scholarly journals Structure of the Core of the Type Three Secretion System Export Apparatus

2018 ◽  
Author(s):  
Lucas Kuhlen ◽  
Patrizia Abrusci ◽  
Steven Johnson ◽  
Joseph Gault ◽  
Justin Deme ◽  
...  
Keyword(s):  
Integral Membrane Protein ◽  
Integral Membrane Proteins ◽  
Structural Elements ◽  
Secretion Systems ◽  
Core Component ◽  
The Core ◽  
Cryo Electron Microscopy ◽  
Type Three Secretion System ◽  
Structure Assembly ◽  
Type Three Secretion

SummaryExport of proteins through type three secretion systems is critical for bacterial motility and virulence of many major bacterial pathogens. Three putative integral membrane proteins (FliP/FliQ/FliR) are suggested to form the core of an export gate in the inner membrane, but their structure, assembly and location within the final nanomachine remain unclear. We here present the structure of this complex at 4.2 Å by cryo-electron microscopy. None of the subunits adopt canonical integral membrane protein topologies and common helix-turn-helix structural elements allow them to form a helical assembly with 5:4:1 stoichiometry. Fitting of the structure into reconstructions of intact secretion systems localize the export gate as a core component of the periplasmic portion of the machinery, and cross-linking experiments confirm this observation. This study thereby identifies the export gate as a key element of the secretion channel and implies that it primes the helical architecture of the components assembling downstream.One Sentence SummaryThe core of the T3SS export gate forms a supra-membrane helical assembly

scholarly journals The structure of an injectisome export gate demonstrates conservation of architecture in the core export gate between flagellar and virulence type three secretion systems

2019 ◽  
Author(s):  
Steven Johnson ◽  
Lucas Kuhlen ◽  
Justin C. Deme ◽  
Patrizia Abrusci ◽  
Susan M. Lea
Keyword(s):  
Electron Microscopy ◽  
Membrane Proteins ◽  
Complex Analysis ◽  
Secretion Systems ◽  
Core Complex ◽  
The Core ◽  
Cryo Electron Microscopy ◽  
Equivalent Complex ◽  
Type Three Secretion ◽  
Resolution Structure

AbstractExport of proteins through type three secretion systems (T3SS) is critical for motility and virulence of many major bacterial pathogens. Proteins are exported though a genetically defined export gate complex consisting of three proteins. We have recently shown at 4.2 Å that the flagellar complex of these three putative membrane proteins (FliPQR in flagellar systems, SctRST in virulence systems) assemble into an extra-membrane helical assembly that likely seeds correct assembly of the rod above. Here we present the structure of an equivalent complex from the more fragile Shigella virulence system at 3.5 Å by cryo-electron microscopy. This higher resolution structure reveals further detail and confirms the prediction of structural conservation in this core complex. Analysis of particle heterogeneity also reveals details of how the SctS/FliQ subunits sequentially assemble in the complex.

scholarly journals The Structure of an Injectisome Export Gate Demonstrates Conservation of Architecture in the Core Export Gate between Flagellar and Virulence Type III Secretion Systems

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Steven Johnson ◽  
Lucas Kuhlen ◽  
Justin C. Deme ◽  
Patrizia Abrusci ◽  
Susan M. Lea
Keyword(s):  
Type Iii Secretion ◽  
Complex Analysis ◽  
Secretion Systems ◽  
Core Complex ◽  
Type Iii ◽  
The Core ◽  
Cryo Electron Microscopy ◽  
Type Iii Secretion Systems ◽  
Equivalent Complex ◽  
Resolution Structure

ABSTRACT Export of proteins through type III secretion systems (T3SS) is critical for motility and virulence of many major bacterial pathogens. Proteins are exported through a genetically defined export gate complex consisting of three proteins. We have recently shown at 4.2 Å that the flagellar complex of these three putative membrane proteins (FliPQR in flagellar systems, SctRST in virulence systems) assembles into an extramembrane helical assembly that likely seeds correct assembly of the rod. Here we present the structure of an equivalent complex from the Shigella virulence system at 3.5 Å by cryo-electron microscopy. This higher-resolution structure yields a more precise description of the structure and confirms the prediction of structural conservation in this core complex. Analysis of particle heterogeneity also suggests how the SctS/FliQ subunits sequentially assemble in the complex. IMPORTANCE Although predicted on the basis of sequence conservation, the work presented here formally demonstrates that all classes of type III secretion systems, flagellar or virulence, share the same architecture at the level of the core structures. This absolute conservation of the unusual extramembrane structure of the core export gate complex now allows work to move to focusing on both mechanistic studies of type III but also on fundamental studies of how such a complex is assembled.

scholarly journals Regulation of the Locus of Enterocyte Effacement in Attaching and Effacing Pathogens

2017 ◽  
Vol 200 (2) ◽  
Author(s):  
R. Christopher D. Furniss ◽  
Abigail Clements
Keyword(s):  
Secretion System ◽  
Effector Proteins ◽  
The Core ◽  
Genetic Feature ◽  
Gut Epithelium ◽  
Type Three Secretion System ◽  
Gut Mucosa ◽  
Enterocyte Effacement ◽  
Type Three Secretion ◽  
Complex Regulatory Network

ABSTRACTAttaching and effacing (AE) pathogens colonize the gut mucosa using a type three secretion system (T3SS) and a suite of effector proteins. The locus of enterocyte effacement (LEE) is the defining genetic feature of the AE pathogens, encoding the T3SS and the core effector proteins necessary for pathogenesis. Extensive research has revealed a complex regulatory network that senses and responds to a myriad of host- and microbiota-derived signals in the infected gut to control transcription of the LEE. These signals include microbiota-liberated sugars and metabolites in the gut lumen, molecular oxygen at the gut epithelium, and host hormones. Recent research has revealed that AE pathogens also recognize physical signals, such as attachment to the epithelium, and that the act of effector translocation remodels gene expression in infecting bacteria. In this review, we summarize our knowledge to date and present an integrated view of how chemical, geographical, and physical cues regulate the virulence program of AE pathogens during infection.

scholarly journals Genetic code expansion enables visualization of Salmonella type three secretion system components and secreted effectors

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Moirangthem Kiran Singh ◽  
Parisa Zangoui ◽  
Yuki Yamanaka ◽  
Linda J Kenney
Keyword(s):  
Genetic Code ◽  
Unnatural Amino Acids ◽  
Click Reaction ◽  
Host Cells ◽  
Secretion Systems ◽  
Cellular Functions ◽  
Small Proteins ◽  
Type Three Secretion System ◽  
Genetic Code Expansion ◽  
Type Three Secretion

Type three secretion systems enable bacterial pathogens to inject effectors into the cytosol of eukaryotic hosts to reprogram cellular functions. It is technically challenging to label effectors and the secretion machinery without disrupting their structure/function. Herein, we present a new approach for labeling and visualization of previously intractable targets. Using genetic code expansion, we site-specifically labeled SsaP, the substrate specificity switch, and SifA, a here-to-fore unlabeled secreted effector. SsaP was secreted at later infection times; SsaP labeling demonstrated the stochasticity of injectisome and effector expression. SifA was labeled after secretion into host cells via fluorescent unnatural amino acids or non-fluorescent labels and a subsequent click reaction. We demonstrate the superiority of imaging after genetic code expansion compared to small molecule tags. It provides an alternative for labeling proteins that do not tolerate N- or C-terminal tags or fluorophores and thus is widely applicable to other secreted effectors and small proteins.

scholarly journals The flagellar substrate specificity switch protein FlhB assembles onto the extra-membrane export gate to regulate type three secretion

2019 ◽  
Author(s):  
Lucas Kuhlen ◽  
Steven Johnson ◽  
Andreas Zeitler ◽  
Sandra Bäurle ◽  
Justin C. Deme ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Conformational Changes ◽  
Complex Structure ◽  
Transmembrane Helices ◽  
Secretion Systems ◽  
Cryo Electron Microscopy ◽  
Type 3 Secretion System ◽  
Membrane Location ◽  
Type 3 ◽  
Type Three Secretion

AbstractExport of proteins through type three secretion systems (T3SS) is critical for motility and virulence of many major bacterial pathogens. Proteins are transported through an export gate complex consisting of three proteins (FliPQR in flagellar systems, SctRST in virulence systems) that were initially annotated as membrane proteins, but which we have recently shown assemble into an extra-membranous helical assembly. A fourth putative membrane protein (FlhB/SctU) is essential to the export process, and also functions to “switch” secretion substrate specificity once the growing hook/needle structures reach their determined length. Here we present the structure of an export gate containing the switch protein from a Vibrio polar flagellar system at 3.2 Å resolution by cryo-electron microscopy. The structure reveals that the FlhB/SctU further extends the helical export gate assembly with its four putative transmembrane helices adopting an out-of-membrane location, wrapped around the other export gate components at the base of the structure. The unusual topology of the switch protein helices creates a loop that wraps around the bottom of the closed export gate complex. Structure-informed mutagenesis suggests that this loop is critical in gating secretion and we propose that a series of conformational changes in the type 3 secretion system trigger opening of the export gate through the interactions between FlhB/SctU and FliPQR/SctRST.

An inhibitory mechanism of action of coiled‐coil peptides against type three secretion system from enteropathogenicEscherichia coli

2019 ◽  
Vol 25 (3) ◽  
Author(s):  
Mariano Larzábal ◽  
Hector A. Baldoni ◽  
Fernando D. Suvire ◽  
Lucrecia M. Curto ◽  
Gabriela E. Gomez ◽  
...  
Keyword(s):  
Mechanism Of Action ◽  
Coiled Coil ◽  
Secretion System ◽  
Inhibitory Mechanism ◽  
Type Three Secretion System ◽  
Type Three Secretion

scholarly journals Structures of Rhodopseudomonas palustris RC-LH1 complexes with open or closed quinone channels

2021 ◽  
Vol 7 (3) ◽  
pp. eabe2631
Author(s):  
David J. K. Swainsbury ◽  
Pu Qian ◽  
Philip J. Jackson ◽  
Kaitlyn M. Faries ◽  
Dariusz M. Niedzwiedzki ◽  
...  
Keyword(s):  
Binding Sites ◽  
Rhodopseudomonas Palustris ◽  
Ring Closure ◽  
Light Harvesting Complex ◽  
The Core ◽  
Quinone Binding ◽  
Cryo Electron Microscopy ◽  
Unique Protein ◽  
Center Light ◽  
Resolution Structure

The reaction-center light-harvesting complex 1 (RC-LH1) is the core photosynthetic component in purple phototrophic bacteria. We present two cryo–electron microscopy structures of RC-LH1 complexes from Rhodopseudomonas palustris. A 2.65-Å resolution structure of the RC-LH114-W complex consists of an open 14-subunit LH1 ring surrounding the RC interrupted by protein-W, whereas the complex without protein-W at 2.80-Å resolution comprises an RC completely encircled by a closed, 16-subunit LH1 ring. Comparison of these structures provides insights into quinone dynamics within RC-LH1 complexes, including a previously unidentified conformational change upon quinone binding at the RC QB site, and the locations of accessory quinone binding sites that aid their delivery to the RC. The structurally unique protein-W prevents LH1 ring closure, creating a channel for accelerated quinone/quinol exchange.

scholarly journals Avolition as the core negative symptom in schizophrenia: relevance to pharmacological treatment development

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Gregory P. Strauss ◽  
Lisa A. Bartolomeo ◽  
Philip D. Harvey
Keyword(s):  
Negative Symptom ◽  
Negative Symptoms ◽  
Clinical Trial Design ◽  
Core Component ◽  
Treatment Target ◽  
Global Improvement ◽  
Treatment Development ◽  
The Core ◽  
Current Review ◽  
Symptom Domains

AbstractNegative symptoms have long been considered a core component of schizophrenia. Modern conceptualizations of the structure of negative symptoms posit that there are at least two broad dimensions (motivation and pleasure and diminished expression) or perhaps five separable domains (avolition, anhedonia, asociality, blunted affect, alogia). The current review synthesizes a body of emerging research indicating that avolition may have a special place among these dimensions, as it is generally associated with poorer outcomes and may have distinct neurobiological mechanisms. Network analytic findings also indicate that avolition is highly central and interconnected with the other negative symptom domains in schizophrenia, and successfully remediating avolition results in global improvement in the entire constellation of negative symptoms. Avolition may therefore reflect the most critical treatment target within the negative symptom construct. Implications for targeted treatment development and clinical trial design are discussed.

scholarly journals Lin28, a major translation reprogramming factor, gains access to YB-1-packaged mRNA through its cold-shock domain

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Anastasiia Samsonova ◽  
Krystel El Hage ◽  
Bénédicte Desforges ◽  
Vandana Joshi ◽  
Marie-Jeanne Clément ◽  
...  
Keyword(s):  
Cancer Progression ◽  
Cold Shock ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Rna Binding Protein ◽  
Large Set ◽  
Core Component ◽  
The Core ◽  
Cold Shock Domain ◽  
Global Translation

AbstractThe RNA-binding protein Lin28 (Lin28a) is an important pluripotency factor that reprograms translation and promotes cancer progression. Although Lin28 blocks let-7 microRNA maturation, Lin28 also binds to a large set of cytoplasmic mRNAs directly. However, how Lin28 regulates the processing of many mRNAs to reprogram global translation remains unknown. We show here, using a structural and cellular approach, a mixing of Lin28 with YB-1 (YBX1) in the presence of mRNA owing to their cold-shock domain, a conserved β-barrel structure that binds to ssRNA cooperatively. In contrast, the other RNA binding-proteins without cold-shock domains tested, HuR, G3BP-1, FUS and LARP-6, did not mix with YB-1. Given that YB-1 is the core component of dormant mRNPs, a model in which Lin28 gains access to mRNPs through its co-association with YB-1 to mRNA may provide a means for Lin28 to reprogram translation. We anticipate that the translational plasticity provided by mRNPs may contribute to Lin28 functions in development and adaptation of cancer cells to an adverse environment.

scholarly journals Enhancing Postgraduate Learning and Teaching: Postgraduate Summer School in Dairy Science

2014 ◽  
Vol 2014 ◽  
pp. 1-7 ◽  
Author(s):  
Pietro Celi ◽  
Gianfranco Gabai ◽  
Massimo Morgante ◽  
Luigi Gallo
Keyword(s):  
Summer School ◽  
Research Collaboration ◽  
Learning Experience ◽  
Scientific Investigation ◽  
Core Component ◽  
Learning And Teaching ◽  
Transferable Skills ◽  
The Core ◽  
Dairy Science ◽  
The University

Dairy science is a multidisciplinary area of scientific investigation and Ph.D. students aiming to do research in the field of animal and/or veterinary sciences must be aware of this. Ph.D. students often have vast spectra of research interests, and it is quite challenging to satisfy the expectation of all of them. The aim of this study was to establish an international Ph.D. training program based on research collaboration between the University of Sydney and the University of Padova. The core component of this program was a two-week Postgraduate Summer School in Dairy Science, which was held at the University of Padova, for Ph.D. students of both universities. Therefore, we designed a program that encompassed seminars, workshops, laboratory practical sessions, and farm visits. Participants were surveyed using a written questionnaire. Overall, participants have uniformly praised the Summer School calling it a rewarding and valuable learning experience. The Ph.D. Summer School in Dairy Science provided its participants a positive learning experience, provided them the opportunity to establish an international network, and facilitated the development of transferable skills.

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