A Novel Role of Listeria monocytogenes Membrane Vesicles in Inhibition of Autophagy and Cell Death.

Frontiers in Cellular and Infection Microbiology
Svitlana VdovikovaSun N Wai

Abstract

Bacterial membrane vesicle (MV) production has been mainly studied in Gram-negative species. In this study, we show that Listeria monocytogenes, a Gram-positive pathogen that causes the food-borne illness listeriosis, produces MVs both in vitro and in vivo. We found that a major virulence factor, the pore-forming hemolysin listeriolysin O (LLO), is tightly associated with the MVs, where it resides in an oxidized, inactive state. Previous studies have shown that LLO may induce cell death and autophagy. To monitor possible effects of LLO and MVs on autophagy, we performed assays for LC3 lipidation and LDH sequestration as well as analysis by confocal microscopy of HEK293 cells expressing GFP-LC3. The results revealed that MVs alone did not affect autophagy whereas they effectively abrogated autophagy induced by pure LLO or by another pore-forming toxin from Vibrio cholerae, VCC. Moreover, Listeria monocytogenes MVs significantly decreased Torin1-stimulated macroautophagy. In addition, MVs protected against necrosis of HEK293 cells caused by the lytic action of LLO. We explored the mechanisms of LLO-induced autophagy and cell death and demonstrated that the protective effect of MVs involves an inhibition of LLO-induced pore format...Continue Reading

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Methods Mentioned

BETA
density gradient centrifugation
Assay
scanning electron microscopy
confocal microscopy
lipidation
fluorescence microscopy
ELISA
atomic force microscopy
density-gradient centrifugation
flow cytometry

Software Mentioned

Image J
FACSDIVA
NanoSight
Elements Microscope Imaging
NIS
Nanoparticle Tracking Analysis
ScanAsyst
JACop
Digital micrograph
Quantity One

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