In vivo structure of the Legionella type II secretion system by electron cryotomography

AbstractThe type II secretion system (T2SS) is a multi-protein envelope-spanning assembly that translocates a wide range of virulence factors, enzymes and effectors through the outer membrane (OM) of many Gram-negative bacteria. Here, using electron cryotomography and subtomogram averaging methods, we present the first in situ structure of an intact T2SS, imaged within the human pathogen Legionella pneumophila. Although the T2SS has only limited sequence and component homology with the evolutionarily-related Type IV pilus (T4P) system, we show that their overall architectures are remarkably similar. Despite similarities, there are also differences, including for instance that the T2SS-ATPase complex is usually present but disengaged from the inner membrane, the T2SS has a much longer periplasmic vestibule, and it has a short-lived flexible pseudopilus. Placing atomic models of the components into our ECT map produced a complete architectural model of the intact T2SS that provides new insights into the structure and function of its components, its position within the cell envelope, and the interactions between its different subcomplexes. Overall, these structural results strongly support the piston model for substrate extrusion.

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In vivo structure of the Legionella type II secretion system by electron cryotomography

Nature Microbiology ◽

10.1038/s41564-019-0603-6 ◽

2019 ◽

Vol 4 (12) ◽

pp. 2101-2108 ◽

Cited By ~ 6

Author(s):

Debnath Ghosal ◽

Ki Woo Kim ◽

Huaixin Zheng ◽

Mohammed Kaplan ◽

Hilary K. Truchan ◽

...

Keyword(s):

Secretion System ◽

Type Ii Secretion ◽

Type Ii ◽

Type Ii Secretion System ◽

Electron Cryotomography

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Publisher Correction: In vivo structure of the Legionella type II secretion system by electron cryotomography

Nature Microbiology ◽

10.1038/s41564-020-0693-1 ◽

2020 ◽

Vol 5 (4) ◽

pp. 651-651

Author(s):

Debnath Ghosal ◽

Ki Woo Kim ◽

Huaixin Zheng ◽

Mohammed Kaplan ◽

Hilary K. Truchan ◽

...

Keyword(s):

Secretion System ◽

Type Ii Secretion ◽

Type Ii ◽

Type Ii Secretion System ◽

Electron Cryotomography

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In vivo cross-linking of EpsG to EpsL suggests a role for EpsL as an ATPase-pseudopilin coupling protein in the Type II secretion system of Vibrio cholerae

Molecular Microbiology ◽

10.1111/j.1365-2958.2010.07487.x ◽

2010 ◽

Vol 79 (3) ◽

pp. 786-798 ◽

Cited By ~ 41

Author(s):

Miranda D. Gray ◽

Michael Bagdasarian ◽

Wim G. J. Hol ◽

Maria Sandkvist

Keyword(s):

Vibrio Cholerae ◽

Secretion System ◽

Cross Linking ◽

Type Ii Secretion ◽

Type Ii ◽

Type Ii Secretion System ◽

Coupling Protein

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Molecular architecture of theLegionellaDot/Icm type IV secretion system

10.1101/312009 ◽

2018 ◽

Cited By ~ 3

Author(s):

Debnath Ghosal ◽

Yi-Wei Chang ◽

Kwang Cheol Jeong ◽

Joseph P. Vogel ◽

Grant J. Jensen

Keyword(s):

Legionella Pneumophila ◽

Cell Envelope ◽

Secretion System ◽

Host Cells ◽

Molecular Architecture ◽

Intact Cells ◽

Type Iv ◽

Type Ivb Secretion ◽

Electron Cryotomography ◽

First Time

AbstractLegionella pneumophilasurvives and replicates inside host cells by secreting ~300 effectors through the Dot/Icm type IVB secretion system (T4BSS). Understanding this machine’s structure is challenging because of its large number of components (27) and integration into all layers of the cell envelope. Previously we overcame this obstacle by imaging the Dot/Icm T4BSS in its native state within intact cells through electron cryotomography. Here we extend our observations by imaging a stabilized mutant that yielded a higher resolution map. We describe for the first time the presence of a well-ordered central channel that opens up into a windowed large (~32 nm wide) secretion chamber with an unusual 13-fold symmetry. We then dissect the complex by matching proteins to densities for many components, including all those with periplasmic domains. The placement of known and predicted structures of individual proteins into the map reveals the architecture of the T4BSS and provides a roadmap for further investigation of this amazing specialized secretion system.

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GspD, The Type II Secretion System Secretin of Leptospira, Protects Hamsters against Lethal Infection with a Virulent L. interrogans Isolate

Vaccines ◽

10.3390/vaccines8040759 ◽

2020 ◽

Vol 8 (4) ◽

pp. 759

Author(s):

Samantha Paulina Llanos Salinas ◽

Luz Olivia Castillo Sánchez ◽

Giselle Castañeda Miranda ◽

Ernesto Armando Rodríguez Reyes ◽

Liliana Ordoñez López ◽

...

Keyword(s):

Virulent Strain ◽

Lethal Dose ◽

Secretion System ◽

Type Ii Secretion ◽

Leptospira Interrogans ◽

Type Ii ◽

Pathogenic Leptospira ◽

Exact Test ◽

Type Ii Secretion System

The wide variety of pathogenic Leptospira serovars and the weak protection offered by the available vaccines encourage the search for protective immunogens against leptospirosis. We found that the secretin GspD of the type II secretion system (T2S) of Leptospira interrogans serovar Canicola was highly conserved amongst pathogenic serovars and was expressed in vivo during infection, as shown by immunohistochemistry. Convalescent sera of hamsters, dogs, and cows showed the presence of IgG antibodies, recognizing a recombinant version of this protein expressed in Escherichia coli (rGspDLC) in Western blot assays. In a pilot vaccination study, a group of eight hamsters was immunized on days zero and 14 with 50 µg of rGspDLC mixed with Freund’s incomplete adjuvant (FIA). On day 28 of the study, 1,000 LD50 (Lethal Dose 50%) of a virulent strain of Leptospira interrogans serovar Canicola (LOCaS46) were inoculated by an intraoral submucosal route (IOSM). Seventy-five percent protection against disease (p = 0.017573, Fisher’s exact test) and 50% protection against infection were observed in this group of vaccinated hamsters. In contrast, 85% of non-vaccinated hamsters died six to nine days after the challenge. These results suggest the potential usefulness of the T2S secretin GspD of Leptospira as a protective recombinant vaccine against leptospirosis.

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Type IV Fimbrial Biogenesis Is Required for Protease Secretion and Natural Transformation in Dichelobacter nodosus

Journal of Bacteriology ◽

10.1128/jb.00138-07 ◽

2007 ◽

Vol 189 (14) ◽

pp. 5022-5033 ◽

Cited By ~ 43

Author(s):

Xiaoyan Han ◽

Ruth M. Kennan ◽

Dane Parker ◽

John K. Davies ◽

Julian I. Rood

Keyword(s):

Natural Transformation ◽

Secretion System ◽

Extracellular Protease ◽

Classical Type ◽

Type Ii Secretion ◽

Type Ii ◽

Dichelobacter Nodosus ◽

Type Iv ◽

Type Ii Secretion System ◽

Protease Secretion

ABSTRACT The objective of this study was to develop an understanding of the molecular mechanisms by which type IV fimbrial biogenesis, natural transformation, and protease secretion are linked in the ovine foot rot pathogen, Dichelobacter nodosus. We have shown that like the D. nodosus fimbrial subunit FimA, the pilin-like protein PilE and the FimN, FimO, and FimP proteins, which are homologs of PilB, PilC, and PilD from Pseudomonas aeruginosa, are essential for fimbrial biogenesis and natural transformation, indicating that transformation requires an intact type IV fimbrial apparatus. The results also showed that extracellular protease secretion in the fimN, fimO, fimP, and pilE mutants was significantly reduced, which represents the first time that PilB, PilC, and PilE homologs have been shown to be required for the secretion of unrelated extracellular proteins in a type IV fimbriate bacterium. Quantitative real-time PCR analysis of the three extracellular protease genes aprV2, aprV5, and bprV showed that the effects on protease secretion were not mediated at the transcriptional level. Bioinformatic analysis did not identify a classical type II secretion system, and the putative fimbrial biogenesis gene pilQ was the only outer membrane secretin gene identified. Based on these results, it is postulated that in D. nodosus, protease secretion occurs by a type II secretion-related process that directly involves components of the type IV fimbrial biogenesis machinery, which represents the only type II secretion system encoded by the small genome of this highly evolved pathogen.

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Structure, Dynamics and Cellular Insight Into Novel Substrates of the Legionella pneumophila Type II Secretion System

Frontiers in Molecular Biosciences ◽

10.3389/fmolb.2020.00112 ◽

2020 ◽

Vol 7 ◽

Author(s):

Theo J. Portlock ◽

Jessica Y. Tyson ◽

Sarath C. Dantu ◽

Saima Rehman ◽

Richard C. White ◽

...

Keyword(s):

Legionella Pneumophila ◽

Secretion System ◽

Type Ii Secretion ◽

Type Ii ◽

Structure Dynamics ◽

Type Ii Secretion System ◽

Insight Into

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Insights into minor pseudopilin complexes of the Type II secretion system

Acta Crystallographica Section A Foundations and Advances ◽

10.1107/s2053273314094145 ◽

2014 ◽

Vol 70 (a1) ◽

pp. C585-C585

Author(s):

Yichen Zhang ◽

Frederick Faucher ◽

Zongchao Jia

Keyword(s):

Virulence Factors ◽

Secretion System ◽

Stable Complex ◽

Type Ii Secretion ◽

Type Ii ◽

Binding Model ◽

Binary And Ternary Complexes ◽

Type Ii Secretion System ◽

Wide Range ◽

Binding Mechanisms

The type II secretion system (T2SS) is sophisticated multiprotein machinery that enables Gram-negative pathogens to secrete a wide range of exoproteins, named virulence factors, into the extracellular environments. In Pseudomonas aeruginosa, the Xcp T2SS is responsible for secreting many virulence factors that induce severe infections. In T2SS, the recognition and binding of secreted exoproteins are conducted by a structure called the pseudopilius tip, which is formed by four minor pseudopilins, including XcpU, XcpV, XcpW and XcpX. These minor pseudopilins form a quaternary complex, which is also involved in the initiation and regulation of the pseudopilus assembly. Although individual structures of these four pseudopilins have been revealed in different organisms, the substrate recognition and binding mechanisms have not been clearly elucidated due to the lack of systematic studies on the whole structures of several complexes formed by these pseudopilins. As a result, the understanding of the structures of these protein complexes will provide useful information for unveiling the mystery of the recognition and binding mechanisms. The establishment of the substrate binding model requires the preparation of stable complex(es) of substrates and certain minor pseudopilin(s). In this work, we aim to gradually elucidate the secretion mechanisms by assembling each component to build up the whole architecture. The structure of XcpV in complex with XcpW has been determined, and other complexes, especially the XcpU-containing binary and ternary complexes, have been stably established and purified. The identification of these complex structures will significantly promote our understandings of the type II secretion mechanisms.

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