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Infection and Immunity, March 2002, p. 1518-1529, Vol. 70, No. 3
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.3.1518-1529.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Identification of a Mycobacterium tuberculosis Putative Classical Nitroreductase Gene Whose Expression Is Coregulated with That of the acr Gene within Macrophages, in Standing versus Shaking Cultures, and under Low Oxygen Conditions

Anjan Purkayastha,1 Lee Ann McCue,2 and Kathleen A. McDonough1,2*

Department of Biomedical Sciences, University of Albany School of Public Health,1 Wadsworth Center, Albany, New York 12201-20022

Received 8 June 2001/ Returned for modification 29 August 2001/ Accepted 29 November 2001

Tuberculosis remains a leading killer worldwide, and new approaches for its treatment and prevention are urgently needed. This effort will benefit greatly from a better understanding of gene regulation in Mycobacterium tuberculosis, particularly with respect to this pathogen's response to its host environment. We examined the behavior of two promoters from the divergently transcribed M. tuberculosis genes acr/hspX/Rv2031c ({alpha}-crystallin homolog) and Rv2032/acg (acr-coregulated gene) by using a promoter-GFP fusion assay in Mycobacterium bovis BCG. We found that Rv2032 is a novel macrophage-induced gene whose expression is coregulated with that of acr. Relative levels of intracellular induction for both promoters were significantly affected by shallow standing versus shaking bacterial culture conditions prior to macrophage infection, and both promoters were strongly induced under low oxygen conditions. Deletion analyses showed that DNA sequences within a 43-bp region were required for expression of these promoters under all conditions. Multiple sequence alignment and database searches performed with PROBE indicated that Rv2032 is one of eight M. tuberculosis genes of previously unknown function that belong to an unusual superfamily of classical nitroreductases, which may have a role for bacteria within the host environment. These findings show that mycobacterial culture conditions can greatly influence the results and interpretation of subsequent gene regulation experiments. We propose that these differences might be exploited for dissection of the regulatory factors that affect mycobacterial gene expression within the host.

* Corresponding author. Mailing address: Wadsworth Center, 120 New Scotland Ave., PO Box 22002, Albany, NY 12201-2002. Phone: (518) 486-4253. Fax: (518) 474-3181. E-mail: kathleen.mcdonough{at}wadsworth.org.

Infection and Immunity, March 2002, p. 1518-1529, Vol. 70, No. 3
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.3.1518-1529.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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