Overexpression of inhA, but not kasA, confers resistance to isoniazid and ethionamide in mycobacterium smegmatis, M. bovis BCG and M. tuberculosis

Michelle H. Larsen, Catherine Vilchèze, Laurent Kremer, Gurdyal S. Besra, Linda Parsons, Max Salfinger, Leonid Heifets, Manzour H. Hazbon, David Alland, James C. Sacchettini, William R. Jacobs

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Abstract

The inhA and kasA genes of Mycobacterium tuberculosis have each been proposed to encode the primary target of the antibiotic isoniazid (INH). Previous studies investigating whether overexpressed inhA or kasA could confer resistance to INH yielded disparate results. In this work, multicopy plasmids expressing either inhA or kasA genes were transformed into M. smegmatis, M. bovis BCG and three different M. tuberculosis strains. The resulting transformants, as well as previously published M. tuberculosis strains with multicopy inhA or kasAB plasmids, were tested for their resistance to INH, ethionamide (ETH) or thiolactomycin (TLM). Mycobacteria containing inhA plasmids uniformly exhibited 20-fold or greater increased resistance to INH and 10-fold or greater increased resistance to ETH. In contrast, the kasA plasmid conferred no increased resistance to INH or ETH in any of the five strains, but it did confer resistance to thiolactomycin, a known KasA inhibitor. INH is known to increase the expression of kasA in INH-susceptible M. tuberculosis strains. Using molecular beacons, quantified inhA and kasA mRNA levels showed that increased inhA mRNA levels correlated with INH resistance, whereas kasA mRNA levels did not. In summary, analysis of strains harbouring inhA or kasA plasmids yielded the same conclusion: overexpressed inhA, but not kasA, confers INH and ETH resistance to M. smegmatis, M. bovis BCG and M. tuberculosis. Therefore, InhA is the primary target of action of INH and ETH in all three species.

Original languageEnglish (US)
Pages (from-to)453-466
Number of pages14
JournalMolecular Microbiology
Volume46
Issue number2
DOIs
StatePublished - 2002

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Ethionamide
Mycobacterium smegmatis
Isoniazid
Mycobacterium bovis
Mycobacterium tuberculosis
Tuberculosis
Plasmids
Messenger RNA
Mycobacterium
Genes
Anti-Bacterial Agents

ASJC Scopus subject areas

  • Molecular Biology
  • Microbiology

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Overexpression of inhA, but not kasA, confers resistance to isoniazid and ethionamide in mycobacterium smegmatis, M. bovis BCG and M. tuberculosis. / Larsen, Michelle H.; Vilchèze, Catherine; Kremer, Laurent; Besra, Gurdyal S.; Parsons, Linda; Salfinger, Max; Heifets, Leonid; Hazbon, Manzour H.; Alland, David; Sacchettini, James C.; Jacobs, William R.

In: Molecular Microbiology, Vol. 46, No. 2, 2002, p. 453-466.

Research output: Contribution to journalArticle

Larsen, Michelle H. ; Vilchèze, Catherine ; Kremer, Laurent ; Besra, Gurdyal S. ; Parsons, Linda ; Salfinger, Max ; Heifets, Leonid ; Hazbon, Manzour H. ; Alland, David ; Sacchettini, James C. ; Jacobs, William R. / Overexpression of inhA, but not kasA, confers resistance to isoniazid and ethionamide in mycobacterium smegmatis, M. bovis BCG and M. tuberculosis. In: Molecular Microbiology. 2002 ; Vol. 46, No. 2. pp. 453-466.
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abstract = "The inhA and kasA genes of Mycobacterium tuberculosis have each been proposed to encode the primary target of the antibiotic isoniazid (INH). Previous studies investigating whether overexpressed inhA or kasA could confer resistance to INH yielded disparate results. In this work, multicopy plasmids expressing either inhA or kasA genes were transformed into M. smegmatis, M. bovis BCG and three different M. tuberculosis strains. The resulting transformants, as well as previously published M. tuberculosis strains with multicopy inhA or kasAB plasmids, were tested for their resistance to INH, ethionamide (ETH) or thiolactomycin (TLM). Mycobacteria containing inhA plasmids uniformly exhibited 20-fold or greater increased resistance to INH and 10-fold or greater increased resistance to ETH. In contrast, the kasA plasmid conferred no increased resistance to INH or ETH in any of the five strains, but it did confer resistance to thiolactomycin, a known KasA inhibitor. INH is known to increase the expression of kasA in INH-susceptible M. tuberculosis strains. Using molecular beacons, quantified inhA and kasA mRNA levels showed that increased inhA mRNA levels correlated with INH resistance, whereas kasA mRNA levels did not. In summary, analysis of strains harbouring inhA or kasA plasmids yielded the same conclusion: overexpressed inhA, but not kasA, confers INH and ETH resistance to M. smegmatis, M. bovis BCG and M. tuberculosis. Therefore, InhA is the primary target of action of INH and ETH in all three species.",
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AU - Kremer, Laurent

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AU - Salfinger, Max

AU - Heifets, Leonid

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