Comment on "A G protein coupled receptor is a plasma membrane receptor for the plant hormone abscisic acid"

Liu et al. (Reports, 23 March 2007, p. 1712) reported that the Arabidopsis thaliana gene GCR2 encodes a seven-transmembrane, G protein-coupled receptor for abscisic acid. We argue that GCR2 is not likely to be a transmembrane protein nor a G protein-coupled receptor. Instead, GCR2 is most likely a plant homolog of bacterial lanthionine synthetases.     

Molecular identification of a taste receptor gene for trehalose in Drosophila

The molecular nature of sweet taste receptors has not been fully explored. Employing a differential screening strategy, we identified a taste receptor gene, Tre1, that controls the taste sensitivity to trehalose in Drosophila melanogaster. The Tre1 gene encodes a novel protein with similarity to G protein-coupled seven-transmembrane receptors. Disruption of the Tre1 gene lowered the taste sensitivity to trehalose, whereas sensitivities to other sugars were unaltered. Overexpression of the Tre1 gene restored the taste sensitivity to trehalose in the Tre1 deletion mutant. The Tre1 gene is expressed in taste sensory cells. These results provide direct evidence that Tre1 encodes a putative taste receptor for trehalose in Drosophila.     

A role for thrombin receptor signaling in endothelial cells during embryonic development

The coagulation protease thrombin triggers fibrin formation, platelet activation, and other cellular responses at sites of tissue injury. We report a role for PAR1, a protease-activated G protein-coupled receptor for thrombin, in embryonic development. Approximately half of Par1-/- mouse embryos died at midgestation with bleeding from multiple sites. PAR1 is expressed in endothelial cells, and a PAR1 transgene driven by an endothelial-specific promoter prevented death of Par1-/- embryos. Our results suggest that the coagulation cascade and PAR1 modulate endothelial cell function in developing blood vessels and that thrombin's actions on endothelial cells-rather than on platelets, mesenchymal cells, or fibrinogen-contribute to vascular development and hemostasis in the mouse embryo.     

Lutropin-choriogonadotropin receptor: an unusual member of the G protein-coupled receptor family

A complementary DNA (cDNA) for the rat luteal lutropin-choriogonadotropin receptor (LH-CG-R) was isolated with the use of a DNA probe generated in a polymerase chain reaction with oligonucleotide primers based on peptide sequences of purified receptor protein. As would be predicted from the cDNA sequence, the LH-CG-R consists of a 26-residue signal peptide, a 341-residue extracellular domain displaying an internal repeat structure characteristic of members of the leucine-rich glycoprotein (LRG) family, and a 333-residue region containing seven transmembrane segments. This membrane-spanning region displays sequence similarity with all members of the G protein-coupled receptor family. Hence, the LH-CG-R gene may have evolved by recombination of LRG and G protein-coupled receptor genes. Cells engineered to express LH-CG-R cDNA bind human choriogonadotropin with high affinity and show an increase in cyclic adenosine monophosphate when exposed to hormone. As revealed by RNA blot analysis and in situ hybridization, the 4.4-kilobase cognate messenger RNA is prominently localized in the rat ovary.     

Lysophosphatidylcholine as a ligand for the immunoregulatory receptor G2A

Although the biological actions of the cell membrane and serum lipid lysophosphatidylcholine (LPC) in atherosclerosis and systemic autoimmune disease are well recognized, LPC has not been linked to a specific cell-surface receptor. We show that LPC is a high-affinity ligand for G2A, a lymphocyte-expressed G protein-coupled receptor whose genetic ablation results in the development of autoimmunity. Activation of G2A by LPC increased intracellular calcium concentration, induced receptor internalization, activated ERK mitogen-activated protein kinase, and modified migratory responses of Jurkat T lymphocytes. This finding implicates a role for LPC-G2A interaction in the etiology of inflammatory autoimmune disease and atherosclerosis.     

Beta-arrestin 2 mediates endocytosis of type III TGF-beta receptor and down-regulation of its signaling

beta-Arrestins bind to activated seven transmembrane-spanning (7TMS) receptors (G protein-coupled receptors) after the receptors are phosphorylated by G protein-coupled receptor kinases (GRKs), thereby regulating their signaling and internalization. Here, we demonstrate an unexpected and analogous role of beta-arrestin 2 (betaarr2) for the single transmembrane-spanning type III transforming growth factor-beta (TGF-beta) receptor (TbetaRIII, also referred to as betaglycan). Binding of betaarr2 to TbetaRIII was also triggered by phosphorylation of the receptor on its cytoplasmic domain (likely at threonine 841). However, such phosphorylation was mediated by the type II TGF-beta receptor (TbetaRII), which is itself a kinase, rather than by a GRK. Association with betaarr2 led to internalization of both receptors and down-regulation of TGF-beta signaling. Thus, the regulatory actions of beta-arrestins are broader than previously appreciated, extending to the TGF-beta receptor family as well.     

High-resolution crystal structure of an engineered human beta2-adrenergic G protein-coupled receptor

Heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors constitute the largest family of eukaryotic signal transduction proteins that communicate across the membrane. We report the crystal structure of a human beta2-adrenergic receptor-T4 lysozyme fusion protein bound to the partial inverse agonist carazolol at 2.4 angstrom resolution. The structure provides a high-resolution view of a human G protein-coupled receptor bound to a diffusible ligand. Ligand-binding site accessibility is enabled by the second extracellular loop, which is held out of the binding cavity by a pair of closely spaced disulfide bridges and a short helical segment within the loop. Cholesterol, a necessary component for crystallization, mediates an intriguing parallel association of receptor molecules in the crystal lattice. Although the location of carazolol in the beta2-adrenergic receptor is very similar to that of retinal in rhodopsin, structural differences in the ligand-binding site and other regions highlight the challenges in using rhodopsin as a template model for this large receptor family.     

Crystal structure of a lipid G protein-coupled receptor

The lyso-phospholipid sphingosine 1-phosphate modulates lymphocyte trafficking, endothelial development and integrity, heart rate, and vascular tone and maturation by activating G protein-coupled sphingosine 1-phosphate receptors. Here, we present the crystal structure of the sphingosine 1-phosphate receptor 1 fused to T4-lysozyme (S1P(1)-T4L) in complex with an antagonist sphingolipid mimic. Extracellular access to the binding pocket is occluded by the amino terminus and extracellular loops of the receptor. Access is gained by ligands entering laterally between helices I and VII within the transmembrane region of the receptor. This structure, along with mutagenesis, agonist structure-activity relationship data, and modeling, provides a detailed view of the molecular recognition and requirement for hydrophobic volume that activates S1P(1), resulting in the modulation of immune and stromal cell responses.     

The sex peptide receptor in the Asian gypsy moth,Lymantria dispar,is involved in development and stress resistance

The G protein-coupled receptor (GPCR) regulates downstream genes by binding to a heterotrimeric G protein. However, the function of sex peptide receptor (SPR) in lepidopteran species is mostly unknown. Understanding the physiological functions of SPR in insects is essential for exploring new insecticidal targets. In the present study, the functions of an SPR in Lymantria dispar (Asian gypsy moth; LdSPR) were investigated. The expression of LdSPR was the highest in the 6th instar larval stage, and there was a large difference in expression between male and female adults. After LdSPR gene silencing, L. dispar larvae showed increased sensitivity to high temperature, starvation, and oxidative stress, indicating that LdSPR enhances stress resistance. These results enrich our knowledge of the function of the insect SPRs, which will lead to a better understanding of other insect GPCR family members and the identification of new targets for the development of environmentally friendly pesticides.     

Essential regulation of CNS angiogenesis by the orphan G protein-coupled receptor GPR124

The orphan G protein-coupled receptor (GPCR) GPR124/tumor endothelial marker 5 is highly expressed in central nervous system (CNS) endothelium. Here, we show that complete null or endothelial-specific GPR124 deletion resulted in embryonic lethality from CNS-specific angiogenesis arrest in forebrain and neural tube. Conversely, GPR124 overexpression throughout all adult vascular beds produced CNS-specific hyperproliferative vascular malformations. In vivo, GPR124 functioned cell-autonomously in endothelium to regulate sprouting, migration, and developmental expression of the blood-brain barrier marker Glut1, whereas in vitro, GPR124 mediated Cdc42-dependent directional migration to forebrain-derived, vascular endothelial growth factor-independent cues. Our results demonstrate CNS-specific angiogenesis regulation by an endothelial receptor and illuminate functions of the poorly understood adhesion GPCR subfamily. Further, the functional tropism of GPR124 marks this receptor as a therapeutic target for CNS-related vascular pathologies.     

Structure of an agonist-bound human A2A adenosine receptor

Activation of G protein-coupled receptors upon agonist binding is a critical step in the signaling cascade for this family of cell surface proteins. We report the crystal structure of the A(2A) adenosine receptor (A(2A)AR) bound to an agonist UK-432097 at 2.7 angstrom resolution. Relative to inactive, antagonist-bound A(2A)AR, the agonist-bound structure displays an outward tilt and rotation of the cytoplasmic half of helix VI, a movement of helix V, and an axial shift of helix III, resembling the changes associated with the active-state opsin structure. Additionally, a seesaw movement of helix VII and a shift of extracellular loop 3 are likely specific to A(2A)AR and its ligand. The results define the molecule UK-432097 as a "conformationally selective agonist" capable of receptor stabilization in a specific active-state configuration.     

Structure and functional expression of a human interleukin-8 receptor

Interleukin-8 (IL-8) is a member of a family of pro-inflammatory cytokines. Although the best characterized activities of IL-8 include the chemoattraction and activation of neutrophils, other members of this family have a wide range of specific actions including the chemotaxis and activation of monocytes, the selective chemotaxis of memory T cells, the inhibition of hematopoietic stem cell proliferation, and the induction of neutrophil infiltration in vivo. A complementary DNA encoding the IL-8 receptor from human neutrophils has now been isolated. The amino acid sequence shows that the receptor is a member of the superfamily of receptors that couple to guanine nucleotide binding proteins (G proteins). The sequence is 29% identical to that of receptors for the other neutrophil chemoattractants, fMet-Leu-Phe and C5a. Mammalian cells transfected with the IL-8 receptor cDNA clone bind IL-8 with high affinity and respond specifically to IL-8 by transiently mobilizing calcium. The IL-8 receptor may be part of a subfamily of related G protein-coupled receptors that transduce signals for the IL-8 family of pro-inflammatory cytokines.     

Molecular characterization of a functional cDNA encoding the rat substance P receptor

Substance P is a member of the tachykinin peptide family and participates in the regulation of diverse biological processes. The polymerase chain reaction and conventional library screening were used to isolate a complementary DNA (cDNA) encoding the rat substance P receptor from brain and submandibular gland. By homology analysis, this receptor belongs to the G protein-coupled receptor superfamily. The receptor cDNA was expressed in a mammalian cell line and the ligand binding properties of the encoded receptor were pharmacologically defined by Scatchard analysis and tachykinin peptide displacement as those of a substance P receptor. The distribution of the messenger RNA for this receptor is highest in urinary bladder, submandibular gland, striatum, and spinal cord, which is consistent with the known distribution of substance P receptor binding sites. Thus, this receptor appears to mediate the primary actions of substance P in various brain regions and peripheral tissues.     

Cloning of complementary DNA encoding a functional human interleukin-8 receptor

Interleukin-8 (IL-8) is an inflammatory cytokine that activates neutrophil chemotaxis, degranulation, and the respiratory burst. Neutrophils express receptors for IL-8 that are coupled to guanine nucleotide-binding proteins (G proteins); binding of IL-8 to its receptor induces the mobilization of intracellular calcium stores. A cDNA clone from HL-60 neutrophils, designated p2, has now been isolated that encodes a human IL-8 receptor. When p2 is expressed in oocytes from Xenopus laevis, the oocytes bind 125I-labeled IL-8 specifically and respond to IL-8 by mobilizing calcium stores with an EC50 of 20 nM. This IL-8 receptor has 77% amino acid identity with a second human neutrophil receptor isotype that binds IL-8 with higher affinity. It also exhibits 69% amino acid identity with a protein reported to be an N-formyl peptide receptor from rabbit neutrophils, but less than 30% identity with all other known G protein-coupled receptors, including the human N-formyl peptide receptor.     

Structures of the CXCR4 chemokine GPCR with small-molecule and cyclic peptide antagonists

Chemokine receptors are critical regulators of cell migration in the context of immune surveillance, inflammation, and development. The G protein-coupled chemokine receptor CXCR4 is specifically implicated in cancer metastasis and HIV-1 infection. Here we report five independent crystal structures of CXCR4 bound to an antagonist small molecule IT1t and a cyclic peptide CVX15 at 2.5 to 3.2 angstrom resolution. All structures reveal a consistent homodimer with an interface including helices V and VI that may be involved in regulating signaling. The location and shape of the ligand-binding sites differ from other G protein-coupled receptors and are closer to the extracellular surface. These structures provide new clues about the interactions between CXCR4 and its natural ligand CXCL12, and with the HIV-1 glycoprotein gp120.     

斑马鱼尾加压素Ⅱ受体基因组织表达的研究

尾加压素Ⅱ受体(UrotensinⅡreceptor,UT)是一种7次跨膜G蛋白偶联受体,是目前发现的具有最强收缩血管作用的尾加压素Ⅱ的特异性受体。斑马鱼中存在4种不同的UT-like基因(uts2r1,uts2r2,uts2r3和uts2r4),分别位于斑马鱼4条不同染色体上。这4种uts2r基因核苷酸序列各不相同,uts2r1位于3号染色体上,其与12号染色体上的uts2r2基因蛋白序列相似度最高为47%,与6号染色体上的uts2r3和16号染色体上的uts2r4蛋白相似度分别是41.6%和33%。通过设计特异引物检测每种uts2r基因在斑马鱼成鱼各组织中的表达情况,结果显示3号染色体上的uts2r1基因在肾脏中表达量最高,6号染色体上的uts2r2基因在脊髓中表达量最高,12号染色体上的uts2r3基因在心脏中表达量最高,而16号染色体上的uts2r4基因在背肌中表达量最高。     

Snapshot of activated G proteins at the membrane: the Galphaq-GRK2-Gbetagamma complex

G protein-coupled receptor kinase 2 (GRK2) plays a key role in the desensitization of G protein-coupled receptor signaling by phosphorylating activated heptahelical receptors and by sequestering heterotrimeric G proteins. We report the atomic structure of GRK2 in complex with Galphaq and Gbetagamma, in which the activated Galpha subunit of Gq is fully dissociated from Gbetagamma and dramatically reoriented from its position in the inactive Galphabetagamma heterotrimer. Galphaq forms an effector-like interaction with the GRK2 regulator of G protein signaling (RGS) homology domain that is distinct from and does not overlap with that used to bind RGS proteins such as RGS4.     

利用WGCNA鉴定谷子内源脱落酸响应禾生指梗霉胁迫的共表达基因

【目的】脱落酸（ABA）作为一类逆境激素,在植物生长发育、生物胁迫和非生物胁迫中发挥着重要作用。脱落酸受体蛋白PYR/PYL/PCAR及SNF1相关的蛋白激酶（SnRK2）是介导脱落酸信号转导的重要调控因子。本研究通过预测脱落酸及其信号转导途径中关键基因在谷子白发病致病菌禾生指梗霉（Sclerospora graminicola）中的调控作用,为谷子内源脱落酸响应禾生指梗霉侵染的互作研究提供参考。【方法】通过对禾生指梗霉侵染的晋谷21号谷子进行转录组测序和脱落酸含量测定,基于谷子全基因组对脱落酸信号转导通路上的PYL和SnRK2家族基因进行鉴定、分析,利用测定的转录组构建加权基因共表达网络（WGCNA）,并与禾生指梗霉侵染引起的寄主内源脱落酸含量进行关联,预测脱落酸及其下游信号转导基因PYL和SnRK2在谷子与禾生指梗霉互作调控中的关键核心基因;利用qRT-PCR技术对候选基因进行验证。【结果】谷子中存在禾本科中较为保守的PYL和SnRK2家族基因各11个,且在PYL和SnRK2家族基因的启动子上均预测到脱落酸响应元件。在禾生指梗霉侵染后,寄主内源脱落酸在第一、第二时期大量积累,含量显著高于对照组,分别为22.50和18.08 ng·mL^-1,而在第三、第四和第五时期脱落酸含量下降,低于对照组。在基因共表达网络分析中,利用18 535个基因共构建了34个基因共表达模块。通过对脱落酸含量和PYL、SnRK2家族基因的关联分析,预测到MEpaleturquoise和MEbrown模块为核心候选模块。利用GO功能富集和模块关键基因的挖掘共预测到1个PYL家族基因Seita.1G030500和2个SnRK2家族基因Seita.2G394500、Seita.3G03200,以及3个核心基因Seita.4G105600、Seita.6G218100和Seita.9G138400,共6个基因可能在脱落酸及其信号转导调控过程中参与谷子与禾生指梗霉的互作。对预测到的3个核心基因在水稻和拟南芥数据中进行比对,鉴定到Seita.4G105600为转导蛋白/WD40重复超家族蛋白、Seita.6G218100为WRKY57转录因子、Seita.9G138400为TIFY转录因子。qRT-PCR分析表明Seita.2G394500、Seita.4G105600和Seita.6G218100基因在谷子白发病早期表达均上调。【结论】谷子在受到禾生指梗霉侵染后脱落酸会在体内大量积累,预测到1个PYL家族基因、2个SnRK2家族基因、2个转录因子基因和1个WD40家族蛋白基因参与谷子内源脱落酸响应禾生指梗霉侵染过程。qRT-PCR结果表明1个SnRK2家族基因、1个WD40家族蛋白基因和1个WRKY57转录因子基因共3个基因可能在谷子脱落酸响应禾生指梗霉侵染过程中发挥重要作用。