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

ABSTRACT 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 i...

Descripción completa

Guardado en:

Detalles Bibliográficos
Autores principales:	Brian C. Russo, Jeffrey K. Duncan, Marcia B. Goldberg
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	IpaC Salmonella Shigella flexneri SipC topology type 3 secretion Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/81d33601ba294611b2a8188cfc568889
Etiquetas:	Agregar Etiqueta Sin Etiquetas, Sea el primero en etiquetar este registro!

id	oai:doaj.org-article:81d33601ba294611b2a8188cfc568889
record_format	dspace
spelling	oai:doaj.org-article:81d33601ba294611b2a8188cfc5688892021-11-15T15:55:24ZTopological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection10.1128/mBio.00877-192150-7511https://doaj.org/article/81d33601ba294611b2a8188cfc5688892019-06-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00877-19https://doaj.org/toc/2150-7511ABSTRACT 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.Brian C. RussoJeffrey K. DuncanMarcia B. GoldbergAmerican Society for MicrobiologyarticleIpaCSalmonellaShigella flexneriSipCtopologytype 3 secretionMicrobiologyQR1-502ENmBio, Vol 10, Iss 3 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	IpaC Salmonella Shigella flexneri SipC topology type 3 secretion Microbiology QR1-502
spellingShingle	IpaC Salmonella Shigella flexneri SipC topology type 3 secretion Microbiology QR1-502 Brian C. Russo Jeffrey K. Duncan Marcia B. Goldberg Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection
description	ABSTRACT 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.
format	article
author	Brian C. Russo Jeffrey K. Duncan Marcia B. Goldberg
author_facet	Brian C. Russo Jeffrey K. Duncan Marcia B. Goldberg
author_sort	Brian C. Russo
title	Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection
title_short	Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection
title_full	Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection
title_fullStr	Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection
title_full_unstemmed	Topological Analysis of the Type 3 Secretion System Translocon Pore Protein IpaC following Its Native Delivery to the Plasma Membrane during Infection
title_sort	topological analysis of the type 3 secretion system translocon pore protein ipac following its native delivery to the plasma membrane during infection
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/81d33601ba294611b2a8188cfc568889
work_keys_str_mv	AT briancrusso topologicalanalysisofthetype3secretionsystemtransloconporeproteinipacfollowingitsnativedeliverytotheplasmamembraneduringinfection AT jeffreykduncan topologicalanalysisofthetype3secretionsystemtransloconporeproteinipacfollowingitsnativedeliverytotheplasmamembraneduringinfection AT marciabgoldberg topologicalanalysisofthetype3secretionsystemtransloconporeproteinipacfollowingitsnativedeliverytotheplasmamembraneduringinfection
_version_	1718427227339620352

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

Ejemplares similares