Dataset Information

MxiA, MxiC and IpaD Regulate Substrate Selection and Secretion Mode in the T3SS of Shigella flexneri.

ABSTRACT: Type III secretion systems (T3SSs) are central virulence devices for many Gram-negative bacterial pathogens of humans, animals & plants. Upon physical contact with eukaryotic host cells, they translocate virulence-mediating proteins, known as effectors, into them during infection. T3SSs are gated from the outside by host-cell contact and from the inside via two cytoplasmic negative regulators, MxiC and IpaD in Shigella flexneri, which together control the effector secretion hierarchy. Their absence leads to premature and increased secretion of effectors. Here, we investigated where and how these regulators act. We demonstrate that the T3SS inner membrane export apparatus protein MxiA plays a role in substrate selection. Indeed, using a genetic screen, we identified two amino acids located on the surface of MxiA's cytoplasmic region (MxiAC) which, when mutated, upregulate late effector expression and, in the case of MxiAI674V, also secretion. The cytoplasmic region of MxiA, but not MxiAN373D and MxiAI674V, interacts directly with the C-terminus of MxiC in a two-hybrid assay. Efficient T3S requires a cytoplasmic ATPase and the proton motive force (PMF), which is composed of the ?? and the ?pH. MxiA family proteins and their regulators are implicated in utilization of the PMF for protein export. However, our MxiA point mutants show similar PMF utilisation to wild-type, requiring primarily the ??. On the other hand, lack of MxiC or IpaD, renders the faster T3S seen increasingly dependent on the ?pH. Therefore, MxiA, MxiC and IpaD act together to regulate substrate selection and secretion mode in the T3SS of Shigella flexneri.

SUBMITTER: Shen DK

PROVIDER: S-EPMC4865121 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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MxiA, MxiC and IpaD Regulate Substrate Selection and Secretion Mode in the T3SS of Shigella flexneri.

Shen Da-Kang DK Blocker Ariel J AJ

PloS one 20160512 5

Type III secretion systems (T3SSs) are central virulence devices for many Gram-negative bacterial pathogens of humans, animals & plants. Upon physical contact with eukaryotic host cells, they translocate virulence-mediating proteins, known as effectors, into them during infection. T3SSs are gated from the outside by host-cell contact and from the inside via two cytoplasmic negative regulators, MxiC and IpaD in Shigella flexneri, which together control the effector secretion hierarchy. Their abse ...[more]

PMID: 27171191

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Project description:Shigella flexneri is an intracellular human pathogen that invades colonic cells and causes bloody diarrhea. S. flexneri evolved from commensal Escherichia coli, and genome comparisons reveal that S. flexneri has lost approximately 20% of its genes through the process of pathoadaptation, including a disproportionate number of genes associated with the turnover of the nucleotide-based second messenger cyclic di-GMP (c-di-GMP); however, the remaining c-di-GMP turnover enzymes are highly conserved. c-di-GMP regulates many behavioral changes in other bacteria in response to changing environmental conditions, including biofilm formation, but this signaling system has not been examined in S. flexneri. In this study, we expressed VCA0956, a constitutively active c-di-GMP synthesizing diguanylate cyclase (DGC) from Vibrio cholerae, in S. flexneri to determine if virulence phenotypes were regulated by c-di-GMP. We found that expressing VCA0956 in S. flexneri increased c-di-GMP levels, and this corresponds with increased biofilm formation and reduced acid resistance, host cell invasion, and plaque size. We examined the impact of VCA0956 expression on the S. flexneri transcriptome and found that genes related to acid resistance were repressed, and this corresponded with decreased survival to acid shock. We also found that individual S. flexneri DGC mutants exhibit reduced biofilm formation and reduced host cell invasion and plaque size, as well as increased resistance to acid shock. This study highlights the importance of c-di-GMP signaling in regulating S. flexneri virulence phenotypes. IMPORTANCE The intracellular human pathogen Shigella causes dysentery, resulting in as many as one million deaths per year. Currently, there is no approved vaccine for the prevention of shigellosis, and the incidence of antimicrobial resistance among Shigella species is on the rise. Here, we explored how the widely conserved c-di-GMP bacterial signaling system alters Shigella behaviors associated with pathogenesis. We found that expressing or removing enzymes associated with c-di-GMP synthesis results in changes in Shigella's ability to form biofilms, invade host cells, form lesions in host cell monolayers, and resist acid stress.

Dataset Information

MxiA, MxiC and IpaD Regulate Substrate Selection and Secretion Mode in the T3SS of Shigella flexneri.

Publications

MxiA, MxiC and IpaD Regulate Substrate Selection and Secretion Mode in the T3SS of Shigella flexneri.

Similar Datasets

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