A unique fold of phospholipase C-β mediates dimerization and interaction with Gαq

Singer, Alex U.; Waldo, Gary L.; Harden, T. Kendall; Sondek, John

doi:10.1038/nsb731

Cited by 92 publications

(96 citation statements)

References 28 publications

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“…Addition of this subunit proved unnecessary for study of the CG6986 receptor: Coexpression of G␣16 did not alter proctolin sensitivity in HEK cells (data not shown), but did increase responses to other peptides (E.C.J., unpublished work). We assume that the increase in calcium-dependent fluorescence after receptor activation in HEK cells is the result of Gq coupling and concomitant activation of the phospholipase C pathway (34). There is substantial evidence implicating phosphoinositol metabolism as a component of proctolin signaling (11,12,18,20), but other pathways have also been proposed (35)(36)(37).…”

Section: Discussionmentioning

confidence: 99%

Identification and characterization of a G protein-coupled receptor for the neuropeptide proctolin in Drosophila melanogaster

Johnson

Garczynski

Park

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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Proctolin is a bioactive neuropeptide that modulates interneuronal and neuromuscular synaptic transmission in a wide variety of arthropods. We present several lines of evidence to propose that the orphan G protein-coupled receptor CG6986 of Drosophila is a proctolin receptor. When expressed in mammalian cells, CG6986 confers second messenger activation after proctolin application, with an EC 50 of 0.3 nM. In competition-based studies, the CG6986 receptor binds proctolin with high affinity (IC50 ‫؍‬ 4 nM). By microarray analysis, CG6986 transcript is consistently detected in head mRNA of different genotypes, and under different environmental conditions. By blot analysis, anti-CG6986 antibodies detect a band in tissue homogenates similar to the predicted size of the protein. Proctolin receptor immunosignals are found in the hindgut, heart, and in distinct neuronal populations of the CNS; such patterns correlate with previous demonstrations of proctolin biological activity, and in several instances, with areas of proctolin peptide immunosignals. The identification of a bona fide proctolin receptor provides the basis for a mechanistic analysis of this critical synaptic modulator.T he pentapeptide proctolin (RYLPT) was the first neuropeptide to be isolated and sequenced from insects (1). It was purified based on its myotropic actions on the insect hindgut, and was subsequently found to have wide distribution throughout the arthropods (2-4). In the crab Cancer, proctolin has potent effects on defined neural circuits within the stomatogastric system (5) and at the neuromuscular junction (6). In insects, proctolin is a major peptide cotransmitter and it modulates contractions of both somatic and visceral muscles (7,8). Proctolin-like immunosignals are found within a small number of neurons in the larval CNS of Drosophila and these include efferents to bodywall muscles, heart, and viscera (9).The mechanisms of actions of proctolin have also been investigated. Proctolin elevates levels of inositol trisphosphate (IP 3 ) and cAMP, and increases calcium entry through voltagegated ion channels (10-13). Several studies have suggested the existence of multiple proctolin receptor subtypes based on binding studies and activities of proctolin (14, 15) or its structural analogues (16-19). However, more recent studies (15,20,21) of proctolin binding support a model featuring a single proctolin receptor. The identification of a specific proctolin receptor will help address issues regarding possibilities of multiple receptors, the precise nature of proctolin-dependent signaling pathways, and phylogenetic differences in proctolin signaling. The significance of such studies is substantial because proctolin exerts such a widespread role as a synaptic modulator in arthropod physiology.Analysis of the completed Drosophila genomic sequences (22, 23) identified Ͼ100 genes encoding G protein-coupled receptors (GPCRs). Further analysis indicated that 44 such receptors are likely to have peptide ligands and that the majority of these are derive...

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Section: Discussionmentioning

confidence: 99%

Identification and characterization of a G protein-coupled receptor for the neuropeptide proctolin in Drosophila melanogaster

Johnson

Garczynski

Park

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

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“…The structure of the isolated C-terminus of turkey PLC-β2 has been solved [51]. It is composed of three long helices forming a coiled-coil that dimerizes along its long axis in an anti-parallel orientation.…”

Section: C2 Domain and C-terminal Extensionmentioning

confidence: 99%

“…This peptide was therefore defined as the signal transfer region of Gβ. Extensive studies with other peptides identified a second signal transfer region (Gβ [42][43][44][45][46][47][48][49][50][51][52][53][54] ) in the seventh blade of Gβ, and revealed segments in the second, fifth and seventh blade that hold the Gβγ/PLC-β2 complex together during activation ( Figure 5) [67,68]. In addition, it was found that the geranylgeranylated moiety attached to the C-terminal segment of the Gγ, which is proximal to the membrane and anchors Gβγ to lipids, binds to the PLC-β2 [69,70].…”

Section: Plc-β2 Binding Site(s) In Gβγmentioning

confidence: 99%

“…More striking is the identification of soluble fragments of Gβγ, Gβ [86][87][88][89][90][91][92][93][94][95][96][97][98][99][100][101][102][103][104][105] and Gβ [42][43][44][45][46][47][48][49][50][51][52][53][54] , that are able to convey full activation of PLC-β2 [66,67,79]. Additionally, Gβγ has been shown to stimulate the phosphodiesterase step of the PI(4,5)P 2 hydrolysis reaction which is not dependent on membrane surfaces, and further biochemical studies have shown that in the presence of Gβγ subunits, product inhibition does not occur suggesting that activation is through an enhanced release of product [19].…”

Section: Constructing An Activation Model Of Plc-β2 By Gβγmentioning

confidence: 99%

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Stimulation of phospholipase Cβ by membrane interactions, interdomain movement, and G protein binding — How many ways can you activate an enzyme?

Drin

Scarlata

2007

Cellular Signalling

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Signaling proteins are usually composed of one or more conserved structural domains. These domains are usually regulatory in nature by binding to specific activators or effectors, or species that regulate cellular location, etc. Inositol-specific mammalian phospholipase C (PLC) enzymes are multidomain proteins whose activities are controlled by regulators, such as G proteins, as well as membrane interactions. One of these domains has been found to bind membranes, regulators, and activate the catalytic region. The recently solved structure of a major region of PLC-β2 together with the structure of PLC-δ1 and a wealth of biochemical studies poises the system towards an understanding of the mechanism through which their regulations occurs.

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“…The idea that PLC-␤ may function as a dimer also agrees with the concentration-dependent activity described for PLC-␤2 (13) and with the observation of fluorescence resonance energy transfer between differently fluorophore-labeled PLC-␤2 molecules. 2 The structure of the carboxyl-terminal tail of turkey PLC-␤2 was recently solved at the atomic level by Singer et al (14). The tail is a long, three-stranded coiled coil.…”

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confidence: 99%

Mutations in the Carboxyl-terminal Domain of Phospholipase C-β1 Delineate the Dimer Interface and a Potential GαqInteraction Site

Ilkaeva

Kinch

Paulssen³

et al. 2002

Journal of Biological Chemistry

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The carboxyl-terminal domain of phospholipase C-␤ is required for its stimulation by G␣ q and for its G␣ q -specific GTPase-activating protein (GAP) activity. We subjected this domain to a combination of deletion and alanine/glycine scanning mutagenesis to detect mutations that would inhibit either responsiveness to G q or G q GAP activity. Most mutations that altered either response or GAP activity diminished both in parallel. Many of these mutations map at the interface at which the carboxyl-terminal domain was recently shown to form a dimer (Singer, A. U., et al. (2001) Nat. Struct. Biol., 9, 32-36). Most others clustered in an area that is a plausible G␣ q binding site. In addition, one mutation that differentially inhibited GAP activity relative to responsiveness to G␣ q mapped in this region at a location modeled to be in close contact with the switch II region of G␣ q . This is the site at which RGS proteins are thought to exert their GAP activity. Last, a deletion mutation differentially inhibited the response of phospholipase C-␤1 to G␣ q without blocking GAP activity. Its location in the molecule suggests that moving the attachment point of the catalytic domain can disrupt its ability to be activated by G␣ q .

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A unique fold of phospholipase C-β mediates dimerization and interaction with Gαq

Cited by 92 publications

References 28 publications

Identification and characterization of a G protein-coupled receptor for the neuropeptide proctolin in Drosophila melanogaster

Identification and characterization of a G protein-coupled receptor for the neuropeptide proctolin in Drosophila melanogaster

Stimulation of phospholipase Cβ by membrane interactions, interdomain movement, and G protein binding — How many ways can you activate an enzyme?

Mutations in the Carboxyl-terminal Domain of Phospholipase C-β1 Delineate the Dimer Interface and a Potential GαqInteraction Site

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