Mycobacterium tuberculosis SufR responds to nitric oxide via its 4Fe-4S cluster and regulates Fe-S cluster biogenesis for persistence in mice

The persistence of Mycobacterium tuberculosis (Mtb) is a major problem in managing tuberculosis (TB). Host-generated nitric oxide (NO) is perceived as one of the signals by Mtb to reprogram metabolism and respira-tion for persistence. However, the mechanisms involved in NO sensing and reorganizing Mtb's physiology are not fully understood. Since NO damages iron-sulfur (Fe-S) clusters of essential enzymes, the mechanism(s) involved in regulating Fe-S cluster biogenesis could help Mtb persist in host tissues. Here, we show that a transcription factor SufR (Rv1460) senses NO via its 4Fe-4S cluster and promotes persistence of Mtb by mobilizing the Fe-S cluster biogenesis system; suf operon (Rv1460-Rv1466). Analysis of anaerobically purified SufR by UV-visible spectroscopy, circular dichroism, and iron-sulfide estimation confirms the presence of a 4Fe-4S cluster. Atmo-spheric O-2 and H2O2 gradually degrade the 4Fe-4S cluster of SufR. Furthermore, electron paramagnetic reso-nance (EPR) analysis demonstrates that NO directly targets SufR 4Fe-4S cluster by forming a protein-bound dinitrosyl-iron-dithiol complex. DNase I footprinting, gel-shift, and in vitro transcription assays confirm that SufR directly regulates the expression of the suf operon in response to NO. Consistent with this, RNA-sequencing of Mtb Delta sufR demonstrates deregulation of the suf operon under NO stress. Strikingly, NO inflicted irreversible damage upon Fe-S clusters to exhaust respiratory and redox buffering capacity of Mtb Delta sufR. Lastly, Mtb Delta sufR failed to recover from a NO-induced non-growing state and displayed persistence defect inside immune-activated macrophages and murine lungs in a NO-dependent manner. Data suggest that SufR is a sensor of NO that sup-ports persistence by reprogramming Fe-S cluster metabolism and bioenergetics.

Automated summary

The study reveals that the transcription factor SufR senses NO and promotes the persistence of Mycobacterium tuberculosis by mobilizing the Fe-S cluster biogenesis system. NO directly damages Fe-S clusters, resulting in the inability of Mtb to recover from a non-growing state, ultimately leading to a persistence defect in immune-activated host cells and murine lungs.