The ins and outs of pertussis toxin

Locht, Camille; Coutte, Loïc; Mielcarek, Nathalie

doi:10.1111/j.1742-4658.2011.08237.x

Cited by 161 publications

(150 citation statements)

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“…The constitutive activities of L32A and D333A were also supported by independent PTX experiments. PTX inhibits GPCR-mediated activation of G i (47). The cAMP levels for D333A and L32A mutants were much lower than that for wild type even after PTX inhibition (Fig.…”

Section: G␣ I Signaling Properties Are Modulated By the Predicted Sinmentioning

confidence: 99%

Dynamic Coupling and Allosteric Networks in the α Subunit of Heterotrimeric G Proteins

Yao¹,

Malik

Griggs

et al. 2016

Journal of Biological Chemistry

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G protein ␣ subunits cycle between active and inactive conformations to regulate a multitude of intracellular signaling cascades. Important structural transitions occurring during this cycle have been characterized from extensive crystallographic studies. However, the link between observed conformations and the allosteric regulation of binding events at distal sites critical for signaling through G proteins remain unclear. Here we describe molecular dynamics simulations, bioinformatics analysis, and experimental mutagenesis that identifies residues involved in mediating the allosteric coupling of receptor, nucleotide, and helical domain interfaces of G␣ i . Most notably, we predict and characterize novel allosteric decoupling mutants, which display enhanced helical domain opening, increased rates of nucleotide exchange, and constitutive activity in the absence of receptor activation. Collectively, our results provide a framework for explaining how binding events and mutations can alter internal dynamic couplings critical for G protein function.Heterotrimeric G proteins are key mediators of intracellular signaling pathways that control diverse cellular processes ranging from movement and division to differentiation and neuronal activity (1). G proteins consist of three subunits: G␣, G␤, and G␥. Bound with GDP, G␣ forms an inactive complex with its G␤␥ subunit partners. Interaction with activated receptor (GPCR) 3 promotes the exchange of GDP for GTP on G␣ and its separation from G␤␥. Both isolated G␣ and G␤␥ can then bind and activate or inhibit downstream effectors. GTP hydrolysis deactivates G␣, which subsequently reassociates with G␤␥ completing the cycle. This cycle is further regulated by two classes of additional proteins called regulators of G protein signaling. These function as either GTPase-activating proteins (which promote GTP hydrolysis) or GDP dissociation inhibitors (GDIs, which hinder exchange of GDP for GTP) (2). Important conformational transitions occurring at each stage of this regulated cycle have been characterized from extensive crystallographic studies. These include GDP, GTP analogue, G␤␥, GTPase-activating protein, GDI and most recently GPCR bound complex structures of G␣. However, the link between the observed conformations and the atomic level mechanisms involved in coupling receptor association, G protein activation, and effector interaction remain unclear.All G␣ proteins consist of a catalytic GTP binding Ras-like domain (termed RasD) and a heterotrimeric G protein specific helical domain (HD). Recent principal component analysis (PCA) of 53 available G␣ crystallographic structures identified three major conformationally distinct groups ( Fig. 1 and Ref. 3). These groups correspond to structures with bound GTP analogues, GDP, and GDI (red, green, and blue points in Fig. 1a, respectively). The major variation in the accumulated structures is the concerted displacements of three nucleotide-binding site loops termed the switch regions (SI, SII, and SIII), as well as a relatively small...

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Section: G␣ I Signaling Properties Are Modulated By the Predicted Sinmentioning

confidence: 99%

Dynamic Coupling and Allosteric Networks in the α Subunit of Heterotrimeric G Proteins

Yao¹,

Malik

Griggs

et al. 2016

Journal of Biological Chemistry

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“…PTX is also an adjuvant that acts on dendritic cells (DCs) to induce DCs activation, IL-12 production, and the subsequent differentiation of T cells by modulating cellular signaling pathways [7][8][9][10][11][12]. Although many cell surface sialoglycoproteins may serve as PTX receptors, specific receptors for PTX have not been identified [13]. PTX induces intracellular signals in part through TLR4, and JNK, p38, ERK1/2, and NF-κB are all activated in response to PTX, similar to the response to LPS [6,14].…”

Section: Introductionmentioning

confidence: 99%

Differential IL‐10 production by DCs determines the distinct adjuvant effects of LPS and PTX in EAE induction

et al. 2014

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Pertussis toxin (PTX) is used as an adjuvant to induce EAE in mice as a model of MS. Although LPS and PTX are similar in their ability to mature dendritic cells (DCs) and in their adjuvant properties, here we demonstrate that LPS does not induce EAE development. We also demonstrate that DCs treated with PTX (PTX-DCs) are able to induce EAE, whereas DCs treated with LPS (LPS-DCs) fail to induce EAE. We determine that a key difference between LPS-DCs and PTX-DCs is that LPS-DCs produce larger amounts of IL-10. IL-10−/− -DCs treated with LPS promote stronger IFN-γ and IL-17 production and T-cell proliferation than WT DCs treated with LPS in a coculture system. Finally, we demonstrate that EAE can be successfully induced when IL-10 −/− -DCs treated with LPS are used as an adjuvant, whereas the use of PTX-DCs overexpressing IL-10 as an adjuvant markedly controls EAE development. These results indicate that the inability of LPS to induce EAE is based on the induction of high levels of IL-10 in the targeted DCs, providing insight into the mechanisms responsible for the induction of EAE.Keywords: Dendritic cells r EAE r IL-10 r LPS r Pertussis toxin (PTX)Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionMS is a chronic autoimmune disease of the CNS. MS is characterized by inflammation, demyelination, and axonal pathology and is thought to occur in genetically predisposed individuals following exposure to an environmental trigger that activates myelin-specific pathogenic T cells that can cross the blood-brain barrier. EAE is Correspondence: Prof. Bing Sun e-mail: bsun@sibs.ac.cn; Yong-Jun.Liu@BaylorHealth.edu a mouse model for studying MS. MS and EAE were previously thought to be mediated by Th1 cells; however, numerous recent studies have provided strong evidence that IL-17-producing T cells play a dominant role in the pathogenesis of EAE [1][2][3]. Pertussis toxin (PTX) is an exotoxin that is produced by Bordetella pertussis. PTX blocks chemokine receptors such as CXCR4 and CCR5. PTX treatment causes a decrease in CXCR4 * These authors contributed equally to this work.C 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu Eur. J. Immunol. 2014. 44: 1352-1362 Cellular immune response 1353 surface expression, inhibits G protein associated signaling, and blocks stromal cell derived factor 1 alpha mediated chemotaxis by activating the TCR signaling network [4]. However, the detailed mechanism by which PTX blocks CCR5 is unclear, although receptor crosstalk between an unknown PTX receptor and chemokine receptors has been suggested [4]. PTX has been widely used as an adjuvant to induce Th1/Th17-mediated organ-specific autoimmune diseases in animal models, including experimental autoimmune uveitis (EAU) and EAE. PTX has profound effects on the immune system and is thought to induce EAE by facilitating the "opening" of the blood-brain barrier [5,6]. PTX is also an adjuvant that acts on dendritic cells (DCs) to induce DCs activation, IL-12 ...

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“…T4SSs are perhaps the most versatile of the macromolecular transport systems, being essential for host colonization by many medically important microbes [8][9][10][11][12][13] , the horizontal transfer of genetic material between bacteria and from bacteria to plants 2 as well as the special cases of the Helicobacter pylori DNA uptake 14 and Neisseria gonorrhoeae DNA release 15 systems. The prototypical T4SS is that of the plant pathogen Agrobacterium tumefaciens that uses its Vir system to cause tumours in most dicotyledonous species 16 .…”

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confidence: 99%