Identification of interactions among host and bacterial proteins and evaluation of their role early during Shigella flexneri infection

Miller, Kelly A.; Garza-Mayers, Anna Cristina; Leung, Yiuka; Goldberg, Marcia B.

doi:10.1099/mic.0.000637

Cited by 12 publications

(11 citation statements)

References 32 publications

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“…To assess the ability of the type 3 secretion system to secrete proteins in the absence of eukaryotic cells, the T3SS was artificially activated by the addition of Congo red (7, 14, 27). Overnight cultures were back diluted, induced where appropriate with 1.2% arabinose, and cultured for 2 h at 37°C.…”

Section: Methodsmentioning

confidence: 99%

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

Russo

Duncan

Goldberg

2019

mBio

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Many 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 the Shigella translocon 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 enterica serovar Typhimurium also requires the T3SS for effector protein delivery into eukaryotic cells. Although the sequence of IpaC is closely related to the Salmonella translocon 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. IMPORTANCE The 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 the Shigella translocon 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.

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Section: Methodsmentioning

confidence: 99%

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

Russo

Duncan

Goldberg

2019

mBio

Self Cite

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“…To assess the ability of the type 3 secretion system to secrete proteins in the absence of eukaryotic cells, the T3SS was artificially activated by addition of Congo red (7,14,27). Overnight cultures were back diluted, induced where appropriate with 1.2% arabinose, and cultured for 2 hours at 37°C.…”

Section: Methodsmentioning

confidence: 99%

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

Russo

Duncan²,

Goldberg

2019

Preprint

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word count: 248 words Text word count: 5,371 words Abstract 1Many Gram-negative bacterial pathogens require a type 3 secretion system 2 (T3SS) to deliver effector proteins into eukaryotic cells. Contact of the tip complex of the 3 T3SS with a target eukaryotic cell initiates the secretion of the two bacterial proteins that 4 assemble into the translocon pore in the plasma membrane. The translocon pore 5 functions to regulate effector protein secretion and is the conduit for effector protein 6 translocation across the plasma membrane. To generate insights into how the 7 translocon pore regulates effector protein secretion, we defined the topology of the 8 Shigella translocon pore protein IpaC in the plasma membrane following its native 9 delivery by the T3SS. Using single-cysteine substitution mutagenesis and site-directed 10 labeling with a membrane-impermeant chemical probe, we mapped residues accessible 11 from the extracellular surface of the cell. Our data support a model in which the N-12 terminus of IpaC is extracellular and the C-terminus of IpaC is intracellular. These 13 findings resolve previously conflicting data on IpaC topology that were based on non-14

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“…Recent results also indicated that Toca-1, besides interacting with IcsB, also interacts with several other S. flexneri effectors, namely IpaB, OspC3 and IpgD. The function of these interactions and possible role in the evasion of the LAP-like response, autophagy and other aspects of S. flexneri pathogenesis remains to be further investigated ( Miller et al., 2018 ).…”

Section: Interactions Of Bacterial Pathogens With Lapmentioning

confidence: 99%

LAPped in Proof: LC3‐Associated Phagocytosis and the Arms Race Against Bacterial Pathogens

Grijmans

Kooij

Varela

et al. 2022

Front. Cell. Infect. Microbiol.

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Cells of the innate immune system continuously patrol the extracellular environment for potential microbial threats that are to be neutralized by phagocytosis and delivery to lysosomes. In addition, phagocytes employ autophagy as an innate immune mechanism against pathogens that succeed to escape the phagolysosomal pathway and invade the cytosol. In recent years, LC3-associated phagocytosis (LAP) has emerged as an intermediate between phagocytosis and autophagy. During LAP, phagocytes target extracellular microbes while using parts of the autophagic machinery to label the cargo-containing phagosomes for lysosomal degradation. LAP contributes greatly to host immunity against a multitude of bacterial pathogens. In the pursuit of survival, bacteria have developed elaborate strategies to disarm or circumvent the LAP process. In this review, we will outline the nature of the LAP mechanism and discuss recent insights into its interplay with bacterial pathogens.

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Identification of interactions among host and bacterial proteins and evaluation of their role early during Shigella flexneri infection

Cited by 12 publications

References 32 publications

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

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

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

LAPped in Proof: LC3‐Associated Phagocytosis and the Arms Race Against Bacterial Pathogens

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