A Cytotoxic Type III Secretion Effector of Vibrio parahaemolyticus Targets Vacuolar H-ATPase Subunit c and Ruptures Host Cell Lysosomes

Duke University, United States of America
PLoS Pathogens (Impact Factor: 7.56). 07/2012; 8(7):e1002803. DOI: 10.1371/journal.ppat.1002803
Source: PubMed


Vibrio parahaemolyticus is one of the human pathogenic vibrios. During the infection of mammalian cells, this pathogen exhibits cytotoxicity that is dependent on its type III secretion system (T3SS1). VepA, an effector protein secreted via the T3SS1, plays a major role in the T3SS1-dependent cytotoxicity of V. parahaemolyticus. However, the mechanism by which VepA is involved in T3SS1-dependent cytotoxicity is unknown. Here, we found that protein transfection of VepA into HeLa cells resulted in cell death, indicating that VepA alone is cytotoxic. The ectopic expression of VepA in yeast Saccharomyces cerevisiae interferes with yeast growth, indicating that VepA is also toxic in yeast. A yeast genome-wide screen identified the yeast gene VMA3 as essential for the growth inhibition of yeast by VepA. Although VMA3 encodes subunit c of the vacuolar H(+)-ATPase (V-ATPase), the toxicity of VepA was independent of the function of V-ATPases. In HeLa cells, knockdown of V-ATPase subunit c decreased VepA-mediated cytotoxicity. We also demonstrated that VepA interacted with V-ATPase subunit c, whereas a carboxyl-terminally truncated mutant of VepA (VepAΔC), which does not show toxicity, did not. During infection, lysosomal contents leaked into the cytosol, revealing that lysosomal membrane permeabilization occurred prior to cell lysis. In a cell-free system, VepA was sufficient to induce the release of cathepsin D from isolated lysosomes. Therefore, our data suggest that the bacterial effector VepA targets subunit c of V-ATPase and induces the rupture of host cell lysosomes and subsequent cell death.

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    • "The effector Vop Q (Park et al., 2004a) is necessary for the formation of autophagic vesicles in the process of V. parahaemolyticus infection (Matsuda et al., 2012). Many researchers have confirmed that the V. parahaemolyticus strain in which T3SS1 is deleted can be easily engulfed and degraded by macrophages, causing the apoptosis of the infected cells (Burdette et al., 2009; Sreelatha et al., 2013). "
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    ABSTRACT: Vibrio parahaemolyticus, a Gram-negative motile bacterium that inhabits marine and estuarine environments throughout the world, is a major food-borne pathogen that causes life-threatening diseases in humans after the consumption of raw or undercooked seafood. The global occurrence of V. parahaemolyticus accentuates the importance of investigating its virulence factors and their effects on the human host. This review describes the virulence factors of V. parahaemolyticus reported to date, including hemolysin, urease, two type III secretion systems and two type VI secretion systems, which both cause both cytotoxicity in cultured cells and enterotoxicity in animal models. We describe various types of detection methods, based on virulence factors, that are used for quantitative detection of V. parahaemolyticus in seafood. We also discuss some useful preventive measures and therapeutic strategies for the diseases mediated by V. parahaemolyticus, which can reduce, to some extent, the damage to humans and aquatic animals attributable to V. parahaemolyticus. This review extends our understanding of the pathogenic mechanisms of V. parahaemolyticus mediated by virulence factors and the diseases it causes in its human host. It should provide new insights for the diagnosis, treatment, and prevention of V. parahaemolyticus infection.
    Full-text · Article · Mar 2015 · Frontiers in Microbiology
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    • "There are currently three known effector proteins that are translocated into cells by T3SS1. These include VopS (vp1686), which AMPylates Rho-family GTPases and produces a cell rounding phenotype during infection (Casselli et al., 2008; Yarbrough et al., 2009), VopQ (VepA, vp1680), which induces PI3-kinase independent autophagy by creating gated lysosomal channels (Burdette et al., 2009; Matsuda et al., 2012; Sreelatha et al., 2013), and VPA0450, an inositol phosphatase that disrupts the integrity of the host cell membrane (Broberg et al., 2010). Although the structural components of the T3SS are often conserved between bacterial genera (Cornelis, 2006; Galán and Wolf-Watz, 2006), effector proteins are substantially more variable. "
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    ABSTRACT: Vibrio parahaemolyticus is an emerging bacterial pathogen capable of causing inflammatory gastroenteritis, wound infections, and septicemia. As a food-borne illness, infection is most frequently associated with the consumption of raw or undercooked seafood, particularly shellfish. It is the primary cause of Vibrio-associated food-borne illness in the United States and the leading cause of food-borne illness in Japan. The larger of its two chromosomes harbors a set of genes encoding type III section system 1 (T3SS1), a virulence factor present in all V. parahaemolyticus strains that is similar to the Yersinia ysc T3SS. T3SS1 translocates effector proteins into eukaryotic cells where they induce changes to cellular physiology and modulate host-pathogen interactions. T3SS1 is also responsible for cytotoxicity toward several different cultured cell lines as well as mortality in a mouse model. Herein we used RNA-seq to obtain global transcriptome patterns of V. parahaemolyticus under conditions that either induce [growth in Dulbecco's Modified Eagle Medium (DMEM) media, in trans expression of transcriptional regulator exsA] or repress T3SS1 expression (growth in LB-S media, in trans exsD expression) and during infection of HeLa cells over time. Comparative transcriptomic analysis demonstrated notable differences in the expression patterns under inducing conditions and was also used to generate an expression profile of V. parahaemolyticus during infection of HeLa cells. In addition, we identified several new genes that are associated with T3SS1 expression and may warrant further study.
    Full-text · Article · Jan 2014 · Frontiers in Cellular and Infection Microbiology
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    • "Similarly, the cytotoxicity of enniatin mycotoxins (Ivanova et al., 2012), and venom toxins from cobra (Feofanov et al., 2005) and South American rattlesnake (Hayashi et al., 2008), have been connected with LMP. Additionally, Vibrio parahaemolyticus VepA was recently identified as a new type of LMP-inducing protein (Matsuda et al., 2012). After inoculation, VepA binds to the cytoplasmic tail of the channel-forming subunit c of vacuolar H + -ATPase and triggers leakage of lysosomal hydrolases into the cytosol in a manner that depends on the subunit c. "
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    ABSTRACT: Lysosomes serve as the cellular recycling centre and are filled with numerous hydrolases that can degrade most cellular macromolecules. Lysosomal membrane permeabilization and the consequent leakage of the lysosomal content into the cytosol leads to so-called "lysosomal cell death". This form of cell death is mainly carried out by the lysosomal cathepsin proteases and can have necrotic, apoptotic or apoptosis-like features depending on the extent of the leakage and the cellular context. This article summarizes our current knowledge on lysosomal cell death with an emphasis on the upstream mechanisms that lead to lysosomal membrane permeabilization.
    Full-text · Article · May 2013 · Journal of Cell Science
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