Mycobacterium tuberculosis DosS binds HS through its Fe heme iron to regulate the DosR dormancy regulon.

Mycobacterium tuberculosis (Mtb) senses and responds to host-derived gasotransmitters NO and CO via heme-containing sensor kinases DosS and DosT and the response regulator DosR. Hydrogen sulfide (HS) is an important signaling molecule in mammals, but its role in Mtb physiology is unclear. We have previously shown that exogenous HS can modulate expression of genes in the Dos dormancy regulon via an unknown mechanism(s). Here, we test the hypothesis that Mtb senses and responds to HS via the DosS/T/R system. Using UV-Vis and EPR spectroscopy, we show that HS binds directly to the ferric (Fe) heme of DosS (K = 5.30 μM) but not the ferrous (Fe) form. No interaction with DosT(Fe-O) was detected. We found that the binding of sulfide can slowly reduce the DosS heme iron to the ferrous form. Steered Molecular Dynamics simulations show that HS, and not the charged HS species, can enter the DosS heme pocket. We also show that HS increases DosS autokinase activity and subsequent phosphorylation of DosR, and HS-mediated increases in Dos regulon gene expression is lost in Mtb lacking DosS. Finally, we demonstrate that physiological levels of HS in macrophages can induce DosR regulon genes via DosS. Overall, these data reveal a novel mechanism whereby Mtb senses and responds to a third host gasotransmitter, HS, via DosS(Fe). These findings highlight the remarkable plasticity of DosS and establish a new paradigm for how bacteria can sense multiple gasotransmitters through a single heme sensor kinase.