Antisense RNA to ahpC, an oxidative stress defence gene involved in isoniazid resistance, indicates that AhpC of Mycobacterium bovis has virulence properties

Microbiology. 1998 Oct;144 ( Pt 10):2687-95. doi: 10.1099/00221287-144-10-2687.

Abstract

Antisense RNA is a versatile tool for reducing gene expression. It was used to determine if ahpC, a gene that is involved in defence against oxidative stress and isoniazid (INH) resistance, is important for virulence of Mycobacterium bovis, a member of the Mycobacterium tuberculosis complex. Antisense RNA constructs of ahpC were made using different strength promoters in front of a reversed coding sequence of ahpC. These constructs were electroporated into a virulent wild-type M. bovis strain and a moderately virulent INH-resistant M. bovis strain that was catalase/peroxidase-negative. Down-regulation of protein synthesis occurred and this was visualized by immunoblotting. All strains containing antisense RNA were markedly less virulent than their parent strains in guinea pigs. M. bovis with an up-regulated ahpC gene was more resistant to cumene hydroperoxide than its parent strain, which had a wild-type ahpC promoter. These results agree with a model of INH resistance in which overexpression of AhpC compensates in some INH-resistant strains for loss of catalase/peroxidase by maintaining the ability to defend against oxidative stress mediated through organic peroxides. In addition, normal expression of AhpC is crucial for maintaining the virulence of wild-type M. bovis, which has normal catalase/peroxidase levels.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Antitubercular Agents / pharmacology
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism
  • Benzene Derivatives / pharmacology
  • Cell Division / drug effects
  • Drug Resistance, Microbial / genetics
  • Female
  • Gene Expression Regulation, Bacterial
  • Guinea Pigs
  • Hydrogen Peroxide / pharmacology
  • Isoniazid / pharmacology
  • Mycobacterium bovis / drug effects
  • Mycobacterium bovis / genetics
  • Mycobacterium bovis / pathogenicity*
  • Oxidative Stress / drug effects
  • Oxidative Stress / genetics
  • Oxidoreductases / genetics
  • Oxidoreductases / metabolism*
  • Peroxidases / genetics
  • Peroxidases / metabolism
  • Peroxiredoxins
  • Promoter Regions, Genetic / genetics
  • RNA, Antisense / genetics*
  • Spleen / microbiology
  • Tuberculosis / microbiology*
  • Tuberculosis / pathology
  • Virulence

Substances

  • Antitubercular Agents
  • Bacterial Proteins
  • Benzene Derivatives
  • RNA, Antisense
  • Hydrogen Peroxide
  • Oxidoreductases
  • Peroxidases
  • Peroxiredoxins
  • catalase HPI
  • cumene hydroperoxide
  • Isoniazid