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Journal of Bacteriology, July 2001, p. 4033-4039, Vol. 183, No. 13
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.13.4033-4039.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Mapping of Mycobacterium tuberculosis katG Promoters and Their Differential Expression in Infected Macrophages

Sharon Master, Thomas C. Zahrt, Jian Song, and Vojo Deretic*

Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan

Received 29 December 2000/Accepted 9 April 2001

Intracellular pathogenic bacteria, including Mycobacterium tuberculosis, frequently have multitiered defense mechanisms ensuring their survival in host phagocytic cells. One such defense determinant in M. tuberculosis is the katG gene, which encodes an enzyme with catalase, peroxidase, and peroxynitritase activities. KatG is considered to be important for protection against reactive oxygen and nitrogen intermediates produced by phagocytic cells. However, KatG also activates the front-line antituberculosis drug isoniazid, hence rendering M. tuberculosis exquisitely sensitive to this compound. In this context, katG expression represents a double-edged sword, as it is an important virulence determinant but at the same time its activity levels determine sensitivity to INH. Thus, it is important to delineate the regulation and expression of katG, as this not only can aid understanding of how M. tuberculosis survives and persists in the host but also may provide information of relevance for better management of INH therapy. Here, we report the first extensive analysis of the katG promoter activity examined both in vitro and in vivo. Using S1 nuclease protection analysis, we mapped the katG mRNA 5' ends and demonstrated that two promoters, P1furA and P1katG, control transcription of katG. The furA and katG genes are cotranscribed from P1furA. Both P1furA and P1katG promoters show induction upon challenge with hydrogen peroxide and cumene hydroperoxide. Studies carried out using the transcriptional fusions P1furA-gfp, P1katG-gfp, and P1furA-P1katG-gfp confirmed the existence of two katG promoters. In addition, we showed that both promoters are expressed in vivo during intracellular growth of virulent M. tuberculosis H37Rv. P1furA is induced early upon infection, and P1katG becomes active only upon extended growth in macrophages. These studies delineate the transcriptional organization of the furA-katG region and indicate differential regulation in vivo of the two katG promoters. These phenomena most likely reflect the differing demands at sequential stages of the infection cycle and may provide information for improved understanding of host-pathogen interactions in tuberculosis and for further optimization of INH chemotherapy.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, University of Michigan Medical School, Medical Science Building II, Ann Arbor, MI 48109. Phone: (734) 763-1580. Fax: (734) 647-6243. E-mail: deretic{at}umich.edu.

Journal of Bacteriology, July 2001, p. 4033-4039, Vol. 183, No. 13
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.13.4033-4039.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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