VgrG2 of type VI secretion system 2 of Vibrio parahaemolyticus induces autophagy in macrophages.

Ying Yu,Yan Zhang,Lihua Fang,Hongxia Sheng,Weihuan Fang

doi:10.3389/fmicb.2015.00168

Ying Yu, Yan Zhang + Show 3 more

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https://doi.org/10.3389/fmicb.2015.00168

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Abstract

Type VI secretion system (T6SS) is a macromolecular transenvelope machine encoded within the genomes of several proteobacteria species. Vibrio parahaemolyticus contains two putative T6SS systems, VpT6SS1 and VpT6SS2, both contributing to adherence to Caco-2 and/or HeLa cells. However, it remains unknown if these systems are involved in cellular responses. In order to exclude the effects of other virulence factors known to induce cytotoxicity or autophagy, a triple deletion mutant dTTT (with deletion of tdh, and T3SS1 and T3SS2 structural protein genes) was used as the parent strain to construct deletion mutants of T6SS genes. The mutant dTTT-ΔicmF2, but not dTTT-ΔicmF1, reduced autophagic response upon 4 h of infection of the macrophage. Further attempt was made to search for the possible effector proteins that might be responsible for direct induction of autophagy by deletion of the genes encoding Hcp2 and VgrG2, two putative translocons of T6SS2 of V. parahaemolyticus. Deletion of either hcp2 or vgrG2 did reduce the autophagic response. However, increased LC3-II lipidation was seen only in the macrophage cells transfected with pVgrG2, but not with pHcp2. Chloroquinine treatment increased accumulation of LC3-II, suggesting that VgrG2 enhanced autophagic flux. The fact that vgrG2 deletion led to reduced level of intracellular cAMP suggests a possible role of cAMP signaling in autophagic responses to the bacterium. We conclude that VgrG2 of V. parahaemolyticus induces autophagy in macrophages.

Highlights

Vibrio parahaemolyticus is one of the leading causes of human foodborne gastroenteritis in China due to consumption of raw or under-cooked seafood (Liu et al, 2004)
By examining LC3 lipidation and EGFP-LC3 punctation in macrophages infected with V. parahaemolyticus, we provide the first evidence that the VgrG2, a translocon of VpT6SS2, induces autophagy
When infection continued up to 4 h, T3SS1 was found to be involved in autophagy shown as reduced LC3-II transformation in the macrophage cells infected with vcrD1 deletion mutant (Figure 1A), as previously reported (Burdette et al, 2009a; Zhou et al, 2010)

Summary

Introduction

Vibrio parahaemolyticus is one of the leading causes of human foodborne gastroenteritis in China (about one-third of the foodborne outbreaks reported between 1991 and 2001) due to consumption of raw or under-cooked seafood (Liu et al, 2004). Studies have linked gastroenteritis to the presence of thermostable direct hemolysin (TDH), TDH-related hemolysin (TRH) and two sets of type III secretion systems (T3SS1 and T3SS2), which are able to induce general cytotoxicity or enterotoxicity to host cells (Kaper et al, 1984; Park et al, 2004; Yeung and Boor, 2004; Okada et al, 2009). Deletion of icmF associated proteins did not affect expression of the translocon proteins but prevents their translocation (Pukatzki et al, 2006; Suarez et al, 2008). Of the two sets of putative T6SS in V. parahaemolyticus (VpT6SS), we found that VpT6SS1 is present in majority of clinical isolates (90.9%), but less in environmental or food isolates (25.0%) while VpT6SS2 exists in all isolates, and both systems contribute different aspects of adherence to Caco-2 and/or HeLa cells (Yu et al, 2012)

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