Dataset Information

The Salmonella effector SteA contributes to the control of membrane dynamics of Salmonella-containing vacuoles.

ABSTRACT: Salmonella enterica serovar Typhimurium is a bacterial pathogen causing gastroenteritis in humans and a typhoid-like systemic disease in mice. S. Typhimurium virulence is related to its capacity to multiply intracellularly within a membrane-bound compartment, the Salmonella-containing vacuole (SCV), and depends on type III secretion systems that deliver bacterial effector proteins into host cells. Here, we analyzed the cellular function of the Salmonella effector SteA. We show that, compared to cells infected by wild-type S. Typhimurium, cells infected by ?steA mutant bacteria displayed fewer Salmonella-induced filaments (SIFs), an increased clustering of SCVs, and morphologically abnormal vacuoles containing more than one bacterium. The increased clustering of SCVs and the appearance of vacuoles containing more than one bacterium were suppressed by inhibition of the activity of the microtubule motor dynein or kinesin-1. Clustering and positioning of SCVs are controlled by the effectors SseF and SseG, possibly by helping to maintain a balanced activity of microtubule motors on the bacterial vacuoles. Deletion of steA in S. Typhimurium ?sseF or ?sseG mutants revealed that SteA contributes to the characteristic scattered distribution of ?sseF or ?sseG mutant SCVs in infected cells. Overall, this shows that SteA participates in the control of SCV membrane dynamics. Moreover, it indicates that SteA is functionally linked to SseF and SseG and suggests that it might contribute directly or indirectly to the regulation of microtubule motors on the bacterial vacuoles.

SUBMITTER: Domingues L

PROVIDER: S-EPMC4097631 | biostudies-literature | 2014 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

The Salmonella effector SteA contributes to the control of membrane dynamics of Salmonella-containing vacuoles.

Domingues Lia L Holden David W DW Mota Luís Jaime LJ

Infection and immunity 20140428 7

Salmonella enterica serovar Typhimurium is a bacterial pathogen causing gastroenteritis in humans and a typhoid-like systemic disease in mice. S. Typhimurium virulence is related to its capacity to multiply intracellularly within a membrane-bound compartment, the Salmonella-containing vacuole (SCV), and depends on type III secretion systems that deliver bacterial effector proteins into host cells. Here, we analyzed the cellular function of the Salmonella effector SteA. We show that, compared to ...[more]

PMID: 24778114

Similar Datasets

Project description:BackgroundBoth typhoidal and non-typhoidal Salmonella infections remain a considerable cause of morbidity and mortality globally, and impose a major socio-economic burden worldwide. A key property of all pathogenic Salmonella strains is the ability to invade host cells and reside within an intracellular, vacuolar compartment called the Salmonella-containing vacuole (SCV). Although the SCV is involved in both immune-evasion and intracellular replication and spread within the host, information about the host:pathogen interactions at this interface are limited, in part due to the technical difficulties involved in purification of these vacuoles. While a number of column- or gradient-based methods have been applied, cross-contamination with other host cell organelles or rupture of the labile SCV membrane has further complicated efforts to successfully isolate SCVs.ResultsHere, we report the isolation of intact SCVs using carbon-coated, paramagnetic nanoparticles. The approach permits rapid isolation of intact SCVs from human macrophages in vitro without involving numerous purification steps. Bacteria are pre-labeled with modified nanoparticles prior to infection, and at various times post-infection, host cells are lysed and intact pathogen-containing phagosomes are recovered after application of a mild magnetic field. Purified, intact SCVs isolated using this method were shown to display high levels of co-association of internalized Salmonella with the standard SCV markers Rab5 and LAMP-1 using both microscopic and protein based methods.ConclusionThe method described is highly efficient, robust and permits rapid isolation of intact SCVs from human macrophages without involving numerous purification steps. The method can also be applied to other intracellular pathogens that reside within a vacuole-like compartment within host cells. Future work using the approach should aid in identification and characterization of host factors associated with the membranes of such intracellular pathogens, which could potentially serve as pharmaceutical targets against intracellular pathogens residing within vacuoles.

Project description:UnlabelledFollowing 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 ?sseFG 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.ImportanceUpon 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.

Dataset Information

The Salmonella effector SteA contributes to the control of membrane dynamics of Salmonella-containing vacuoles.

Publications

The Salmonella effector SteA contributes to the control of membrane dynamics of Salmonella-containing vacuoles.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets