High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.

The ability of the tubercle bacillus to arrest phagosome maturation is considered one major mechanism that allows its survival within host macrophages. To identify mycobacterial genes involved in this process, we developed a high throughput phenotypic cell-based assay enabling individual sub-cellula...

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Autores principales:	Priscille Brodin, Yannick Poquet, Florence Levillain, Isabelle Peguillet, Gerald Larrouy-Maumus, Martine Gilleron, Fanny Ewann, Thierry Christophe, Denis Fenistein, Jichan Jang, Mi-Seon Jang, Sei-Jin Park, Jean Rauzier, Jean-Philippe Carralot, Rachel Shrimpton, Auguste Genovesio, Jesus A Gonzalo-Asensio, Germain Puzo, Carlos Martin, Roland Brosch, Graham R Stewart, Brigitte Gicquel, Olivier Neyrolles
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2010
Materias:	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5
Acceso en línea:	https://doaj.org/article/3fbdc7f8e69e49e48b8fb602f57de815
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spelling	oai:doaj.org-article:3fbdc7f8e69e49e48b8fb602f57de8152021-11-18T06:01:38ZHigh content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.1553-73661553-737410.1371/journal.ppat.1001100https://doaj.org/article/3fbdc7f8e69e49e48b8fb602f57de8152010-09-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/20844580/pdf/?tool=EBIhttps://doaj.org/toc/1553-7366https://doaj.org/toc/1553-7374The ability of the tubercle bacillus to arrest phagosome maturation is considered one major mechanism that allows its survival within host macrophages. To identify mycobacterial genes involved in this process, we developed a high throughput phenotypic cell-based assay enabling individual sub-cellular analysis of over 11,000 Mycobacterium tuberculosis mutants. This very stringent assay makes use of fluorescent staining for intracellular acidic compartments, and automated confocal microscopy to quantitatively determine the intracellular localization of M. tuberculosis. We characterised the ten mutants that traffic most frequently into acidified compartments early after phagocytosis, suggesting that they had lost their ability to arrest phagosomal maturation. Molecular analysis of these mutants revealed mainly disruptions in genes involved in cell envelope biogenesis (fadD28), the ESX-1 secretion system (espL/Rv3880), molybdopterin biosynthesis (moaC1 and moaD1), as well as in genes from a novel locus, Rv1503c-Rv1506c. Most interestingly, the mutants in Rv1503c and Rv1506c were perturbed in the biosynthesis of acyltrehalose-containing glycolipids. Our results suggest that such glycolipids indeed play a critical role in the early intracellular fate of the tubercle bacillus. The unbiased approach developed here can be easily adapted for functional genomics study of intracellular pathogens, together with focused discovery of new anti-microbials.Priscille BrodinYannick PoquetFlorence LevillainIsabelle PeguilletGerald Larrouy-MaumusMartine GilleronFanny EwannThierry ChristopheDenis FenisteinJichan JangMi-Seon JangSei-Jin ParkJean RauzierJean-Philippe CarralotRachel ShrimptonAuguste GenovesioJesus A Gonzalo-AsensioGermain PuzoCarlos MartinRoland BroschGraham R StewartBrigitte GicquelOlivier NeyrollesPublic Library of Science (PLoS)articleImmunologic diseases. AllergyRC581-607Biology (General)QH301-705.5ENPLoS Pathogens, Vol 6, Iss 9, p e1001100 (2010)
institution	DOAJ
collection	DOAJ
language	EN
topic	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5
spellingShingle	Immunologic diseases. Allergy RC581-607 Biology (General) QH301-705.5 Priscille Brodin Yannick Poquet Florence Levillain Isabelle Peguillet Gerald Larrouy-Maumus Martine Gilleron Fanny Ewann Thierry Christophe Denis Fenistein Jichan Jang Mi-Seon Jang Sei-Jin Park Jean Rauzier Jean-Philippe Carralot Rachel Shrimpton Auguste Genovesio Jesus A Gonzalo-Asensio Germain Puzo Carlos Martin Roland Brosch Graham R Stewart Brigitte Gicquel Olivier Neyrolles High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
description	The ability of the tubercle bacillus to arrest phagosome maturation is considered one major mechanism that allows its survival within host macrophages. To identify mycobacterial genes involved in this process, we developed a high throughput phenotypic cell-based assay enabling individual sub-cellular analysis of over 11,000 Mycobacterium tuberculosis mutants. This very stringent assay makes use of fluorescent staining for intracellular acidic compartments, and automated confocal microscopy to quantitatively determine the intracellular localization of M. tuberculosis. We characterised the ten mutants that traffic most frequently into acidified compartments early after phagocytosis, suggesting that they had lost their ability to arrest phagosomal maturation. Molecular analysis of these mutants revealed mainly disruptions in genes involved in cell envelope biogenesis (fadD28), the ESX-1 secretion system (espL/Rv3880), molybdopterin biosynthesis (moaC1 and moaD1), as well as in genes from a novel locus, Rv1503c-Rv1506c. Most interestingly, the mutants in Rv1503c and Rv1506c were perturbed in the biosynthesis of acyltrehalose-containing glycolipids. Our results suggest that such glycolipids indeed play a critical role in the early intracellular fate of the tubercle bacillus. The unbiased approach developed here can be easily adapted for functional genomics study of intracellular pathogens, together with focused discovery of new anti-microbials.
format	article
author	Priscille Brodin Yannick Poquet Florence Levillain Isabelle Peguillet Gerald Larrouy-Maumus Martine Gilleron Fanny Ewann Thierry Christophe Denis Fenistein Jichan Jang Mi-Seon Jang Sei-Jin Park Jean Rauzier Jean-Philippe Carralot Rachel Shrimpton Auguste Genovesio Jesus A Gonzalo-Asensio Germain Puzo Carlos Martin Roland Brosch Graham R Stewart Brigitte Gicquel Olivier Neyrolles
author_facet	Priscille Brodin Yannick Poquet Florence Levillain Isabelle Peguillet Gerald Larrouy-Maumus Martine Gilleron Fanny Ewann Thierry Christophe Denis Fenistein Jichan Jang Mi-Seon Jang Sei-Jin Park Jean Rauzier Jean-Philippe Carralot Rachel Shrimpton Auguste Genovesio Jesus A Gonzalo-Asensio Germain Puzo Carlos Martin Roland Brosch Graham R Stewart Brigitte Gicquel Olivier Neyrolles
author_sort	Priscille Brodin
title	High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
title_short	High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
title_full	High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
title_fullStr	High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
title_full_unstemmed	High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
title_sort	high content phenotypic cell-based visual screen identifies mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.
publisher	Public Library of Science (PLoS)
publishDate	2010
url	https://doaj.org/article/3fbdc7f8e69e49e48b8fb602f57de815
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High content phenotypic cell-based visual screen identifies Mycobacterium tuberculosis acyltrehalose-containing glycolipids involved in phagosome remodeling.

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