Lighting up Clostridium Difficile: reporting gene expression using fluorescent Lov domains

Buckley, A. M., Jukes, C., Candlish, D., Irvine, J. J., Spencer, J., Fagan, R. P., Roe, A. J. , Christie, J. , Fairweather, N. F. and Douce, G. R. (2016) Lighting up Clostridium Difficile: reporting gene expression using fluorescent Lov domains. Scientific Reports, 6, 23463. (doi: 10.1038/srep23463) (PMID:26996606) (PMCID:PMC4800718)

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Abstract

The uses of fluorescent reporters derived from green fluorescent protein have proved invaluable for the visualisation of biological processes in bacteria grown under aerobic conditions. However, their requirement for oxygen has limited their application in obligate anaerobes such as Clostridium difficile. Fluorescent proteins derived from Light, Oxygen or Voltage sensing (LOV) domains have been shown to bridge this limitation, but their utility as translational fusions to monitor protein expression and localisation in a strict anaerobic bacterium has not been reported. Here we demonstrate the utility of phiLOV in three species of Clostridium and its application as a marker of real-time protein translation and dynamics through genetic fusion with the cell division protein, FtsZ. Time lapse microscopy of dividing cells suggests that Z ring assembly arises through the extension of the FtsZ arc starting from one point on the circumference. Furthermore, through incorporation of phiLOV into the flagella subunit, FliC, we show the potential of bacterial LOV-based fusion proteins to be successfully exported to the extracellular environment.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Douce, Dr Gillian and Christie, Professor John and Candlish, Mrs Denise and Roe, Professor Andrew and Spencer, Dr Janice and Buckley, Dr Anthony
Authors: Buckley, A. M., Jukes, C., Candlish, D., Irvine, J. J., Spencer, J., Fagan, R. P., Roe, A. J., Christie, J., Fairweather, N. F., and Douce, G. R.
College/School:College of Medical Veterinary and Life Sciences > Institute of Infection Immunity and Inflammation
Journal Name:Scientific Reports
Publisher:Nature Publishing Group
ISSN:2045-2322
ISSN (Online):2045-2322
Published Online:21 March 2016
Copyright Holders:Copyright © 2016 The Authors
First Published:First published in Scientific Reports 6:23463
Publisher Policy:Reproduced under a creative commons license

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
492111Genetic and phenotypic characterisation of emerging virulent Clostridium difficileGillian DouceWellcome Trust (WELLCOME)086418/B/08/ZIII - BACTERIOLOGY
529641Role of the phase variable cell wall protein CwpV in C. difficile infectionGillian DouceWellcome Trust (WELLCOME)090969/B/09/ZIII - BACTERIOLOGY