<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network

ABSTRACT Following infection of mammalian cells, Salmonella enterica serovar Typhimurium (S. Typhimurium) replicates within membrane-bound compartments known as Salmonella-containing vacuoles (SCVs). The Salmonella pathogenicity island 2 type III secretion system (SPI-2 T3SS) translocates approximat...

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Autores principales:	Xiu-Jun Yu, Mei Liu, David W. Holden
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2016
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/136b6510cad34a2fb9b53439e92517b6
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spelling	oai:doaj.org-article:136b6510cad34a2fb9b53439e92517b62021-11-15T15:50:18Z<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network10.1128/mBio.00474-162150-7511https://doaj.org/article/136b6510cad34a2fb9b53439e92517b62016-09-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00474-16https://doaj.org/toc/2150-7511ABSTRACT Following infection of mammalian cells, Salmonella enterica serovar Typhimurium (S. Typhimurium) replicates within membrane-bound compartments known as Salmonella-containing vacuoles (SCVs). The Salmonella pathogenicity island 2 type III secretion system (SPI-2 T3SS) translocates approximately 30 different effectors across the vacuolar membrane. SseF and SseG are two such effectors that are required for SCVs to localize close to the Golgi network in infected epithelial cells. In a yeast two-hybrid assay, SseG and an N-terminal variant of SseF interacted directly with mammalian ACBD3, a multifunctional cytosolic Golgi network-associated protein. Knockdown of ACBD3 by small interfering RNA (siRNA) reduced epithelial cell Golgi network association of wild-type bacteria, phenocopying the effect of null mutations of sseG or sseF. Binding of SseF to ACBD3 in infected cells required the presence of SseG. A single-amino-acid mutant of SseG and a double-amino-acid mutant of SseF were obtained that did not interact with ACBD3 in Saccharomyces cerevisiae. When either of these was produced together with the corresponding wild-type effector by Salmonella in infected cells, they enabled SCV-Golgi network association and interacted with ACBD3. However, these properties were lost and bacteria displayed an intracellular replication defect when cells were infected with Salmonella carrying both mutant genes. Knockdown of ACBD3 resulted in a replication defect of wild-type bacteria but did not further attenuate the growth defect of a ΔsseFG mutant strain. We propose a model in which interaction between SseF and SseG enables both proteins to bind ACBD3, thereby anchoring SCVs at the Golgi network and facilitating bacterial replication. IMPORTANCE Upon invasion of epithelial cells, the majority of vacuoles containing Salmonella enterica migrate to the perinuclear region-located Golgi network and remain in this region of the cell during the first few rounds of bacterial replication, forming a clustered microcolony of vacuoles. This process requires the action of SseF and SseG, two effector proteins that are translocated by the Salmonella SPI-2 type III secretion system. However, little is known about how they function. Here, we show that both proteins interact with the mammalian Golgi network-associated protein ACBD3. To our knowledge, the SseF-SseG-ACBD3 interaction is the first example of a tethering complex between a pathogen-containing vacuole and a host cell organelle.Xiu-Jun YuMei LiuDavid W. HoldenAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 7, Iss 4 (2016)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Xiu-Jun Yu Mei Liu David W. Holden <italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network
description	ABSTRACT Following infection of mammalian cells, Salmonella enterica serovar Typhimurium (S. Typhimurium) replicates within membrane-bound compartments known as Salmonella-containing vacuoles (SCVs). The Salmonella pathogenicity island 2 type III secretion system (SPI-2 T3SS) translocates approximately 30 different effectors across the vacuolar membrane. SseF and SseG are two such effectors that are required for SCVs to localize close to the Golgi network in infected epithelial cells. In a yeast two-hybrid assay, SseG and an N-terminal variant of SseF interacted directly with mammalian ACBD3, a multifunctional cytosolic Golgi network-associated protein. Knockdown of ACBD3 by small interfering RNA (siRNA) reduced epithelial cell Golgi network association of wild-type bacteria, phenocopying the effect of null mutations of sseG or sseF. Binding of SseF to ACBD3 in infected cells required the presence of SseG. A single-amino-acid mutant of SseG and a double-amino-acid mutant of SseF were obtained that did not interact with ACBD3 in Saccharomyces cerevisiae. When either of these was produced together with the corresponding wild-type effector by Salmonella in infected cells, they enabled SCV-Golgi network association and interacted with ACBD3. However, these properties were lost and bacteria displayed an intracellular replication defect when cells were infected with Salmonella carrying both mutant genes. Knockdown of ACBD3 resulted in a replication defect of wild-type bacteria but did not further attenuate the growth defect of a ΔsseFG mutant strain. We propose a model in which interaction between SseF and SseG enables both proteins to bind ACBD3, thereby anchoring SCVs at the Golgi network and facilitating bacterial replication. IMPORTANCE Upon invasion of epithelial cells, the majority of vacuoles containing Salmonella enterica migrate to the perinuclear region-located Golgi network and remain in this region of the cell during the first few rounds of bacterial replication, forming a clustered microcolony of vacuoles. This process requires the action of SseF and SseG, two effector proteins that are translocated by the Salmonella SPI-2 type III secretion system. However, little is known about how they function. Here, we show that both proteins interact with the mammalian Golgi network-associated protein ACBD3. To our knowledge, the SseF-SseG-ACBD3 interaction is the first example of a tethering complex between a pathogen-containing vacuole and a host cell organelle.
format	article
author	Xiu-Jun Yu Mei Liu David W. Holden
author_facet	Xiu-Jun Yu Mei Liu David W. Holden
author_sort	Xiu-Jun Yu
title	<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network
title_short	<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network
title_full	<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network
title_fullStr	<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network
title_full_unstemmed	<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network
title_sort	<italic toggle="yes">salmonella</italic> effectors ssef and sseg interact with mammalian protein acbd3 (gcp60) to anchor <italic toggle="yes">salmonella</italic>-containing vacuoles at the golgi network
publisher	American Society for Microbiology
publishDate	2016
url	https://doaj.org/article/136b6510cad34a2fb9b53439e92517b6
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<italic toggle="yes">Salmonella</italic> Effectors SseF and SseG Interact with Mammalian Protein ACBD3 (GCP60) To Anchor <italic toggle="yes">Salmonella</italic>-Containing Vacuoles at the Golgi Network

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