Development of a repressible mycobacterial promoter system based on two transcriptional repressors.
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.
Boldrin
Francesca
F
Department of Histology, Microbiology and Medical Biotechnologies, University of Padova, Via Gabelli, 63 35100 Padova, Italy.
Casonato
Stefano
S
Dainese
Elisa
E
Sala
Claudia
C
Dhar
Neeraj
N
Palù
Giorgio
G
Riccardi
Giovanna
G
Cole
Stewart T
ST
Manganelli
Riccardo
R
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
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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