G-protein-coupled receptors: Evolving views on physiological signalling: Symposium on G-Protein-Coupled Receptors: Evolving Concepts and New Techniques

Department of Pharmacology, Vanderbilt University Medical Center, 23rd Avenue South @ Pierce, Room 442 Robinson Research Building, Nashville, Tennessee 37232-6600, USA.
EMBO Reports (Impact Factor: 9.06). 10/2006; 7(9):866-9. DOI: 10.1038/sj.embor.7400788
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Available from: Michael Holinstat, Aug 04, 2014
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    • "3.2. G˛13 interaction with Itk is partially dependent on GDP-and GTP-bound conformation G␣ is normally bound to GDP in its inactive state, and with GTP in its active state (Holinstat et al., 2006). The GTP-bound G␣13 can interact with downstream effectors to regulate downstream signaling (Hart et al., 1998; Meigs et al., 2002; Yamaguchi et al., 2002; Niu et al., 2001; Gong et al., 2010). "
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    ABSTRACT: Tec family kinases play critical roles in the activation of immune cells. In particular, Itk is important for the activation of T cells via the T cell Receptor (TcR), however, molecules that cooperate with Itk to activate downstream targets remain little explored. Here we show that Itk interacts with the heterotrimeric G-protein α subunit Gα13 during TcR triggering. This interaction requires membrane localization of both partners, and is partially dependent on GDP- and GTP-bound states of Gα13. Furthermore, we find that Itk interacts with Gα13 via the zinc binding regions within its Tec homology domain. The interaction between Itk and Gα13 also results in tyrosine phosphorylation of Gα13, however this is not required for the interaction. Itk enhances Gα13 mediated activation of serum response factor (SRF) transcriptional activity dependent on its ability to interact with Gα13, but its kinase activity is not required to enhance SRF activity. These data reveal a new pathway regulated by Itk in cells, and suggest cross talk between Itk and G-protein signaling downstream of the TcR.
    The international journal of biochemistry & cell biology 02/2013; 45(6). DOI:10.1016/j.biocel.2013.02.011 · 4.05 Impact Factor
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    • "The GTP- and GDP-bound complexes define the active and inactive states of the G proteins, respectively. The binding of specific ligands to transmembrane receptors activates the heterotrimeric G protein subunits that are responsible for the flow of information in many eukaryotic signal transduction pathways [5]. The traditional G proteins coupled receptors (GPCRs) share a characteristic topological structure of seven transmembrane domains and recognize diverse extracellular signals. "
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    ABSTRACT: Background Adaptive responses in fungi result from the interaction of membrane receptors and extracellular ligands. Many different classes of receptors have been described in eukaryotic cells. Recently a new family of receptors classified as belonging to the progesterone-adiponectin receptor (PAQR) family has been identified. These receptors have the seven transmembrane domains characteristic of G-protein coupled receptors, but their activity has not been associated directly to G proteins. They share sequence similarity to the eubacterial hemolysin III proteins. Results A new receptor, SsPAQR1 (Sporothrixschenckiiprogesterone-adiponectinQ receptor1), was identified as interacting with Sporothrix schenckii G protein alpha subunit SSG-2 in a yeast two-hybrid assay. The receptor was identified as a member of the PAQR family. The cDNA sequence revealed a predicted ORF of 1542 bp encoding a 514 amino acids protein with a calculated molecular weight of 57.8 kDa. Protein domain analysis of SsPAQR1 showed the 7 transmembrane domains (TM) characteristic of G protein coupled receptors and the presence of the distinctive motifs that characterize PAQRs. A yeast-based assay specific for PAQRs identified progesterone as the agonist. S. schenckii yeast cells exposed to progesterone (0.50 mM) showed an increase in intracellular levels of 3′, 5′ cyclic adenosine monophosphate (cAMP) within the first min of incubation with the hormone. Different progesterone concentrations were tested for their effect on the growth of the fungus. Cultures incubated at 35°C did not grow at concentrations of progesterone of 0.05 mM or higher. Cultures incubated at 25°C grew at all concentrations tested (0.01 mM-0.50 mM) with growth decreasing gradually with the increase in progesterone concentration. Conclusion This work describes a receptor associated with a G protein alpha subunit in S. schenckii belonging to the PAQR family. Progesterone was identified as the ligand. Exposure to progesterone increased the levels of cAMP in fungal yeast cells within the first min of incubation suggesting the connection of this receptor to the cAMP signalling pathway. Progesterone inhibited the growth of both the yeast and mycelium forms of the fungus, with the yeast form being the most affected by the hormone.
    BMC Microbiology 09/2012; 12(1):194. DOI:10.1186/1471-2180-12-194 · 2.73 Impact Factor
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    • "Information is conveyed to the interior of the cell following the binding of ligands to receptors. The heterotrimeric G proteins constitute a family of GTPases that transmit messages received at cell surface receptors (GPCR) to cytoplasmic effector proteins inside the cell [5]. Heterotrimeric G proteins are made up of three subunits: the GTP-binding α subunit and the tightly associated complex of β and γ subunits. "
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    ABSTRACT: Important biological processes require selective and orderly protein-protein interactions at every level of the signalling cascades. G proteins are a family of heterotrimeric GTPases that effect eukaryotic signal transduction through the coupling of cell surface receptors to cytoplasmic effector proteins. They have been associated with growth and pathogenicity in many fungi through gene knock-out studies. In Sporothrix schenckii, a pathogenic, dimorphic fungus, we previously identified a pertussis sensitive G alpha subunit, SSG-1. In this work we inquire into its interactions with other proteins. Using the yeast two-hybrid technique, we identified protein-protein interactions between SSG-1 and other important cellular proteins. The interactions were corroborated using co-immuneprecipitation. Using these techniques we identified a Fe/Mn superoxide dismutase (SOD), a glyceraldehyde-3-P dehydrogenase (GAPDH) and two ion transport proteins, a siderophore-iron transporter belonging to the Major Facilitator Superfamily (MFS) and a divalent-cation transporter of the Nramp (natural resistance-associated macrophage protein) family as interacting with SSG-1. The cDNA's encoding these proteins were sequenced and bioinformatic macromolecular sequence analyses were used for the correct classification and functional assignment. This study constitutes the first report of the interaction of a fungal G alpha inhibitory subunit with SOD, GAPDH, and two metal ion transporters. The identification of such important proteins as partners of a G alpha subunit in this fungus suggests possible mechanisms through which this G protein can affect pathogenicity and survival under conditions of environmental stress or inside the human host. The two ion transporters identified in this work are the first to be reported in S. schenckii and the first time they are identified as interacting with fungal G protein alpha subunits. The association of G protein alpha subunits to transport molecules reinforces the role of G proteins in the response to environmental signals and also highlights the involvement of fungal G protein alpha subunits in nutrient sensing in S. schenckii. These interactions suggest that these permeases could function as transceptors for G proteins in fungi.
    BMC Microbiology 12/2010; 10(1):317. DOI:10.1186/1471-2180-10-317 · 2.73 Impact Factor
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