Heme Oxygenase-1-derived Carbon Monoxide Induces the Mycobacterium tuberculosis Dormancy Regulon

Abstract

The mechanisms that allow Mycobacterium tuberculosis (Mtb) to persist in human tissue for decades and to then abruptly cause disease are not clearly understood. Regulatory elements thought to assist Mtb to enter such a state include the heme two-component sensor kinases DosS and DosT and the cognate response regulator DosR. We have demonstrated previously that O2, nitric oxide (NO), and carbon monoxide (CO) are regulatory ligands of DosS and DosT. Here, we show that in addition to O2 and NO, CO induces the complete Mtb dormancy (Dos) regulon. Notably, we demonstrate that CO is primarily sensed through DosS to induce the Dos regulon, whereas DosT plays a less prominent role. We also show that Mtb infection of macrophage cells significantly increases the expression, protein levels, and enzymatic activity of heme oxygenase-1 (HO-1, the enzyme that produces CO), in an NO-independent manner. Furthermore, exploiting HO-1+/+ and HO-1-/- bone marrow-derived macrophages, we demonstrate that physiologically relevant levels of CO induce the Dos regulon. Finally, we demonstrate that increased HO-1 mRNA and protein levels are produced in the lungs of Mtb-infected mice. Our data suggest that during infection, O2, NO, and CO are being sensed concurrently rather than independently via DosS and DosT. We conclude that CO, a previously unrecognized host factor, is a physiologically relevant Mtb signal capable of inducing the Dos regulon, which introduces a new paradigm for understanding the molecular basis of Mtb persistence. The mechanisms that allow Mycobacterium tuberculosis (Mtb) to persist in human tissue for decades and to then abruptly cause disease are not clearly understood. Regulatory elements thought to assist Mtb to enter such a state include the heme two-component sensor kinases DosS and DosT and the cognate response regulator DosR. We have demonstrated previously that O2, nitric oxide (NO), and carbon monoxide (CO) are regulatory ligands of DosS and DosT. Here, we show that in addition to O2 and NO, CO induces the complete Mtb dormancy (Dos) regulon. Notably, we demonstrate that CO is primarily sensed through DosS to induce the Dos regulon, whereas DosT plays a less prominent role. We also show that Mtb infection of macrophage cells significantly increases the expression, protein levels, and enzymatic activity of heme oxygenase-1 (HO-1, the enzyme that produces CO), in an NO-independent manner. Furthermore, exploiting HO-1+/+ and HO-1-/- bone marrow-derived macrophages, we demonstrate that physiologically relevant levels of CO induce the Dos regulon. Finally, we demonstrate that increased HO-1 mRNA and protein levels are produced in the lungs of Mtb-infected mice. Our data suggest that during infection, O2, NO, and CO are being sensed concurrently rather than independently via DosS and DosT. We conclude that CO, a previously unrecognized host factor, is a physiologically relevant Mtb signal capable of inducing the Dos regulon, which introduces a new paradigm for understanding the molecular basis of Mtb persistence. Mycobacterium tuberculosis (Mtb) 3The abbreviations used are: Mtb, Mycobacterium tuberculosis; NO, nitric oxide; TB, tuberculosis; IFN-γ, interferon γ; iNOS, inducible nitric-oxide synthase; CO, carbon monoxide; HO-1, heme oxygenase-1; BMM, bone marrow-derived macrophage; Q-PCR, quantitative PCR. causes approximately two million deaths annually, and it is estimated that one third of the world's population is latently infected with Mtb (1Dye C. Scheele S. Dolin P. Pathania V. Raviglione M.C. J. Am. Med. Soc. 1999; 282: 677-686Crossref PubMed Scopus (2731) Google Scholar). This clinically latent state of disease can last for decades during which Mtb remains unresponsive to drug therapy. Evidence suggests roles for at least two signals, oxygen tension and nitric oxide (NO), in assisting Mtb to enter and maintain a latent state (2Cosma C.L. Sherman D.R. Ramakrishnan L. Annu. Rev. Microbiol. 2003; 57: 641-676Crossref PubMed Scopus (305) Google Scholar, 3MacMicking J.D. North R.J. LaCourse R. Mudgett J.S. Shah S.K. Nathan C.F. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 5243-5248Crossref PubMed Scopus (913) Google Scholar, 4Wayne L.G. Sohaskey C.D. Annu. Rev. Microbiol. 2001; 55: 139-163Crossref PubMed Scopus (656) Google Scholar). Using the murine model for latent tuberculosis (TB), interferon γ (IFN-γ), tumor necrosis factor-α, inducible nitric-oxide synthase (iNOS), and, thus, NO were shown to be continuously expressed and necessary to prevent reactivation of TB (5Flynn J.L. Scanga C.A. Tanaka K.E. Chan J. J. Immunol. 1998; 160: 1796-1803PubMed Google Scholar). Several studies have demonstrated a dramatic overlap between gene expression profiles of Mtb cultured under hypoxic conditions and upon treatment with NO (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar, 7Ohno H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar). These findings led to the identification of the 48-member Mtb dormancy (Dos) regulon, which is under the control of the two heme sensor kinases DosS and DosT, and the response regulator DosR (9Roberts D.M. Liao R.P. Wisedchaisri G. Hol W.G. Sherman D.R. J. Biol. Chem. 2004; 279: 23082-23087Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar). Notably, mycobacterial dosR mutants were shown to be impaired for survival during hypoxia (8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar, 10Boon C. Dick T. J. Bacteriol. 2002; 184: 6760-6767Crossref PubMed Scopus (220) Google Scholar), whereas the Mtb Beijing lineage was shown to constitutively express the Dos regulon 50-fold higher than non-Beijing strains (11Reed M.B. Gagneux S. Deriemer K. Small P.M. Barry III, C.E. J. Bacteriol. 2007; 189: 2583-2589Crossref PubMed Scopus (192) Google Scholar). Intriguingly, the Mtb dosR mutant is attenuated in guinea pigs (12Malhotra V. Sharma D. Ramanathan V.D. Shakila H. Saini D.K. Chakravorty S. Das T.K. Li Q. Silver R.F. Narayanan P.R. Tyagi J.S. FEMS Microbiol. Lett. 2004; 231: 237-245Crossref PubMed Scopus (109) Google Scholar), but not mice (13Rustad T.R. Harrell M.I. Liao R. Sherman D.R. PLoS ONE. 2008; 3: e1502Crossref PubMed Scopus (374) Google Scholar). Recently, we (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar) and others (15Ioanoviciu A. Yukl E.T. Moenne-Loccoz P. de Montellano P.R. Biochemistry. 2007; 46: 4250-4260Crossref PubMed Scopus (69) Google Scholar, 16Sardiwal S. Kendall S.L. Movahedzadeh F. Rison S.C. Stoker N.G. Djordjevic S. J. Mol. Biol. 2005; 353: 929-936Crossref PubMed Scopus (102) Google Scholar, 17Sousa E.H. Tuckerman J.R. Gonzalez G. Gilles-Gonzalez M.A. Protein Sci. 2007; 16: 1708-1719Crossref PubMed Scopus (124) Google Scholar) have shown that DosS and DosT are heme proteins that in addition to binding O2 and NO are able to bind CO. The latter findings led us to hypothesize that CO could be a third physiologically relevant ligand whose binding might induce the Dos regulon. CO is produced in vivo by heme oxygenase (HO), which is a cytoprotective enzyme that degrades heme to generate CO (18Maines M.D. Annu. Rev. Pharmacol. Toxicol. 1997; 37: 517-554Crossref PubMed Scopus Google Scholar). Notably, HO-1 through the and of (18Maines M.D. Annu. Rev. Pharmacol. Toxicol. 1997; 37: 517-554Crossref PubMed Scopus Google Scholar, J. J. Am. Soc. 2007; PubMed Scopus Google Scholar, N.G. A. Biol. Med. 2005; PubMed Scopus Google Scholar). HO-1 shown to of the of tumor necrosis and NO (18Maines M.D. Annu. Rev. Pharmacol. Toxicol. 1997; 37: 517-554Crossref PubMed Scopus Google Scholar, J. J. Am. Soc. 2007; PubMed Scopus Google Scholar, N.G. A. Biol. Med. 2005; PubMed Scopus Google Scholar). activity in the of by have for the host and the a a for CO in the of A. A. J. V. G. S. A. C.D. M. S. Med. 2007; PubMed Scopus Google Scholar). CO the of a of the of CO and was N.G. A. Biol. Med. 2005; PubMed Scopus Google Scholar). to the of HO-1, and CO, not in Mtb we hypothesize that host HO-1 CO, which induces the Mtb Dos regulon. we the of the Mtb Dos regulon in response to CO and the of DosS and DosT in signal to the Mtb Dos regulon. We also the expression of the Mtb Dos regulon in response to levels of CO we HO-1 levels in and in the lungs of Mtb-infected and mice. mice were the HO-1-/- mice and HO-1+/+ and were The were by the and at the of at and of The mice were and to be of and were used for at of Mtb mutant strains were Sherman of and is by Sherman (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar), and and are by (9Roberts D.M. Liao R.P. Wisedchaisri G. Hol W.G. Sherman D.R. J. Biol. Chem. 2004; 279: 23082-23087Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar). was to be attenuated for in the model for in dormancy (8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar, T.R. Harrell M.I. Liao R. Sherman D.R. PLoS ONE. 2008; 3: e1502Crossref PubMed Scopus (374) Google Scholar) and is not capable of inducing the Mtb Dos regulon under hypoxic conditions in response to NO (8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar). The expression of was in whereas expression of in response to hypoxia (9Roberts D.M. Liao R.P. Wisedchaisri G. Hol W.G. Sherman D.R. J. Biol. Chem. 2004; 279: 23082-23087Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar). Mtb was the were infected with of Mtb in of via Mtb was used in and used in were produced and at the for at the P. S. P. S. 2007; PubMed Scopus Google Scholar). for of the Mtb Dos in were HO-1-/- mice and HO-1+/+ were infected with a of infection of was and in to previously (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar, Mol. Biol. 1998; Google Scholar). The expression of a gene of in HO-1+/+ was to the gene of in HO-1-/- was Mtb of were HO-1-/- mice and HO-1+/+ were infected with a of infection of were by with to of were with and were by of was and via quantitative cells macrophage were in a and infected with Mtb of infection cells were and in were and and the protein of the was the protein was J. S. S. A. H. Li S. R. J. A. M.B. A. J. Exp. Med. 2007; PubMed Scopus Google Scholar). cells in tissue were infected with Mtb of infection cells were used to activity was G. J. M. K. F. G.M. J. Biol. Chem. Full Text PDF PubMed Google Scholar). infected of were with in and to These were and in were in by in with HO-1 with and with the to a were of the and the were also with and with to previously H. M. H. L. 2005; PubMed Scopus Google Scholar). are expressed and are at least of with was used for CO the of the Mtb Dos We have that the state of the DosS and DosT heme the ligands O2, NO, and CO, DosS and DosT activity (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar). the of CO the complete Dos regulon is not we to CO could induce the Mtb Dos of We used expression and the response of Mtb cells to CO. The demonstrate that CO the complete Dos regulon We the the mutant and that of the Dos regulon by CO was via DosR the of the Dos regulon by hypoxia (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar), NO H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar), and CO is by the We a of CO and expression via an (9Roberts D.M. Liao R.P. Wisedchaisri G. Hol W.G. Sherman D.R. J. Biol. Chem. 2004; 279: 23082-23087Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar) for Dos regulon We that of at CO we the for the of expression to CO and that expression not expression of to levels but increased upon treatment with a of CO not of the Mtb Dos regulon in response to CO. Q-PCR, expression was Mtb cells to of CO. Mtb DosS is the sensor of CO. Mtb, and cells were independently to CO by and expression are expressed in data demonstrate to hypoxia and NO, to of CO induces the complete Mtb Dos regulon in a and we have CO a third signal that induces the Mtb Dos regulon. Mtb DosS the of to is not in response to to hypoxia (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar), NO H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar), CO we to the of DosS and DosT in the Dos regulon. we cells to CO and that to the was impaired in to induce the Dos regulon We also the and and that was attenuated in to induce the Dos regulon with Mtb, whereas was We conclude that CO is sensed through the Mtb and that DosS is the but not the sensor of CO. of the Mtb Dos in by to CO could be a physiologically relevant ligand of Mtb DosS and DosT to the Dos regulon. We HO-1+/+ and HO-1-/- strains rather than of which have of activity J. Biol. Chem. Full Text PDF PubMed Google Scholar), H. Full Text PDF PubMed Scopus Google Scholar) of NO synthase activity D. Am. J. Mol. Biol. 2002; PubMed Scopus Google Scholar), and of and G. S. M. S. J. 2005; PubMed Scopus Google Scholar). the of Mtb infection HO-1 expression, we infected HO-1+/+ mice with Mtb and used to HO-1 Mtb-infected significantly increased HO-1 levels with whereas the of was to studies D. S. Voskuil M.I. Dolganov G. Nathan C. Schoolnik G.K. J. Exp. Med. 2003; 198: PubMed Scopus Google Scholar), we infected HO-1+/+ and HO-1-/- mice with Mtb and for and studies have shown that are to hypoxia and NO (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar, 7Ohno H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar). We a to of in HO-1+/+ cells to HO-1-/- cells a for HO-1 in the of the Dos regulon. data suggest that Mtb-infected HO-1 to CO, which is sensed by the Mtb sensor kinases DosS and DosT and then to DosR to induce the Dos regulon. of HO-1 of the NO is that during Mtb infection is via to generate NO S. M. H. 1997; PubMed Google Scholar). NO is an of HO-1 and and the demonstrate of HO-1 in response to Mtb infection we to the of the NO in HO-1 We infected macrophage cells with Mtb and to HO-1 expression We that Mtb significantly HO-1 expression with cells and the of HO-1 mRNA was with increased HO-1 protein infection with that HO-1 can in a and a in Q. S. G. R. K. A. J. T. F. J. Biol. Chem. 2007; 282: Full Text Full Text PDF PubMed Scopus Google Scholar). Furthermore, demonstrated that HO-1 expression was of Mtb infection and was for at least not the between HO-1 and the NO we also infected the which is in the of PubMed Scopus Google Scholar). Our that HO-1 expression Mtb infection is significantly increased in the at levels with that of the data is the increased HO-1 protein levels in Mtb-infected cells We also the and that HO-1 mRNA and protein levels were significantly increased and in the of the during Mtb Finally, we infected with Mtb and a in HO-1 expression upon Mtb upon of with that HO-1 mRNA and protein levels in upon infection with Mtb and and we to increased HO-1 expression an in enzymatic activity to increased CO levels, and the of the NO in enzymatic activity upon cells were infected with Mtb and used in the enzymatic activity used CO, molecular and The latter is then to by and is an of the of CO. it was that infected with Mtb higher enzymatic activity with control cells were with and cells with the demonstrate that activity was not by a a and suggest that the of HO-1, and CO, upon Mtb infection is of NO and an by which Mtb induces HO-1 in of Mtb-infected a of in and macrophage infection we the that Mtb and to levels of CO to induce the Dos regulon. CO was produced at the of infection, is the for NO J.D. North R.J. LaCourse R. Mudgett J.S. Shah S.K. Nathan C.F. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 5243-5248Crossref PubMed Scopus (913) Google Scholar), we HO-1 mRNA is in lungs Mtb infection with Mtb, HO-1 expression was significantly increased of with demonstrated that cells were with TB a of cells were in the lungs of the mice and The was in cells and we have shown that increased levels of HO-1 and CO are produced in the lungs of Mtb-infected mice. The data suggest and roles for NO and CO in Mtb and CO a ligand for DosS and DosT to the Mtb Dos regulon. The Mtb two-component to at least two physiologically relevant dormancy signals, O2 (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar) and NO H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar). we for the that CO is a third physiologically relevant signal capable of inducing the complete 48-member Mtb Dos regulon. We have also shown that CO is primarily sensed through we have demonstrated that the levels of CO produced via HO-1 were sensed by Mtb and in the of of the Dos regulon. We demonstrated that HO-1 expression and protein levels, enzymatic activity in macrophages, are significantly increased upon Mtb infection and are of the NO we have demonstrated that HO-1 is significantly increased in the lungs of Mtb-infected mice. These findings demonstrate an and previously for HO-1 and, thus, CO. Notably, the of CO, NO, and O2, to induce an of Mtb is an and a paradigm for signal in Recently, (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar) others (15Ioanoviciu A. Yukl E.T. Moenne-Loccoz P. de Montellano P.R. Biochemistry. 2007; 46: 4250-4260Crossref PubMed Scopus (69) Google Scholar, 16Sardiwal S. Kendall S.L. Movahedzadeh F. Rison S.C. Stoker N.G. Djordjevic S. J. Mol. Biol. 2005; 353: 929-936Crossref PubMed Scopus (102) Google Scholar, 17Sousa E.H. Tuckerman J.R. Gonzalez G. Gilles-Gonzalez M.A. Protein Sci. 2007; 16: 1708-1719Crossref PubMed Scopus (124) Google Scholar) shown that DosS and DosT are heme We a model that suggests that the state of Mtb DosS and DosT can be by of NO and CO of disease (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar). a a for CO in A. A. J. V. G. S. A. C.D. M. S. Med. 2007; PubMed Scopus Google Scholar), it was that CO in the to the which can with oxygen to the A. A. J. V. G. S. A. C.D. M. S. Med. 2007; PubMed Scopus Google Scholar). The overlap between model and the model for the Mtb Dos regulon (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar) one of the of the to with and not NO, the can with and we have and and demonstrated that to hypoxia and NO (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar, 7Ohno H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar), of CO induce the complete Dos regulon of CO studies have that M. M. M. and M. tuberculosis of CO G.M. Microbiol. 2003; PubMed Scopus Google Scholar) via CO studies have shown that hypoxia (6Sherman D.R. Voskuil M. Schnappinger D. Liao R. Harrell M.I. Schoolnik G.K. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 7534-7539Crossref PubMed Scopus (643) Google Scholar), NO H. Zhu G. Mohan V.P. Chu D. Kohno S. Jacobs W.R.J. Chan J. Cell. Microbiol. 2003; 5: 637-648Crossref PubMed Scopus (166) Google Scholar, 8Voskuil M.I. Schnappinger D. Visconti K.C. Harrell M.I. Dolganov G.M. Sherman D.R. Schoolnik G.K. J. Exp. Med. 2003; 198: 705-713Crossref PubMed Scopus (775) Google Scholar), CO expression of findings that CO is sensed primarily through with DosT a less prominent with data that DosS a higher for CO than DosT E.H. Tuckerman J.R. Gonzalez G. Gilles-Gonzalez M.A. Protein Sci. 2007; 16: 1708-1719Crossref PubMed Scopus (124) Google Scholar). be with an shown that DosS and DosT (9Roberts D.M. Liao R.P. Wisedchaisri G. Hol W.G. Sherman D.R. J. Biol. Chem. 2004; 279: 23082-23087Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar) in response to hypoxia in binding of O2 to DosS DosT (9Roberts D.M. Liao R.P. Wisedchaisri G. Hol W.G. Sherman D.R. J. Biol. Chem. 2004; 279: 23082-23087Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar). expression is not to O2, NO, CO to and to be constitutively DosS DosT protein levels also of the Dos regulon. the of the of to constitutively expressed is an of CO, and heme a in it was to demonstrate that not CO but also levels of CO are capable of inducing the Mtb Dos regulon upon Using HO-1+/+ and HO-1-/- we demonstrated that CO produced significantly induces expression of the Mtb Dos regulon. not CO, but also and it could be that two rather than CO induce the Dos regulon. heme G.M. Voskuil M.I. Schoolnik G.K. 2002; PubMed Scopus Google Scholar) was shown to induce the Dos regulon, and is to suggest that is capable data studies (14Kumar A. Toledo J.C. Patel R.P. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11568-11573Crossref PubMed Scopus (260) Google Scholar, A. Yukl E.T. Moenne-Loccoz P. de Montellano P.R. Biochemistry. 2007; 46: 4250-4260Crossref PubMed Scopus (69) Google Scholar, 17Sousa E.H. Tuckerman J.R. Gonzalez G. Gilles-Gonzalez M.A. Protein Sci. 2007; 16: 1708-1719Crossref PubMed Scopus (124) Google Scholar) in studies that DosS and DosT are CO and not the heme HO-1 is the of CO and is upon J. A. 2003; PubMed Scopus Google Scholar, R. R. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, R. R. J. Biol. Chem. Full Text PDF PubMed Google Scholar), whereas is the that to CO levels M.D. J. PubMed Scopus Google Scholar). CO primarily of and and a of CO C.A. 2002; PubMed Scopus Google Scholar). of and tumor necrosis which expression, were previously shown to be to prevent reactivation of Mtb in infected mice (5Flynn J.L. Scanga C.A. Tanaka K.E. Chan J. J. Immunol. 1998; 160: 1796-1803PubMed Google Scholar), we the of the NO in HO-1 We independently infected two of which in the response and demonstrated that the of HO-1 in response to Mtb infection via an that is of NO, and IFN-γ, Notably, we a of HO-1 in the lungs of Mtb-infected mice. This is not HO-1 is in response to and which are the causes of a was that response to be of the NO The of NO and CO at the of infection of NO and CO disease levels of O2, NO, and CO could maintain Mtb in a the of the and of CO (18Maines M.D. Annu. Rev. Pharmacol. Toxicol. 1997; 37: 517-554Crossref PubMed Scopus Google Scholar, J. J. Am. Soc. 2007; PubMed Scopus Google Scholar, P. M.I. 2003; PubMed Scopus Google Scholar), of CO and also NO (5Flynn J.L. Scanga C.A. Tanaka K.E. Chan J. J. Immunol. 1998; 160: 1796-1803PubMed Google Scholar) could cause findings the of and a that can be Furthermore, in the and of the for DosS and DosT E.H. Tuckerman J.R. Gonzalez G. Gilles-Gonzalez M.A. Protein Sci. 2007; 16: 1708-1719Crossref PubMed Scopus (124) Google Scholar) and host and are of a that Mtb to disease during disease Mtb also to O2 Mtb was shown to bind NO and with O2 via which might a to a in vivo carbon A. L. D. J. K.E. Lancaster Jr., J.R. Steyn A.J. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: PubMed Scopus (166) Google Scholar). the between TB and CO J. 2007; Google Scholar), the of HO-1 in human K. S. J. Immunol. PubMed Scopus Google Scholar), suggests that the findings in have and the of HO-1 and CO in Mtb remains to be the findings in significantly understanding of the Mtb Dos dormancy being by two in vivo relevant signals, hypoxia and NO, and a of TB We have shown that CO expression profiles to that of Mtb cells to hypoxia NO and that DosS is the sensor of CO. We have also shown that levels of CO induce the Mtb Dos regulon. Furthermore, we have demonstrated that HO-1 expression, and enzymatic activity significantly in response to Mtb infection and that are of the NO we have demonstrated that HO-1 is produced at the of infection, the lungs of Mtb-infected mice. These findings have for understanding the basis for Mtb persistence. We of the Steyn and for of the with