title: Development of a repressible mycobacterial promoter system based on two transcriptional repressors.
abstract: Tightly regulated gene expression systems represent invaluable tools for studying gene function and for the validation of drug targets in bacteria. While several regulated bacterial promoters have been characterized, few of them have been successfully used in mycobacteria. In this article we describe the development of a novel repressible promoter system effective in both fast- and slow-growing mycobacteria based on two chromosomally encoded repressors, dependent on tetracycline (TetR) and pristinamycin (Pip), respectively. This uniqueness results in high versatility and stringency. Using this method we were able to obtain an ftsZ conditional mutant in Mycobacterium smegmatis and a fadD32 conditional mutant in Mycobacterium tuberculosis, confirming their essentiality for bacterial growth in vitro. This repressible promoter system could also be exploited to regulate gene expression during M. tuberculosis intracellular growth.
authors.affiliation: Department of Histology, Microbiology and Medical Biotechnologies, University of Padova, Via Gabelli, 63 35100 Padova, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Boldrin
authors.forename: Stefano
authors.initials: S
authors.lastname: Casonato
authors.forename: Elisa
authors.initials: E
authors.lastname: Dainese
authors.forename: Claudia
authors.initials: C
authors.lastname: Sala
authors.forename: Neeraj
authors.initials: N
authors.lastname: Dhar
authors.forename: Giorgio
authors.initials: G
authors.lastname: Palù
authors.forename: Giovanna
authors.initials: G
authors.lastname: Riccardi
authors.forename: Stewart T
authors.initials: ST
authors.lastname: Cole
authors.forename: Riccardo
authors.initials: R
authors.lastname: Manganelli
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D003598:Cytoskeletal Proteins / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D009154:Mutation; D020102:Mycobacterium smegmatis / Q000235:genetics*; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D009876:Operon; D025762:Pristinamycin / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000378:metabolism*; D013754:Tetracyclines / Q000494:pharmacology
chemical_list: D001426:Bacterial Proteins; D003598:Cytoskeletal Proteins; C071525:FtsZ protein, Bacteria; D025762:Pristinamycin; D012097:Repressor Proteins; D013754:Tetracyclines; C034811:tetracycline resistance-encoding transposon repressor protein; C016229:4-epianhydrotetracycline
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