Type IV Secretion-Dependent Activation of Host MAP Kinases Induces an Increased Proinflammatory Cytokine Response to Legionella pneumophila

The immune system must discriminate between pathogenic and nonpathogenic microbes in order to initiate an appropriate response. Toll-like receptors (TLRs) detect microbial components common to both pathogenic and nonpathogenic bacteria, whereas Nod-like receptors (NLRs) sense microbial components introduced into the host cytosol by the specialized secretion systems or pore-forming toxins of bacterial pathogens. The host signaling pathways that respond to bacterial secretion systems remain poorly understood. Infection with the pathogen Legionella pneumophila, which utilizes a type IV secretion system (T4SS), induced an increased proinflammatory cytokine response compared to avirulent bacteria in which the T4SS was inactivated. This enhanced response involved NF-κB activation by TLR signaling as well as Nod1 and Nod2 detection of type IV secretion. Furthermore, a TLR- and RIP2-independent pathway leading to p38 and SAPK/JNK MAPK activation was found to play an equally important role in the host response to virulent L. pneumophila. Activation of this MAPK pathway was T4SS-dependent and coordinated with TLR signaling to mount a robust proinflammatory cytokine response to virulent L. pneumophila. These findings define a previously uncharacterized host response to bacterial type IV secretion that activates MAPK signaling and demonstrate that coincident detection of multiple bacterial components enables immune discrimination between virulent and avirulent bacteria

Chemical Genetics Reveals Bacterial and Host Cell Functions Critical for Type IV Effector Translocation by Legionella pneumophila

Delivery of effector proteins is a process widely used by bacterial pathogens to subvert host cell functions and cause disease. Effector delivery is achieved by elaborate injection devices and can often be triggered by environmental stimuli. However, effector export by the L. pneumophila Icm/Dot Type IVB secretion system cannot be detected until the bacterium encounters a target host cell. We used chemical genetics, a perturbation strategy that utilizes small molecule inhibitors, to determine the mechanisms critical for L. pneumophila Icm/Dot activity. From a collection of more than 2,500 annotated molecules we identified specific inhibitors of effector translocation. We found that L. pneumophila effector translocation in macrophages requires host cell factors known to be involved in phagocytosis such as phosphoinositide 3-kinases, actin and tubulin. Moreover, we found that L. pneumophila phagocytosis and effector translocation also specifically require the receptor protein tyrosine phosphate phosphatases CD45 and CD148. We further show that phagocytosis is required to trigger effector delivery unless intimate contact between the bacteria and the host is artificially generated. In addition, real-time analysis of effector translocation suggests that effector export is rate-limited by phagocytosis. We propose a model in which L. pneumophila utilizes phagocytosis to initiate an intimate contact event required for the translocation of pre-synthesized effector molecules. We discuss the need for host cell participation in the initial step of the infection and its implications in the L. pneumophila lifestyle. Chemical genetic screening provides a novel approach to probe the host cell functions and factors involved in host–pathogen interactions

Legionella pneumophila Secretes a Mitochondrial Carrier Protein during Infection

The Mitochondrial Carrier Family (MCF) is a signature group of integral membrane proteins that transport metabolites across the mitochondrial inner membrane in eukaryotes. MCF proteins are characterized by six transmembrane segments that assemble to form a highly-selective channel for metabolite transport. We discovered a novel MCF member, termed Legionella nucleotide carrier Protein (LncP), encoded in the genome of Legionella pneumophila, the causative agent of Legionnaire's disease. LncP was secreted via the bacterial Dot/Icm type IV secretion system into macrophages and assembled in the mitochondrial inner membrane. In a yeast cellular system, LncP induced a dominant-negative phenotype that was rescued by deleting an endogenous ATP carrier. Substrate transport studies on purified LncP reconstituted in liposomes revealed that it catalyzes unidirectional transport and exchange of ATP transport across membranes, thereby supporting a role for LncP as an ATP transporter. A hidden Markov model revealed further MCF proteins in the intracellular pathogens, Legionella longbeachae and Neorickettsia sennetsu, thereby challenging the notion that MCF proteins exist exclusively in eukaryotic organisms

Structural studies of T4S systems by electron microscopy

Abstract: Type IV secretion (T4S) systems are large dynamic nanomachines that transport DNA and/or proteins through the membranes of bacteria. Analysis of T4S system architecture is an extremely challenging task taking into account their multi protein organisation and lack of overall global symmetry. Nonetheless the last decade demonstrated an amazing progress achieved by X-ray crystallography and cryo-electron microscopy. In this review we present a structural analysis of this dynamic complex based on recent advances in biochemical, biophysical and structural studies

Measurement and interpretation of same-sign W boson pair production in association with two jets in pp collisions at s = 13 TeV with the ATLAS detector

This paper presents the measurement of fducial and diferential cross sections for both the inclusive and electroweak production of a same-sign W-boson pair in association with two jets (W±W±jj) using 139 fb−1 of proton-proton collision data recorded at a centre-of-mass energy of √s = 13 TeV by the ATLAS detector at the Large Hadron Collider. The analysis is performed by selecting two same-charge leptons, electron or muon, and at least two jets with large invariant mass and a large rapidity diference. The measured fducial cross sections for electroweak and inclusive W±W±jj production are 2.92 ± 0.22 (stat.) ± 0.19 (syst.)fb and 3.38±0.22 (stat.)±0.19 (syst.)fb, respectively, in agreement with Standard Model predictions. The measurements are used to constrain anomalous quartic gauge couplings by extracting 95% confdence level intervals on dimension-8 operators. A search for doubly charged Higgs bosons H±± that are produced in vector-boson fusion processes and decay into a same-sign W boson pair is performed. The largest deviation from the Standard Model occurs for an H±± mass near 450 GeV, with a global signifcance of 2.5 standard deviations

Differential cross-section measurements of the production of four charged leptons in association with two jets using the ATLAS detector

Differential cross-sections are measured for the production of four charged leptons in association with two jets. These measurements are sensitive to final states in which the jets are produced via the strong interaction as well as to the purely-electroweak vector boson scattering process. The analysis is performed using proton-proton collision data collected by ATLAS at √s = 13 TeV and with an integrated luminosity of 140 fb−1. The data are corrected for the effects of detector inefficiency and resolution and are compared to state-of-the-art Monte Carlo event generator predictions. The differential cross-sections are used to search for anomalous weak-boson self-interactions that are induced by dimension-six and dimension-eight operators in Standard Model effective field theory

Search for dark photons in rare Z boson decays with the ATLAS detector

A search for events with a dark photon produced in association with a dark Higgs boson via rare decays of the standard model Z boson is presented, using 139     fb − 1 of √ s = 13     TeV proton-proton collision data recorded by the ATLAS detector at the Large Hadron Collider. The dark boson decays into a pair of dark photons, and at least two of the three dark photons must each decay into a pair of electrons or muons, resulting in at least two same-flavor opposite-charge lepton pairs in the final state. The data are found to be consistent with the background prediction, and upper limits are set on the dark photon’s coupling to the dark Higgs boson times the kinetic mixing between the standard model photon and the dark photon, α D ϵ 2 , in the dark photon mass range of [5, 40] GeV except for the Υ mass window [8.8, 11.1] GeV. This search explores new parameter space not previously excluded by other experiments

Combined measurement of the Higgs boson mass from the H → γγ and H → ZZ∗ → 4ℓ decay channels with the ATLAS detector using √s = 7, 8, and 13 TeV pp collision data

A measurement of the mass of the Higgs boson combining the H → Z Z ∗ → 4 ℓ and H → γ γ decay channels is presented. The result is based on 140     fb − 1 of proton-proton collision data collected by the ATLAS detector during LHC run 2 at a center-of-mass energy of 13 TeV combined with the run 1 ATLAS mass measurement, performed at center-of-mass energies of 7 and 8 TeV, yielding a Higgs boson mass of 125.11 ± 0.09 ( stat ) ± 0.06 ( syst ) = 125.11 ± 0.11     GeV . This corresponds to a 0.09% precision achieved on this fundamental parameter of the Standard Model of particle physics

Modulation of Rab GTPase function by a protein phosphocholine transferase

The intracellular pathogen Legionella pneumophila modulates the activity of host GTPases to direct the transport and assembly of the membrane-bound compartment in which it resides1–6. In vitro studies have suggested that the Legionella protein DrrA post-translationally modifies the GTPase Rab1 by a process called AMPylation7. Here, mass spectrometry was used to investigate post-translational modifications to Rab1 that occur during infection of host cells by Legionella. Consistent with in vitro studies, DrrA-mediated AMPylation of a conserved tyrosine residue in the switch II region of Rab1 was detected during infection. In addition, a modification to an adjacent serine residue in Rab1 was discovered, which was independent of DrrA. The Legionella effector protein AnkX was required for this modification. Biochemical studies determined that AnkX directly mediates the covalent attachment of a phosphocholine moiety to Rab1. This phosphocholine transferase activity used CDP-choline as a substrate and required a conserved histidine residue located in the FIC domain of the AnkX protein. During infection, AnkX modified both Rab1 and Rab35, which explains how this protein modulates membrane transport through both the endocytic and exocytic pathways of the host cell. Thus, phosphocholination of Ra