio)triphosphate binding to Galphai1beta1gamma2. Co-expression of other Galpha isoforms such as Galphas, Galpha11, Galpha14, Galpha16, Galpha12, or Galpha13 did not exhibit any significant effects on ligand binding affinity in this system. These results reveal that BLT1 and coupled trimeric G proteins were functionally reconstituted on BV and that Galphao as well as Galphai couples to BLT1. This expression system should prove highly useful for pharmacological characterization, biosensor chip applications, and also drug discovery directed at highly important targets of the membrane receptor proteins.

Baculoviridae↗
PubMed2003

Plexin-A1 and plexin-B1 specifically interact at their cytoplasmic domains.

Semaphorin 3A (Sema3A) is a member of semaphorins and functions as an axonal repulsive guidance molecule. Neuropilin-1 and plexin-As form receptor complexes for Sema3A and plexin-As are thought to initiate the intracellular signaling cascade. However, the molecule by which plexin-As transduce their signal is not well understood. We searched molecules that interact with intracellular domains of plexin-A1 by yeast two-hybrid screening and identified a 349 amino acid fragment of plexin-B1 as a plexin-A1 interacting protein. We, then, cloned mouse plexin-B1 and confirmed their interaction in a mammalian expression system. Plexin-B1 physically associated with plexin-A1, but not with plexin-A2 or A3. Northern blot analysis showed the expression of both plexin-A1 and B1 in adult brain. We propose that plexin-A1 and B1 interact in the adult brain and transduce Sema3A signaling in cooperation.

Animals↗
PubMed2003

International Union of Pharmacology XXXVII. Nomenclature for leukotriene and lipoxin receptors.

The leukotrienes and lipoxins are biologically active metabolites derived from arachidonic acid. Their diverse and potent actions are associated with specific receptors. Recent molecular techniques have established the nucleotide and amino acid sequences and confirmed the evidence that suggested the existence of different G-protein-coupled receptors for these lipid mediators. The nomenclature for these receptors has now been established for the leukotrienes. BLT receptors are activated by leukotriene B(4) and related hydroxyacids and this class of receptors can be subdivided into BLT(1) and BLT(2). The cysteinyl-leukotrienes (LT) activate another group called CysLT receptors, which are referred to as CysLT(1) and CysLT(2). A provisional nomenclature for the lipoxin receptor has also been proposed. LXA(4) and LXB(4) activate the ALX receptor and LXB(4) may also activate another putative receptor. However this latter receptor has not been cloned. The aim of this review is to provide the molecular evidence as well as the properties and significance of the leukotriene and lipoxin receptors, which has lead to the present nomenclature.

Amino Acid Sequence↗
PubMed2002

Requirement of phosphatidylinositol 3-kinase activation and calcium influx for leukotriene B4-induced enzyme release.

Leukotriene B(4) (LTB(4)) is a potent lipid mediator involved in host defense and inflammatory responses. It causes chemotaxis, generation of reactive oxygen species, and degranulation. However, only little is known of the molecular mechanisms by which LTB(4) induces these biological activities. To analyze the intracellular signaling pathways to mediate lysosomal enzyme release through the cloned LTB(4) receptor (BLT1), we transfected BLT1 to rat basophilic leukemia cells (RBL-2H3). LTB(4) dose-dependently released beta-hexosaminidase, and the release was mostly inhibited when the cells were pretreated with pertussis toxin, indicating that the degranulation is mediated by G(i) proteins. LTB(4) activated phosphatidylinositol 3-kinase (PI3-K) through G(i), and inhibition of PI3-K by wortmannin or LY290042 inhibited degranulation. Granulocytes from PI3-Kgamma-deficient mice showed reduced LTB(4)-induced degranulation, suggesting that this isozyme of PI3-K is involved in the degranulation. LTB(4) also caused calcium release from intracellular stores and calcium influx from the outside milieu through G(i), but only the calcium influx is critical for the lysosomal enzyme release. Calcium influx and PI3-K activation are both downstream events of G(i), since they were inhibited by pertussis toxin. These two events are in essence independent each other, because calcium depletion did not affect PI3-K, and inhibition of PI3-K did not attenuate calcium influx significantly. Thus, our results have clearly shown that LTB(4) binds BLT1 and activates G(i)-like protein, and both PI3-Kgamma activation and a sustained calcium elevation by calcium influx are necessary for enzyme release in these cells.

Animals↗
PubMed2002

Characterization of mouse cysteinyl leukotriene receptors mCysLT1 and mCysLT2: differential pharmacological properties and tissue distribution.

Cysteinyl leukotrienes (LTs) are important proinflammatory mediators. Their precise roles in mice need to be elucidated to interpret mouse models of inflammatory diseases. For this purpose, we cloned and characterized mouse receptors for cysteinyl LTs, mCysLT(1) and mCysLT(2). mCysLT(1) and mCysLT(2) were composed of 339 amino acids with 87.3% identity and 309 amino acids with 73.4% identity to human orthologues, respectively. A pharmacological difference was noted between mouse and human CysLT(2). Pranlukast, a specific inhibitor for human CysLT(1), antagonized mCysLT(2) responses as determined by Ca(2+) elevation and receptor-induced promoter activation. The mRNA expressions of both mCysLTs were higher in C57BL/6 mice than in 129 mice. mCysLT(1) mRNA was expressed mainly in skin, lung, and small intestine. mCysLT(2) was seen more ubiquitously with high expressions in spleen, lung, and small intestine. By in situ hybridization we demonstrated for the first time that mCysLT(1) and mCysLT(2) were expressed in subcutaneous fibroblasts. The different pharmacological characteristics of CysLT(2) between human and mouse and the different distributions of CysLTs between mouse strains suggest that careful choice and interpretation are necessary for a study of CysLTs using animal models.

Amino Acid Sequence↗
PubMed2002

Leukotriene B4 receptors.

Leukotriene B4 (LTB4) is a potent chemotactic agent and activating factor for granulocytes. Two cell surface receptors for LTB4 (BLT1 and BLT2) have been isolated in the last few years. These receptors are G-protein-coupled receptors (GPCR), and they have 45% amino acid identity. BLT1 and BLT2 are high- and low- affinity receptors, respectively. Cells transfected with BLT1 and BLT2 show LTB4-dependent intracellular signal transduction and chemotaxis in vitro. The distribution and pharmacological characteristics of BLT1 and BLT2 are different, suggesting distinct roles for these receptors in vivo. The open reading frame (ORF) of BLT2 overlaps the promoter of BLT1, a so called 'promoter in ORF'. Based on recent publications on BLT1 transgenic and knock out mice phenotypes, it appears that LTB4 plays important roles in inflammation in addition to host defense in vivo.

Animals↗
PubMed2002

Leukotriene receptors: classification, gene expression, and signal transduction.

Leukotrienes (LTs) are potent pro-inflammatory mediators derived from arachidonic acid by the action of 5-lipoxygenase. There are two groups of LTs: LTB(4) and cysteinyl LTs (LTC(4), LTD(4), and LTE(4)). Both of them play important roles in many inflammatory diseases and allergic responses. Recently, their G-protein coupled receptors have been cloned. The identification of these receptors enables us to analyze their gene structures, regulation of expression, and signal transduction in the cells, and it also leads to the development of useful antagonists. Some LT receptors have been disrupted by gene targeting. Such studies may reveal novel functions of leukotrienes, confirming deeper viewpoints for further research.

Animals↗
PubMed2002

Association of eNOS Glu298Asp polymorphism with end-stage renal disease.

Nitric oxide (NO) derived from endothelial cells is profoundly related to the maintenance of physiological vascular tone. Impairment of endothelial NO generation brought about by gene polymorphism is considered the major deterioration factor for progressive renal disease, including diabetic nephropathy. The present study aimed to elucidate the Glu298Asp polymorphism of endothelial NO synthase (eNOS) in patients with end-stage renal disease (ESRD) and its role as a predisposing factor for cardiovascular complications. Glu298Asp in exon 7 of the eNOS gene was determined by polymerase chain reaction, followed by restriction fragment length polymorphism analysis, in ESRD patients (n=185) and compared with that of unrelated healthy individuals (n=304). The occurrence of 298Asp was significantly higher in the ESRD group (P=0.0020; odds ratio [OR] 1.65; 95% confidential interval [CI]: 1.21 to 2.25). In this group, 72 patients had type 2 diabetes mellitus (DM). Although 298Asp did not reach a significant level in the non-DM ESRD subgroup, the occurrence of 298Asp was significantly higher in DM-derived ESRD patients (P=0.0010; OR 2.02; 95% CI: 1.37 to 3.07). The functional effect of the Glu298Asp was examined using Chinese hamster ovary (CHO) cells stably overexpressing either 1917G or 1917T. NO-selective electrode measurements and fluorometric nitrite assay revealed a statistically significant difference in NO production or nitrite accumulation between CHO 1917G and 1917T (P<0.01). These data indicated that Glu298Asp is the predisposing factor in ESRD, especially DM-derived ESRD. The functional difference in NO generation depending on eNOS with either glutamate or aspartate at position 298 was also confirmed in vitro.

Animals↗