Ammunition for the TB Wars: Tuberculosis is a major human disease of global importance resulting from infection with the air-borne pathogen Mycobacterium tuberculosis , which is becoming increasingly resistant to all available drugs. An antituberculosis benzothiazinone compound kills mycobacterium in infected cells and in mice. Makarov et al. (p. 801 ) have identified a sulfur atom and nitro residues important for benzothiazinone's activity and used genetic methods and biochemical analysis to identify its target in blocking arabinogalactan biosynthesis during cell-wall synthesis. The compound affects the same pathway as ethambutol, and thus a benzothiazinone drug has the potential to become an important part of treatment of drug-resistant disease and, possibly, replace the less effective ethambutol in the primary treatment of tuberculosis.

The proteins belonging to the Fur family are global regulators of gene expression involved in the response to several environmental stresses and to the maintenance of divalent cation homeostasis. TheMycobacterium tuberculosisgenome encodes two Fur-like proteins, FurA and a protein formerly annotated FurB. Since in this paper we show that it represents a zinc uptake regulator, we refer to it as Zur. The gene encoding Zur is found in an operon together with the gene encoding a second transcriptional regulator (Rv2358). In a previous work we demonstrated that Rv2358 is responsible for the zinc-dependent repression of the Rv2358-zuroperon, favoring the hypothesis that these genes represent key regulators of zinc homeostasis. In this study we generated azurmutant inM. tuberculosis, examined its phenotype, and characterized the Zur regulon by DNA microarray analysis. Thirty-two genes, presumably organized in 16 operons, were found to be upregulated in thezurmutant. Twenty-four of them belonged to eight putative transcriptional units preceded by a conserved 26-bp palindrome. Electrophoretic mobility shift experiments demonstrated that Zur binds to this palindrome in a zinc-dependent manner, suggesting its direct regulation of these genes. The proteins encoded by Zur-regulated genes include a group of ribosomal proteins, three putative metal transporters, the proteins belonging to early secretory antigen target 6 (ESAT-6) cluster 3, and three additional proteins belonging to the ESAT-6/culture filtrate protein 10 (CFP-10) family known to contain immunodominant epitopes in the T-cell response toM. tuberculosisinfection.

The crystal structure of the mycobacterial DprE1 reveals how the TB drug benzothiazinone BTZ043 blocks this microbial enzyme target.

The 1,5-diarylpyrrole derivative BM212 was previously shown to be active against multidrug-resistant clinical isolates and Mycobacterium tuberculosis residing within macrophages as well as against Mycobacterium avium and other atypical mycobacteria. To determine its mechanism of action, we identified the cellular target. Spontaneous Mycobacterium smegmatis , Mycobacterium bovis BCG, and M. tuberculosis H37Rv mutants that were resistant to BM212 were isolated. By the screening of genomic libraries and by whole-genome sequencing, we found that all the characterized mutants showed mutations in the mmpL3 gene, allowing us to conclude that resistance to BM212 maps to the MmpL3 protein, a member of the MmpL ( m ycobacterial m embrane p rotein, l arge) family. Susceptibility was unaffected by the efflux pump inhibitors reserpine, carbonylcyanide m -chlorophenylhydrazone, and verapamil. Uptake/efflux experiments with [ ¹⁴ C]BM212 demonstrated that resistance is not driven by the efflux of BM212. Together, these data strongly suggest that the MmpL3 protein is the cellular target of BM212.