Tetracycline Controlled Expression System for Mycobacteria
- *Abstract
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Investigators at Weill Cornell Medical College have utilized the prokaryotic tetracycline-controlled gene expression system to direct gene expression in mycobacterium. Initially, the investigators developed a tet-on system of gene expression in mycobacterium in which anhydrotetracycline (atc) induces gene expression; the TetR-controlled gene expression is dependent on the concentration of atc. They used the mycobacterial gene ftsZ to show that these gene expression systems would be useful in constructing conditional knockouts as well as for the analysis of essential mycobacterial genes.
Because tetracyclines must be removed for gene silencing in tet-on gene expression systems, the investigators at WMC have also developed a tet-off system where reverse TetR mutants, that require tetracyclines as co-repressors, inhibit gene expression. The mutant TetR r1.7 inhibited lacZ expression in Mycobacterium smegmatis in the presence of atc. In addition, TetR r1.7 was also able to inhibit the secA1 gene, which decreased the amount of SecA1 protein and inhibited the growth of the conditional mutant. This system allows for the construction of conditional mycobacterium mutants in which expression of genes, such as secA1, can be silenced with the addition of atc.
This tet-off system of gene repression permits investigators to analyze the vulnerability of M. tuberculosis to inhibition of different genes involved in growth and non-replicating endurance in vitro and in vivo. In fact, the investigators at WMC used both the tet-on and tet-off systems to conditionally express the Mycobacterium tuberculosis proteasome during mouse infections. These studies revealed that the proteasome is essential for the ability of Mycobacterium tuberculosis to persist during a chronic infection in mice.
There is a need to identify drug targets that would lead to M. tuberculosis termination even when the mycobacteria are not replicating. The tet-off system-induced repression of genes that allow M. tuberculosis to avoid the host immune system, genes that induce growth of the bacterium as well as genes that allow its persistence even when it is not dividing would mimic drug inhibition of these genes and therefore permit drug target validation. This, then, could lead to the development of drugs that would inhibit M. tuberculosis survival in host cells whether it was replicating or not.
- *Licensing
- Dan-Oscar Antsonda429@cornell.edu212-746-1297
- 其他
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Ehrt S, et al (2005) Controlling gene expression in mycobacteria with anhydrotetracycline and Tet repressor.
http://nar.oxfordjournals.org/cgi/content/full/33/2/e21
Gandotra, S et al (2007) In vivo gene silencing identifies the Mycobacterium tuberculosis proteasome as essential for the bacteria to persist in mice.
http://www.ncbi.nlm.nih.gov/pubmed/18059281?ordinalpos=1&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVDocSum
Guo XV, et al (2007) Silencing essential protein secretion in Mycobacterium smegmatis by using tetracycline repressors.
http://jb.asm.org/cgi/content/full/189/13/4614?view=long&pmid=17483222
- 国家/地区
- 美国
