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Growth of Mycobacterium tuberculosis Biofilms
Published on: February 15, 2012
Trehalose-6-Phosphate-Mediated Toxicity Determines Essentiality of OtsB2 in Mycobacterium tuberculosis In Vitro and
Jan Korte1,2, Marina Alber2, Carolina M Trujillo3
1Institute for Pharmaceutical Biology and Biotechnology, Heinrich-Heine-University Düsseldorf, Düsseldorf, Germany.
Abstract:
Trehalose biosynthesis is considered an attractive target for the development of antimicrobials against fungal, helminthic and bacterial pathogens including Mycobacterium tuberculosis. The most common biosynthetic route involves trehalose-6-phosphate (T6P) synthase OtsA and T6P phosphatase OtsB that generate trehalose from ADP/UDP-glucose and glucose-6-phosphate. In order to assess the drug target potential of T6P phosphatase, we generated a conditional mutant of M. tuberculosis allowing the regulated gene silencing of the T6P phosphatase gene otsB2. We found that otsB2 is essential for growth of M. tuberculosis in vitro as well as for the acute infection phase in mice following aerosol infection. By contrast, otsB2 is not essential for the chronic infection phase in mice, highlighting the substantial remodelling of trehalose metabolism during infection by M. tuberculosis. Blocking OtsB2 resulted in the accumulation of its substrate T6P, which appears to be toxic, leading to the self-poisoning of cells. Accordingly, blocking T6P production in a ΔotsA mutant abrogated otsB2 essentiality. T6P accumulation elicited a global upregulation of more than 800 genes, which might result from an increase in RNA stability implied by the enhanced neutralization of toxins exhibiting ribonuclease activity. Surprisingly, overlap with the stress response caused by the accumulation of another toxic sugar phosphate molecule, maltose-1-phosphate, was minimal. A genome-wide screen for synthetic lethal interactions with otsA identified numerous genes, revealing additional potential drug targets synergistic with OtsB2 suitable for combination therapies that would minimize the emergence of resistance to OtsB2 inhibitors.
Insights
Trehalose biosynthesis is a promising antimicrobial target. Blocking trehalose-6-phosphate phosphatase (OtsB2) in Mycobacterium tuberculosis is lethal during acute infection, causing toxic substrate accumulation and revealing new drug targets.
Area of Science:
- Microbiology
- Biochemistry
- Drug Discovery
Background:
- Trehalose biosynthesis is a potential antimicrobial target against various pathogens, including Mycobacterium tuberculosis.
- The key enzymes are trehalose-6-phosphate (T6P) synthase (OtsA) and T6P phosphatase (OtsB).
Purpose of the Study:
- To evaluate T6P phosphatase (OtsB2) as a drug target in Mycobacterium tuberculosis.
- To investigate the role of OtsB2 during different infection phases and understand the consequences of its inhibition.
Main Methods:
- Generation of a conditional mutant of Mycobacterium tuberculosis for regulated silencing of the otsB2 gene.
- Assessment of bacterial growth in vitro and during acute and chronic infection phases in mice.
- Genome-wide screening for synthetic lethal interactions with otsA.
Main Results:
- otsB2 is essential for Mycobacterium tuberculosis growth in vitro and during acute infection in mice.
- otsB2 is dispensable during the chronic infection phase, indicating metabolic adaptation.
- Inhibition of OtsB2 leads to toxic T6P accumulation and global gene upregulation, suggesting a self-poisoning mechanism.
- Blocking T6P production (ΔotsA mutant) abrogated otsB2 essentiality.
- A screen identified numerous genes synthetic lethal with otsA, suggesting potential combination therapy targets.
Conclusions:
- OtsB2 is a validated drug target for acute Mycobacterium tuberculosis infections.
- Trehalose metabolism undergoes significant remodeling during infection.
- Accumulation of T6P is toxic and triggers a broad cellular response.
- Synergistic drug targets with OtsB2 inhibitors can be identified to combat antimicrobial resistance.
