Dataset Information

Secretion of Salmonella Pathogenicity Island 1-Encoded Type III Secretion System Effectors by Outer Membrane Vesicles in Salmonella enterica Serovar Typhimurium.

ABSTRACT: Outer membrane vesicles (OMVs) are spherical membranous structures released by Gram-negative bacteria. Several bacterial pathogens utilize OMVs as vehicles for the delivery of virulence factors into host cells. Results of our previous study on proteomic analysis revealed that OMVs isolated from Salmonella enterica serovar Typhimurium had virulence effectors that are known to be translocated by Salmonella pathogenicity island 1 (SPI-1)-encoded type III secretion system (T3SS1) into the host cell. In the present study, immunoblot analysis confirmed the secretion of the six T3SS1 effector proteins, namely SipB and SipC (translocators of T3SS1), and SipA, SopA, SopB, and SopE2 (effectors translocated by T3SS1), by OMVs. Results of proteinase K treatment revealed the localization of these T3SS1 effector proteins on the outer surface of OMVs. SipC and SopE2 were secreted by OMVs independent of the three secretion systems T3SS1, T3SS2, and flagella, signifying OMVs to be an alternative delivery system to T3SSs. T3SS1 effectors SipA, SipC, and SopE2 were internalized into the cytoplasm of the host cell by OMVs independent of cellular Salmonella-host cell contact. In epithelial cells, addition of OMVs harboring T3SS1 effectors stimulated the production of F-actin, thereby complementing the attenuated invasion of ?sopE2 into host cells. These results suggest that S. Typhimurium might exploit OMVs as a long-distance vehicle to deliver T3SS1 effectors into the cytoplasm of the host cell independent of bacteria-host cell interaction.

SUBMITTER: Kim SI

PROVIDER: S-EPMC6266720 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Secretion of Salmonella Pathogenicity Island 1-Encoded Type III Secretion System Effectors by Outer Membrane Vesicles in Salmonella enterica Serovar Typhimurium.

Kim Seul I SI Kim Seongok S Kim Eunsuk E Hwang Seo Yeon SY Yoon Hyunjin H

Frontiers in microbiology 20181123

Outer membrane vesicles (OMVs) are spherical membranous structures released by Gram-negative bacteria. Several bacterial pathogens utilize OMVs as vehicles for the delivery of virulence factors into host cells. Results of our previous study on proteomic analysis revealed that OMVs isolated from Salmonella enterica serovar Typhimurium had virulence effectors that are known to be translocated by Salmonella pathogenicity island 1 (SPI-1)-encoded type III secretion system (T3SS1 ...[more]

PMID: 30532744

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Project description:UnlabelledSalmonella enterica serovars cause severe diseases in humans, such as gastroenteritis and typhoid fever. The development of systemic disease is dependent on a type III secretion system (T3SS) encoded by Salmonella pathogenicity island-2 (SPI-2). Translocation of effector proteins across the Salmonella-containing vacuole, via the SPI-2 T3SS, enables bacterial replication within host cells, including macrophages. Here, we investigated the contribution of these effectors to intramacrophage replication of Salmonella enterica serovar Typhimurium using Fluorescence Dilution, a dual-fluorescence tool which allows direct measurement of bacterial replication. Of 32 strains, each carrying single mutations in genes encoding effectors, 10 (lacking sifA, sseJ, sopD2, sseG, sseF, srfH, sseL, spvD, cigR, or steD) were attenuated in replication in mouse bone marrow-derived macrophages. The replication profiles of strains combining deletions in effector genes were also investigated: a strain lacking the genes sseG, sopD2, and srfH showed an increased replication defect compared to single-mutation strains and was very similar to SPI-2 T3SS-deficient bacteria with respect to its replication defect. This strain was substantially attenuated in virulence in vivo and yet retained intracellular vacuole integrity and a functional SPI-2 T3SS. Moreover, this strain was capable of SPI-2 T3SS-mediated delivery of a model antigen for major histocompatibility complex (MHC) class I-dependent T-cell activation. This work establishes a basis for the use of a poly-effector mutant strain as an attenuated vaccine carrier for delivery of heterologous antigens directly into the cytoplasm of host cells.ImportanceLive attenuated strains of Salmonella enterica serotype Typhi have generated much interest in the search for improved vaccines against typhoid fever and as vaccine vectors for the delivery of heterologous antigens. A promising vaccine candidate is the ?aroC ?ssaV S. Typhi strain, which owes its attenuation mainly to lack of a type III secretion system (SPI-2 T3SS). The SPI-2 T3SS is important for bacterial proliferation inside macrophages, but not all of the effectors involved in this process have been identified. Here, we show that 10 effectors of the related strain S. Typhimurium contribute to intracellular replication in macrophages. Moreover, we establish that a poly-effector mutant strain of S. Typhimurium can have a severe replication defect and maintain a functional SPI-2 T3SS, which can be exploited for delivery of heterologous antigens.

Project description:Salmonella enterica serovar Typhimurium induces inflammatory diarrhea and bacterial uptake into intestinal epithelial cells using the Salmonella pathogenicity island 1 (SPI1) type III secretion system (T3SS). HilA activates transcription of the SPI1 structural components and effector proteins. Expression of hilA is activated by HilD, HilC, and RtsA, which act in a complex feed-forward regulatory loop. Many environmental signals and other regulators are integrated into this regulatory loop, primarily via HilD. After the invasion of Salmonella into host intestinal epithelial cells or during systemic replication in macrophages, the SPI T3SS is no longer required or expressed. We have shown that the two-component regulatory system PhoPQ, required for intracellular survival, represses the SPI1 T3SS mostly by controlling the transcription of hilA and hilD Here we show that PinT, one of the PhoPQ-regulated small RNAs (sRNAs), contributes to this regulation by repressing hilA and rtsA translation. PinT base pairs with both the hilA and rtsA mRNAs, resulting in translational inhibition of hilA, but also induces degradation of the rts transcript. PinT also indirectly represses expression of FliZ, a posttranslational regulator of HilD, and directly represses translation of ssrB, encoding the primary regulator of the SPI2 T3SS. Our in vivo mouse competition assays support the concept that PinT controls a series of virulence genes at the posttranscriptional level in order to adapt Salmonella from the invasion stage to intracellular survival.IMPORTANCE Salmonella is one of the most important food-borne pathogens, infecting over one million people in the United States every year. These bacteria use a needle-like device to interact with intestinal epithelial cells, leading to invasion of the cells and induction of inflammatory diarrhea. A complex regulatory network controls expression of the invasion system in response to numerous environmental signals. Here we explore the molecular mechanisms by which the small RNA PinT contributes to this regulation, facilitating inactivation of the system after invasion. PinT controls several important virulence systems in Salmonella, tuning the transition between different stages of infection.

Dataset Information

Secretion of Salmonella Pathogenicity Island 1-Encoded Type III Secretion System Effectors by Outer Membrane Vesicles in Salmonella enterica Serovar Typhimurium.

Publications

Secretion of <i>Salmonella</i> Pathogenicity Island 1-Encoded Type III Secretion System Effectors by Outer Membrane Vesicles in <i>Salmonella enterica</i> Serovar Typhimurium.

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