Lookup NU author(s): Dr Paul Dean
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Bacterial Type Three Secretion System (T3SS) effector proteins are critical determinants of infection for many animal and plant pathogens. However, monitoring of the translocation and delivery of these important virulence determinants has proved to be technically challenging. Here, we used a genetically engineered LOV (light-oxygen-voltage) sensing domain derivative to monitor the expression, translocation and localization of bacterial T3SS effectors. We found theEscherichia coli O157:H7 bacterial effector fusion Tir-LOV was functional following its translocation and localized to the host cell membrane in discrete foci demonstrating that LOV-based reporters can be used to visualize the effector translocation with minimal manipulation and interference. Further evidence for the versatility of the reporter was demonstrated by fusing LOV to the C-terminus of the Shigella flexneri effector IpaB. IpaB-LOV localized preferentially at bacterial poles before translocation. We observed the rapid translocation of IpaB-LOV in a T3SS-dependent manner into host cells, where it localized at the bacterial entry site within membrane ruffles.
Author(s): Gawthorne JA, Audry L, McQuitty C, Dean P, Christie JM, Enninga J, Roe AJ
Publication type: Article
Publication status: Published
Journal: Applied and Environmental Microbiology
Print publication date: 01/05/2016
Online publication date: 26/02/2016
Acceptance date: 26/02/2016
ISSN (print): 0099-2240
ISSN (electronic): 1098-5336
Publisher: American Society for Microbiology
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