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Infection mechanisms of Escherichia Coli by bacteriophage T5, studied by cryo-electron microscopy (infectphage)
Start date: Nov 19, 2010, End date: Nov 18, 2014 PROJECT  FINISHED 

The project proposed here aims at understanding the infection mechanism of bacteria by phages. I will focus my attention on a tailed phage: T5. A global molecular and cellular analysis will be carried out. Electron microscopy techniques including single particle analysis in vitro and electron tomography in situ will be used to analyze the first steps of phage infection.First of all, molecular mechanisms for the anchoring of the T5 phage onto its membrane receptor and for DNA injection will be proposed. To this end, electron microscopy experiments followed by single particles analysis of proteins and complexes of proteins from the tip of the phage tail will be performed. Moreover, it has been recently shown that both phage fixation into its receptor and DNA ejection occur preferentially at the poles or at the septum of the bacteria, for several combinations of bacteria and phages. The correlation of the localization of the receptors and the fixation sites with the organization of the cytoskeleton will be tested by a cellular analysis by electron tomography.Eventually, I intend to understand the mechanisms which lead to the recruitment of the cellular machinery of E. Coli to the purpose of expressing phagic proteins and therefore repressing the expression of the bacterial genome. A structural study in vitro will dissect the role of the different actors (RNA polymerase, phagic supposed transcription factors and phage promoteurs) for expressing the phagic proteins instead of the bacterial ones.This project will be carried out in a multi disciplinary environment. It relies on the expertise of the host lab in virology and involves cutting edge techniques in biochemistry and electron microscopy (single particle analysis and tomography) for which I will bring my knowledge. Dissection of the E.coli infection by a specific phage (T5) will make possible a better general understanding of bacterial infection by phages.

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