Mycobacterium tuberculosis Thioredoxin Reductase Is Essential for Thiol Redox Homeostasis but Plays a Minor Role in Antioxidant Defense

Mycobacterium tuberculosis (Mtb) must cope with exogenous oxidative stress imposed by the host. Unlike other antioxidant enzymes, Mtb’s thioredoxin reductase TrxB2 has been predicted to be essential not only to fight host defenses but also for in vitro growth. However, the specific physiological role of TrxB2 and its importance for Mtb pathogenesis remain undefined. Here we show that genetic inactivation of thioredoxin reductase perturbed several growth-essential processes, including sulfur and DNA metabolism and rapidly killed and lysed Mtb. Death was due to cidal thiol-specific oxidizing stress and prevented by a disulfide reductant. In contrast, thioredoxin reductase deficiency did not significantly increase susceptibility to oxidative and nitrosative stress. In vivo targeting TrxB2 eradicated Mtb during both acute and chronic phases of mouse infection. Deliberately leaky knockdown mutants identified the specificity of TrxB2 inhibitors and showed that partial inactivation of TrxB2 increased Mtb’s susceptibility to rifampicin. These studies reveal TrxB2 as essential thiol-reducing enzyme in Mtb in vitro and during infection, establish the value of targeting TrxB2, and provide tools to accelerate the development of TrxB2 inhibitors.

Author Summary Mycobacterium tuberculosis (Mtb) antioxidant systems represent attractive targets for developing novel tuberculosis therapies. We demonstrate that targeting thioredoxin reductase TrxB2 eradicates Mtb during acute and chronic mouse infections. TrxB2 inactivation caused thiol-specific oxidizing stress, perturbed growth-essential processes and resulted in lytic death. Unexpectedly, TrxB2 deficiency did not cause increased susceptibility to oxidative and nitrosative stress. To uncover the mechanistic consequences of depleting TrxB2, or other growth essential proteins, in viable and growing bacteria, we developed a “leaky” knockdown system, with which partial TrxB2 depletion was achieved. Importantly, these leaky mutants revealed that one of two TrxB2 inhibitors kills Mtb via TrxB2 inactivation. They also demonstrated that TrxB2 depletion results in hypersusceptibility to rifampicin suggesting that a TrxB2 inhibitor will synergize with this frontline anti tuberculosis drug.