<i>In situ</i>
            and high‐resolution cryo‐
            <scp>EM</scp>
            structure of a bacterial type
            <scp>VI</scp>
            secretion system membrane complex

Rapisarda, C.; Cherrak, Yassine; Kooger, Romain; Schmidt, Victoria; Pellarin, Riccardo; Logger, Laureen; Cascales, Éric; Pilhofer, Martin; Durand, Éric; Fronzes, Rémi

doi:10.15252/embj.2018100886

Cited by 80 publications

(62 citation statements)

References 86 publications

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“…First, conformational changes in the baseplate are proposed to trigger sheath contraction (22,69,70). Second, the membrane complex is expected to undergo large conformational changes to enable the passage of the spiked tube (19,71). It is possible that a leucine-to-arginine mutation causes aberrant interactions between VgrG and components of the membrane complex or baseplate, which prevent the necessary conformational changes for T6SS firing.…”

Section: Downloaded Frommentioning

confidence: 99%

The Tip of the VgrG Spike Is Essential to Functional Type VI Secretion System Assembly in Acinetobacter baumannii

Lopez

Feldman

2020

mBio

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The type VI secretion system (T6SS) is a critical weapon in bacterial warfare between Gram-negative bacteria. Although invaluable for niche establishment, this machine represents an energetic burden to its host bacterium. Acinetobacter baumannii is an opportunistic pathogen that poses a serious threat to public health due to its high rates of multidrug resistance. In some A. baumannii strains, the T6SS is transcriptionally downregulated by large multidrug resistance plasmids. Other strains, such as the clinical isolate AbCAN2, express T6SS-related genes but lack T6SS activity under laboratory conditions, despite not harboring these plasmids. This suggests that alternative mechanisms exist to repress the T6SS. Here, we used a transposon mutagenesis approach in AbCAN2 to identify novel T6SS repressors. Our screen revealed that the T6SS of this strain is inhibited by a homolog of VgrG, an essential structural component of all T6SSs reported to date. We named this protein inhibitory VgrG (VgrGi). Biochemical and in silico analyses demonstrated that the unprecedented inhibitory capability of VgrGi is due to a single amino acid mutation in a widely conserved C-terminal domain of unknown function, DUF2345. We also show that unlike in other bacteria, the C terminus of VgrG is essential for functional T6SS assembly in A. baumannii. Our study provides insight into the architectural requirements underlying functional assembly of the T6SS of A. baumannii. We propose that T6SS-inactivating point mutations are beneficial to the host bacterium, since they eliminate the energy cost associated with maintaining a functional T6SS, which appears to be unnecessary for A. baumannii virulence. IMPORTANCE Despite the clinical relevance of A. baumannii, little is known about its fundamental biology. Here, we show that a single amino acid mutation in VgrG, a critical T6SS structural protein, abrogates T6SS function. Given that this mutation was found in a clinical isolate, we propose that the T6SS of A. baumannii is probably not involved in virulence; this idea is supported by multiple genomic analyses showing that the majority of clinical A. baumannii strains lack proteins essential to the T6SS. We also show that, unlike in other species, the C terminus of VgrG is a unique architectural requirement for functional T6SS assembly in A. baumannii, suggesting that over evolutionary time, bacteria have developed changes to their T6SS architecture, leading to specialized systems.

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Section: Downloaded Frommentioning

confidence: 99%

The Tip of the VgrG Spike Is Essential to Functional Type VI Secretion System Assembly in Acinetobacter baumannii

Lopez

Feldman

2020

mBio

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“…T6SS biogenesis needs at least 14 different proteins. These subunits assemble two sub-complexes: a membrane complex anchored in the envelope (Durand et al, 2015;Rapisarda et al, 2019), on which a platform that controls the assembly of a contractile tail tube/sheath structure is docked (Basler et al, 2012;Brunet et al, 2015;Logger et al, 2016;Taylor et al, 2016;Nguyen et al, 2017;Nazarov et al, 2018). The tail tube/sheath complex comprises an inner tube made of Hcp hexamers covered by a sheath and topped by a needle spike constituted of a VgrG trimer and a PAAR-domain protein (Ho et al, 2014;Zoued et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

Structural basis for loading and inhibition of a bacterial T6 SS phospholipase effector by the VgrG spike

et al. 2020

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The bacterial type VI secretion system (T6SS) is a macromolecular machine that injects effectors into prokaryotic and eukaryotic cells. The mode of action of the T6SS is similar to contractile phages: the contraction of a sheath structure pushes a tube topped by a spike into target cells. Effectors are loaded onto the spike or confined into the tube. In enteroaggregative Escherichia coli, the Tle1 phospholipase binds the C-terminal extension of the VgrG trimeric spike. Here, we purify the VgrG-Tle1 complex and show that a VgrG trimer binds three Tle1 monomers and inhibits their activity. Using covalent cross-linking coupled to high-resolution mass spectrometry, we provide information on the sites of contact and further identify the requirement for a Tle1 N-terminal secretion sequence in complex formation. Finally, we report the 2.6-Å-resolution cryo-electron microscopy tri-dimensional structure of the (VgrG) 3 -(Tle1) 3 complex revealing how the effector binds its cargo, and how VgrG inhibits Tle1 phospholipase activity. The inhibition of Tle1 phospholipase activity once bound to VgrG suggests that Tle1 dissociation from VgrG is required upon delivery.

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“…The vast majority of T6SS gene clusters of Proteobacterial species encode three membrane proteins: TssJ, TssL, and TssM (8-10, 53) ( The electron microscopy structure of the fully assembled 1.7-MDa TssJLM MC from enteroaggregative Escherichia coli has been reported (49,68,69). The complex has a rocketlike structure: a large base, that contains the cytoplasmic and membrane domains of TssL and TssM, is followed by arches and pillars which correspond to the TssM periplasmic domains and TssJ (68). The TssJLM complex, which has five-fold symmetry in vivo and after purification, comprises copies of TssJ, and 10 copies of TssL and of TssM (49,58).…”

Section: Architecture Of the Type VI Secretion System The Membrane Comentioning

confidence: 99%

Structure and Activity of the Type VI Secretion System

Cherrak

Flaugnatti

Durand

et al. 2019

Protein Secretion in Bacteria

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The Type VI secretion system (T6SS) is a multiprotein machine that uses a spring-like mechanism to inject effectors into target cells. The injection apparatus is composed of a baseplate on which is built a contractile tail tube/sheath complex. The inner tube, topped by the spike complex, is propelled outside of the cell by the contraction of the sheath. The injection system is anchored to the cell envelope and oriented towards the cell exterior by a trans-envelope complex. Effectors delivered by the T6SS are loaded within the inner tube or on the spike complex, and can target prokaryotic and/or eukaryotic cells. Here, we summarize the structure, assembly and mechanism of action of the T6SS. We also review the function of effectors and their mode of recruitment and delivery.

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In situ and high‐resolution cryo‐ EM structure of a bacterial type VI secretion system membrane complex

Cited by 80 publications

References 86 publications

The Tip of the VgrG Spike Is Essential to Functional Type VI Secretion System Assembly in Acinetobacter baumannii

The Tip of the VgrG Spike Is Essential to Functional Type VI Secretion System Assembly in Acinetobacter baumannii

Structural basis for loading and inhibition of a bacterial T6 SS phospholipase effector by the VgrG spike

Structure and Activity of the Type VI Secretion System

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