title: Essentiality of mmpL3 and impact of its silencing on Mycobacterium tuberculosis gene expression.
abstract: MmpL3 is an inner membrane transporter of Mycobacterium tuberculosis responsible for the export of trehalose momomycolate, a precursor of the mycobacterial outer membrane component trehalose dimycolate (TDM), as well as mycolic acids bound to arabinogalactan. MmpL3 represents an emerging target for tuberculosis therapy. In this paper, we describe the construction and characterization of an mmpL3 knockdown strain of M. tuberculosis. Downregulation of mmpL3 led to a stop in bacterial division and rapid cell death, preceded by the accumulation of TDM precursors. MmpL3 was also shown to be essential for growth in monocyte-derived human macrophages. Using RNA-seq we also found that MmpL3 depletion caused up-regulation of 47 genes and down-regulation of 23 genes (at least 3-fold change and false discovery rate ≤1%). Several genes related to osmoprotection and metal homeostasis were induced, while several genes related to energy production and mycolic acids biosynthesis were repressed suggesting that inability to synthesize a correct outer membrane leads to changes in cellular permeability and a metabolic downshift.
authors.affiliation: Department of Molecular Medicine, University of Padova, Padova, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: Degiacomi
authors.affiliation: Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.
authors.forename: Andrej
authors.initials: A
authors.lastname: Benjak
authors.affiliation: Department of Biochemistry, Faculty of Natural Sciences, Comenius University in Bratislava, Bratislava, Slovakia.
authors.forename: Jan
authors.initials: J
authors.lastname: Madacki
authors.affiliation: Department of Molecular Medicine, University of Padova, Padova, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Boldrin
authors.affiliation: Department of Biology of Padova, Padova, Italy.
authors.forename: Roberta
authors.initials: R
authors.lastname: Provvedi
authors.affiliation: Department of Molecular Medicine, University of Padova, Padova, Italy.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Palù
authors.affiliation: Department of Biochemistry, Faculty of Natural Sciences, Comenius University in Bratislava, Bratislava, Slovakia.
authors.forename: Jana
authors.initials: J
authors.lastname: Kordulakova
authors.affiliation: Global Health Institute, Ecole Polytechnique Fédérale de Lausanne, Lausanne, Switzerland.
authors.forename: Stewart T
authors.initials: ST
authors.lastname: Cole
authors.affiliation: Department of Molecular Medicine, University of Padova, Padova, Italy.
authors.forename: Riccardo
authors.initials: R
authors.lastname: Manganelli
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002470:Cell Survival / Q000276:immunology; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008055:Lipids / Q000737:chemistry; D008264:Macrophages / Q000276:immunology / Q000382:microbiology; D026901:Membrane Transport Proteins / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D059467:Transcriptome; D014376:Tuberculosis / Q000276:immunology / Q000382:microbiology
chemical_list: D001426:Bacterial Proteins; D008055:Lipids; D026901:Membrane Transport Proteins; C000599913:MmpL3 protein, Mycobacterium tuberculosis