scholarly journals Activation ofShigella flexneritype 3 secretion requires a host-induced conformational change to the translocon pore

2019 ◽  
Author(s):  
Brian C. Russo ◽  
Jeffrey K. Duncan ◽  
Alexandra L. Wiscovitch ◽  
Austin C. Hachey ◽  
Marcia B. Goldberg
Keyword(s):  
Intermediate Filaments ◽  
Secretion System ◽  
Human Cells ◽  
Bacterial Virulence ◽  
Secretion Systems ◽  
Virulence Proteins ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Effector Secretion ◽  
Pore Protein

AbstractType 3 secretion systems (T3SSs) are conserved bacterial nanomachines that inject virulence proteins (effectors) into eukaryotic cells during infection. Due to their ability to introduce heterologous protein into human cells, these systems are being developed as therapeutic delivery devices. The T3SS assembles a translocon pore in the plasma membrane and then docks onto the pore. Docking activates effector secretion through the pore and into the host cytosol. Here, usingShigella flexneri, a model pathogen for the study of type 3 secretion, we determined the molecular mechanisms by which host intermediate filaments trigger docking and enable effector secretion. We show that the interaction of intermediate filaments with the translocon pore protein IpaC changed the pore’s conformation in a manner that was required for docking. Intermediate filaments repositioned residues of theShigellapore protein IpaC that are located on the surface of the pore and in the pore channel. Restricting these conformational changes blocked docking in an intermediate filament-dependent manner. These data demonstrate that a host-induced conformational change to the pore enables T3SS docking and effector secretion, providing new mechanistic insight into the regulation of type 3 secretion.Author summaryThe movement of bacterial proteins across membranes is essential for bacterial physiology and bacterial virulence. The type 3 secretion system moves bacterial virulence proteins from the inside of bacterial pathogens into human cells. To do so, the type 3 secretion system forms a pore in the plasma membrane of the target cell, attaches (docks) onto the pore, and delivers virulence proteins through the pore. Docking is essential for establishing a continuous channel from the inside of the bacteria to the inside of the human cell. What enables the type 3 secretion system to dock onto pores is not understood. We show that structural proteins in human cells, intermediate filament proteins, induce structural rearrangements to the type 3 secretion pore that trigger docking and that enable the subsequent delivery of virulence proteins into human cells. Due to the wide prevalence of type 3 secretion systems among human pathogens, these findings are likely to broadly enhance our understanding of type 3 secretion.

scholarly journals Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection

mBio ◽  
2019 ◽  
Vol 10 (3) ◽  
Author(s):  
Brian C. Russo ◽  
Jeffrey K. Duncan ◽  
Marcia B. Goldberg
Keyword(s):  
Plasma Membrane ◽  
Protein Secretion ◽  
Secretion System ◽  
Bacterial Virulence ◽  
Eukaryotic Cells ◽  
Effector Protein ◽  
Virulence Proteins ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Pore Protein

ABSTRACTMany Gram-negative bacterial pathogens require a type 3 secretion system (T3SS) to deliver effector proteins into eukaryotic cells. Contact of the tip complex of the T3SS with a target eukaryotic cell initiates secretion of the two bacterial proteins that assemble into the translocon pore in the plasma membrane. The translocon pore functions to regulate effector protein secretion and is the conduit for effector protein translocation across the plasma membrane. To generate insights into how the translocon pore regulates effector protein secretion, we defined the topology of theShigellatranslocon pore protein IpaC in the plasma membrane following its native delivery by the T3SS. Using single cysteine substitution mutagenesis and site-directed labeling with a membrane-impermeant chemical probe, we mapped residues accessible from the extracellular surface of the cell. Our data support a model in which the N terminus of IpaC is extracellular and the C terminus of IpaC is intracellular. These findings resolve previously conflicting data on IpaC topology that were based on nonnative delivery of IpaC to membranes.Salmonella entericaserovar Typhimurium also requires the T3SS for effector protein delivery into eukaryotic cells. Although the sequence of IpaC is closely related to theSalmonellatranslocon pore protein SipC, the two proteins have unique functional attributes during infection. We showed a similar overall topology for SipC and IpaC and identified subtle topological differences between their transmembrane α-helixes and C-terminal regions. Together, our data suggest that topological differences among distinct translocon pore proteins may dictate organism-specific functional differences of the T3SSs during infection.IMPORTANCEThe type 3 secretion system (T3SS) is a nanomachine required for virulence of many bacterial pathogens that infect humans. The system delivers bacterial virulence proteins into the cytosol of human cells, where the virulence proteins promote bacterial infection. The T3SS forms a translocon pore in the membranes of target cells. This pore is the portal through which bacterial virulence proteins are delivered by the T3SS into the eukaryotic cytosol. The pore also regulates secretion of these virulence proteins. Our work defines the topology of translocon pore proteins in their native context during infection, resolves previously conflicting reports about the topology of theShigellatranslocon pore protein IpaC, and provides new insights into how interactions of the pore with the T3SS likely produce signals that activate secretion of virulence proteins.

scholarly journals Topological analysis of the type 3 secretion system translocon pore protein IpaC following its native delivery to the plasma membrane during infection

2019 ◽  
Author(s):  
Brian C. Russo ◽  
Jeffrey K. Duncan ◽  
Marcia B. Goldberg
Keyword(s):  
Plasma Membrane ◽  
Protein Secretion ◽  
Secretion System ◽  
Bacterial Virulence ◽  
Eukaryotic Cells ◽  
Effector Protein ◽  
Virulence Proteins ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Pore Protein

AbstractMany Gram-negative bacterial pathogens require a type 3 secretion system (T3SS) to deliver effector proteins into eukaryotic cells. Contact of the tip complex of the T3SS with a target eukaryotic cell initiates the secretion of the two bacterial proteins that assemble into the translocon pore in the plasma membrane. The translocon pore functions to regulate effector protein secretion and is the conduit for effector protein translocation across the plasma membrane. To generate insights into how the translocon pore regulates effector protein secretion, we defined the topology of theShigellatranslocon pore protein IpaC in the plasma membrane following its native delivery by the T3SS. Using single-cysteine substitution mutagenesis and site-directed labeling with a membrane-impermeant chemical probe, we mapped residues accessible from the extracellular surface of the cell. Our data support a model in which the N-terminus of IpaC is extracellular and the C-terminus of IpaC is intracellular. These findings resolve previously conflicting data on IpaC topology that were based on non-native delivery of IpaC to membranes.Salmonella entericaserovar Typhimurium also requires the T3SS for effector protein delivery into eukaryotic cells. Although the sequence of IpaC is closely related to theSalmonellatranslocon pore protein SipC, the two proteins have unique functional attributes during infection. We showed a similar overall topology for SipC and IpaC and identified subtle topological differences between their transmembrane α-helixes and C-terminal regions. Together, our data suggest that topological differences among distinct translocon pore proteins may dictate organism-specific functional differences of the T3SSs during infection.ImportanceThe type 3 secretion system (T3SS) is a nanomachine required for virulence of many bacterial pathogens that infect humans. The system delivers bacterial virulence proteins into the cytosol of human cells, where the virulence proteins promote bacterial infection. The T3SS forms a translocon pore in the membrane of target cells. This pore is the portal through which bacterial virulence proteins are delivered by the T3SS into the eukaryotic cytosol. The pore also regulates the secretion of these virulence proteins. Our work defines the topology of translocon pore proteins in their native context during infection, resolves previously conflicting reports about the topology of theShigellatranslocon pore protein IpaC, and provides new insights into how interactions of the pore with the T3SS likely produce signals that activate secretion of virulence proteins.

scholarly journals Topology and Contribution to the Pore Channel Lining of Plasma Membrane-Embedded Shigella flexneri Type 3 Secretion Translocase IpaB

mBio ◽  
2021 ◽  
Author(s):  
Poyin Chen ◽  
Brian C. Russo ◽  
Jeffrey K. Duncan-Lowey ◽  
Natasha Bitar ◽  
Keith T. Egger ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Human Cell ◽  
Shigella Flexneri ◽  
Cellular Function ◽  
Human Cells ◽  
Bacterial Virulence ◽  
Pore Channel ◽  
Secretion Systems ◽  
Virulence Proteins ◽  
Type 3

Type 3 secretion systems are nanomachines employed by many bacteria, including Shigella , which deliver into human cells bacterial virulence proteins that alter cellular function in ways that promote infection. Delivery of Shigella virulence proteins occurs through a pore formed in human cell membranes by the IpaB and IpaC proteins.

scholarly journals The type 3 secretion system requires actin polymerization to open translocon pores

2021 ◽  
Vol 17 (9) ◽  
pp. e1009932
Author(s):  
Brian C. Russo ◽  
Jeffrey K. Duncan-Lowey ◽  
Poyin Chen ◽  
Marcia B. Goldberg
Keyword(s):  
Plasma Membrane ◽  
Actin Polymerization ◽  
Shigella Flexneri ◽  
Coiled Coil ◽  
Model Organism ◽  
Secretion System ◽  
Pore Channel ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Pore Protein

Many bacterial pathogens require a type 3 secretion system (T3SS) to establish a niche. Host contact activates bacterial T3SS assembly of a translocon pore in the host plasma membrane. Following pore formation, the T3SS docks onto the translocon pore. Docking establishes a continuous passage that enables the translocation of virulence proteins, effectors, into the host cytosol. Here we investigate the contribution of actin polymerization to T3SS-mediated translocation. Using the T3SS model organism Shigella flexneri, we show that actin polymerization is required for assembling the translocon pore in an open conformation, thereby enabling effector translocation. Opening of the pore channel is associated with a conformational change to the pore, which is dependent upon actin polymerization and a coiled-coil domain in the pore protein IpaC. Analysis of an IpaC mutant that is defective in ruffle formation shows that actin polymerization-dependent pore opening is distinct from the previously described actin polymerization-dependent ruffles that are required for bacterial internalization. Moreover, actin polymerization is not required for other pore functions, including docking or pore protein insertion into the plasma membrane. Thus, activation of the T3SS is a multilayered process in which host signals are sensed by the translocon pore leading to the activation of effector translocation.

scholarly journals A Primed Subpopulation of Bacteria Enables Rapid Expression of the Type 3 Secretion System in Pseudomonas aeruginosa

mBio ◽  
2021 ◽  
Author(s):  
Christina K. Lin ◽  
Daniel S. W. Lee ◽  
Saria McKeithen-Mead ◽  
Thierry Emonet ◽  
Barbara Kazmierczak
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Secretion System ◽  
Innate Immune ◽  
Secretion Systems ◽  
Content Type ◽  
Acute Infections ◽  
A Cell ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Specific Virulence

The expression of specific virulence traits is strongly associated with Pseudomonas aeruginosa ’s success in establishing acute infections but is thought to carry a cost for bacteria. Producing multiprotein secretion systems or motility organelles is metabolically expensive and can target a cell for recognition by innate immune system receptors that recognize structural components of the type 3 secretion system (T3SS) or flagellum.

scholarly journals The type 3 secretion system requires actin polymerization to open translocon pores

2021 ◽  
Author(s):  
Brian C. Russo ◽  
Jeffrey K. Duncan-Lowey ◽  
Poyin Chen ◽  
Marcia B. Goldberg
Keyword(s):  
Plasma Membrane ◽  
Actin Polymerization ◽  
Shigella Flexneri ◽  
Coiled Coil ◽  
Model Organism ◽  
Secretion System ◽  
Pore Channel ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Pore Protein

Many bacterial pathogens require a type 3 secretion system (T3SS) to establish a niche. Host contact activates bacterial T3SS assembly of a translocon pore in the host plasma membrane. Following pore formation, the T3SS docks onto the translocon pore. Docking establishes a continuous passage that enables the translocation of virulence proteins, effectors, into the host cytosol. Here we investigate the contribution of actin polymerization to T3SS-mediated translocation. Using the T3SS model organism Shigella flexneri, we show that actin polymerization is required for assembling the translocon pore in an open conformation, thereby enabling effector translocation. Opening of the pore channel is associated with a conformational change to the pore, which is dependent upon actin polymerization and a coiled-coil domain in the pore protein IpaC. An IpaC mutant is identified that shows actin polymerization-dependent pore opening is distinct from the previously described actin polymerization-dependent ruffles that are required for bacterial internalization. Moreover, actin polymerization is not required for other pore functions, including docking or pore protein insertion into the plasma membrane. Thus, activation of the T3SS is a multilayered process in which host signals are sensed by the translocon pore leading to the activation of effector translocation.

scholarly journals Orchestrated delivery of Legionella effectors by the Icm/Dot secretion system

2019 ◽  
Author(s):  
Julie Allombert ◽  
Corentin Jaboulay ◽  
Céline Michard ◽  
Claire Andréa ◽  
Xavier Charpentier ◽  
...  
Keyword(s):  
Host Cell ◽  
Legionella Pneumophila ◽  
Translational Control ◽  
Function Analysis ◽  
Secretion System ◽  
Secretion Systems ◽  
Type Iv ◽  
Set Up ◽  
Type 3 ◽  
Effector Secretion

AbstractLegionella pneumophila uses the Icm/Dot Type IV secretion system (T4SS) to translocate a record number (300) of bacterial effectors into the host cell. Despite recent breakthrough progress in determining the structure and the localization of the secretion machinery, it is still a challenge to understand how the delivery of so many effectors is organized to avoid bottleneck effect and to allow effective manipulation of the host cell by L. pneumophila. Here, we demonstrate that secretion of effectors is ordered and so precisely set up that it lines-up with the delivery timing required for the function of the effectors in the cell. We observe notably that the secretion order of 4 effectors targeting Rab1 is fully consistent with the sequence of their actions on Rab1. Importantly, we show that the timed delivery of an effector is not dependent on its concentration, nor on its picking-up by chaperone proteins. Conversely, this control involves c-di-GMP signaling, as a c-di-GMP synthesizing enzyme, namely the diguanylate cyclase Lpl0780/Lpp0809, significantly contributes to accurate triggering of effector secretion via a post-translational control of the T4SS machinery at the bacterial pole.SignificanceType 3, 4 and 6 secretion systems are multiprotein complex known to be crucial for infectious cycle of many bacterial pathogens. Despite considerable progress on several fronts in structure-function analysis of these systems, one of the blackest boxes in our understanding is the signal that triggers the activation of effectors transfer. This is particularly true for the Icm/Dot T4SS in L. pneumophila that deals with the translocation of a record number of 300 effectors. We demonstrate that Icm/Dot secretion is timely fine-tuned and most importantly, that the complex orchestration of so many effector actions relies at least in part on the defined timing of their translocation into the host cell. Also, we highlight for the first time a post-translational control of a T4SS by c-diGMP signaling.

scholarly journals The HrpG/HrpX Regulon of Xanthomonads—An Insight to the Complexity of Regulation of Virulence Traits in Phytopathogenic Bacteria

2021 ◽  
Vol 9 (1) ◽  
pp. 187
Author(s):  
Doron Teper ◽  
Sheo Shankar Pandey ◽  
Nian Wang
Keyword(s):  
Regulatory Network ◽  
Regulatory Networks ◽  
Focal Point ◽  
Genetic Regulation ◽  
Transcriptional Regulators ◽  
Secretion System ◽  
In Planta ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Made In

Bacteria of the genus Xanthomonas cause a wide variety of economically important diseases in most crops. The virulence of the majority of Xanthomonas spp. is dependent on secretion and translocation of effectors by the type 3 secretion system (T3SS) that is controlled by two master transcriptional regulators HrpG and HrpX. Since their discovery in the 1990s, the two regulators were the focal point of many studies aiming to decipher the regulatory network that controls pathogenicity in Xanthomonas bacteria. HrpG controls the expression of HrpX, which subsequently controls the expression of T3SS apparatus genes and effectors. The HrpG/HrpX regulon is activated in planta and subjected to tight metabolic and genetic regulation. In this review, we cover the advances made in understanding the regulatory networks that control and are controlled by the HrpG/HrpX regulon and their conservation between different Xanthomonas spp.

scholarly journals Cytosolic Assembly Among Bacterial Type 3 Secretion System Proteins Revealed by High-Throughput Single-Molecule Tracking

2018 ◽  
Vol 114 (3) ◽  
pp. 534a
Author(s):  
Julian Rocha ◽  
Charles Richardson ◽  
Mingxing Zhang ◽  
Andreas Diepold ◽  
Andreas Gahlmann
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Secretion System ◽  
Single Molecule Tracking ◽  
Type 3 Secretion System ◽  
Type 3 ◽  
Bacterial Type
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