Signature Gene Expression Profiles Discriminate between Isoniazid-, Thiolactomycin-, and Triclosan-Treated
            <i>Mycobacterium tuberculosis</i>

Betts, Joanna; McLaren, Alistair; Lennon, Mark; Kelly, F. M.; Lukey, Pauline T.; Blakemore, Steve J.; Duncan, Ken

doi:10.1128/aac.47.9.2903-2913.2003

Cited by 131 publications

(124 citation statements)

References 33 publications

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“…Glycolysis, ubiquinone metabolism and redox metabolism have the maximum influence. A similar trend is observed in microarray experiments which show that the genes belonging to the TubercuList functional class of 'intermediary metabolism and respiration' are the ones that are highly regulated, along with the genes belonging to the 'conserved hypotheticals' functional class (Betts et al 2003).…”

Section: Metabolic Adjustments Inferred At the Pathway Levelsupporting

confidence: 55%

Modeling metabolic adjustment in Mycobacterium tuberculosis upon treatment with isoniazid

2010

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Complex biological systems exhibit a property of robustness at all levels of organization. Through different mechanisms, the system tries to sustain stress such as due to starvation or drug exposure. To explore whether reconfiguration of the metabolic networks is used as a means to achieve robustness, we have studied possible metabolic adjustments in Mtb upon exposure to isoniazid (INH), a front-line clinical drug. The redundancy in the genome of M. tuberculosis (Mtb) makes it an attractive system to explore if alternate routes of metabolism exist in the bacterium. While the mechanism of action of INH is well studied, its effect on the overall metabolism is not well characterized. Using flux balance analysis, inhibiting the fluxes flowing through the reactions catalyzed by Rv1484, the target of INH, significantly changes the overall flux profiles. At the pathway level, activation or inactivation of certain pathways distant from the target pathway, are seen. Metabolites such as NADPH are shown to reduce drastically, while fatty acids tend to accumulate. The overall biomass also decreases with increasing inhibition levels. Inhibition studies, pathway level clustering and comparison of the flux profiles with the gene expression data indicate the activation of folate metabolism, ubiquinone metabolism, and metabolism of certain amino acids. This analysis provides insights useful for target identification and designing strategies for combination therapy. Insights gained about the role of individual components of a system and their interactions will also provide a basis for reconstruction of whole systems through synthetic biology approaches.Keywords Genome scale metabolic networks Á Applications of flux balance analysis Á Flux profiles Á Robustness through metabolic adjustment Á Incorporating gene expression profiles Abbreviations INH Isoniazid MtbMycobacterium tuberculosis InhA NADH-dependent enoyl [acyl-carrier-protein] reductase NAD(P) Nicotinamide adenine dinucleotide (phosphate)

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Section: Metabolic Adjustments Inferred At the Pathway Levelsupporting

confidence: 55%

Modeling metabolic adjustment in Mycobacterium tuberculosis upon treatment with isoniazid

2010

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“…Moreover, the different cell morphologies prior to lysis suggest that the two mutants may have activated different cell death-inducing pathways. In this regard, it has been shown using the DNA microarray technology that although numerous genes were commonly regulated in M. tuberculosis treated with INH or TLM (inhibiting InhA and KasA, respectively), there were differences that distinguish the modes of action of these two drugs (5). Therefore, the signature gene expression profiles that discriminate between INH-and TLMtreated mycobacteria may help to identify cell death-inducing genes that are specifically activated following InhA or KasA depletion.…”

Section: Discussionmentioning

confidence: 99%

Conditional Depletion of KasA, a Key Enzyme of Mycolic Acid Biosynthesis, Leads to Mycobacterial Cell Lysis

et al. 2005

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Inhibition or inactivation of InhA, a fatty acid synthase II (FASII) enzyme, leads to mycobacterial cell lysis. To determine whether inactivation of other enzymes of the mycolic acid-synthesizing FASII complex also leads to lysis, we characterized the essentiality of two ␤-ketoacyl-acyl carrier protein synthases, KasA and KasB, in Mycobacterium smegmatis. Using specialized transduction for allelic exchange, null kasB mutants, but not kasA mutants, could be generated in Mycobacterium smegmatis, suggesting that unlike kasB, kasA is essential. To confirm the essentiality of kasA, and to detail the molecular events that occur following depletion of KasA, we developed CESTET (conditional expression specialized transduction essentiality test), a genetic tool that combines conditional gene expression and specialized transduction. Using CESTET, we were able to generate conditional null inhA and kasA mutants. We studied the effects of depletion of KasA in M. smegmatis using the former strain as a reference. Depletion of either InhA or KasA led to cell lysis, but with different biochemical and morphological events prior to lysis. While InhA depletion led to the induction of an 80-kDa complex containing both KasA and AcpM, the mycobacterial acyl carrier protein, KasA depletion did not induce the same complex. Depletion of either InhA or KasA led to inhibition of ␣ and epoxy mycolate biosynthesis and to accumulation of ␣-mycolates. Furthermore, scanning electron micrographs revealed that KasA depletion resulted in the cell surface having a "crumpled" appearance, in contrast to the blebs observed on InhA depletion. Thus, our studies support the further exploration of KasA as a target for mycobacterial-drug development.The emergence of multidrug-resistant Mycobacterium tuberculosis has been partly responsible for the global spread of tuberculosis (TB) in the past decade (51). There is an urgent need to develop novel drugs that are active against M. tuberculosis. Existing antimycobacterial agents act either by inhibiting growth or by causing cell death. The antituberculosis drug isoniazid (INH) inhibits the biosynthesis of mycolic acids, major components of cell wall lipids, resulting in cell death by lysis (43,49,50). INH inhibits InhA (1, 21, 32), an enoyl-acyl carrier protein (ACP) reductase which is part of the multienzyme fatty acid synthase II complex (FASII) that synthesizes the meromycolate precursors of mycolic acids. Although the structures of mycolic acids have been well characterized, the genetics and enzymology of mycolic acid biosynthesis have only recently begun to be elucidated (1,2,15,20,32,36,37,42,47). Mycobacteria, unlike most bacteria, have two fatty acid synthases. The eukaryote-like FASI, which is a large multidomain polypeptide containing all enzymatic functions required for de novo fatty acid synthesis, produces in a bimodal fashion saturated fatty acids of palmitate (C 16:0 ) and tetracosanoate (C 24:0 ) or C 26 (6,30). FASII elongates FASI end products to form long-chain fatty acids or meromycolates....

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“…6,7 Recently, there is growing interest in studying the pharmacogenomics of M. tuberculosis by means of microarrays. 5,[8][9][10] The functional-genomics approach will unveil genes physiologically or biochemically relevant to the drug's mode of action, genes linked to its toxic consequence, and genes related to its resistance if they are regulated at the transcriptional level. A genomic criterion based on microarray evidence may well provide a more reliable indicator for clinical drug efficacy.…”

Section: Introductionmentioning

confidence: 99%

Exploring drug action on Mycobacterium tuberculosis using affymetrix oligonucleotide genechips

2006

Tuberculosis

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SummaryDNA microarrays have rapidly emerged as an important tool for Mycobacterium tuberculosis research. While the microarray approach has generated valuable information, a recent survey has found a lack of correlation among the microarray data produced by different laboratories on related issues, raising a concern about the credibility of research findings. The Affymetrix oligonucleotide array has been shown to be more reliable for interrogating changes in gene expression than other platforms. However, this type of array system has not been applied to the pharmacogenomic study of M. tuberculosis. The goal here was to explore the strength of the Affymetrix array system for monitoring drug-induced gene expression in M. tuberculosis, compare with other related studies, and conduct cross-platform analysis. The genome-wide gene expression profiles of M. tuberculosis in response to drug treatments including INH (isoniazid) and ethionamide were obtained using the Affymetrix array system. Up-regulated or down-regulated genes were identified through bioinformatic analysis of the microarray data derived from the hybridization of RNA samples and gene probes. Based on the Affymetrix system, our method identified all drug-induced genes reported in the original reference work as well as some other genes that have not been recognized previously under the same drug treatment. For instance, the Affymetrix system revealed that Rv2524c (fas) was induced by both INH and ethionamide under the given levels of concentration, as suggested by most of the probe sets implementing this gene sequence. This finding is contradictory to previous observations that the expression of fas is not changed by INH treatment. This example illustrates that the determination of expression change for certain genes is probe-dependent, and the appropriate use of multiple probe-set representation is an advantage with the Affymetrix system. Our data also suggest that whereas the up-regulated gene expression pattern reflects the drug's mode of action, the down-regulated pattern is largely non-specific. According to our analysis, the Affymetrix array system is a reliable tool for studying the pharmacogenomics of M. tuberculosis and lends itself well in the research and development of anti-TB drugs.

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Signature Gene Expression Profiles Discriminate between Isoniazid-, Thiolactomycin-, and Triclosan-Treated Mycobacterium tuberculosis

Cited by 131 publications

References 33 publications

Modeling metabolic adjustment in Mycobacterium tuberculosis upon treatment with isoniazid

Modeling metabolic adjustment in Mycobacterium tuberculosis upon treatment with isoniazid

Conditional Depletion of KasA, a Key Enzyme of Mycolic Acid Biosynthesis, Leads to Mycobacterial Cell Lysis

Exploring drug action on Mycobacterium tuberculosis using affymetrix oligonucleotide genechips

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