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A novel antioxidant gene from Mycobacterium tuberculosis.

Ehrt S, Shiloh MU, Ruan J, Choi M, Gunzburg S, Nathan C, Xie Q, Riley LW - J. Exp. Med. (1997)

Bottom Line: Among the major antimicrobial products of macrophages are reactive intermediates of the oxidation of nitrogen (RNI) and the reduction of oxygen (ROI).Expression of noxR1 conferred upon Escherichia coli and Mycobacterium smegmatis enhanced ability to resist RNI and ROI, whether the bacteria were exposed to exogenous compounds in medium or to endogenous products in macrophages.These studies provide the first identification of an RNI resistance mechanism in mycobacteria, point to a new mechanism for resistance to ROI, and raise the possibility that inhibition of the noxR1 pathway might enhance the ability of macrophages to control tuberculosis.

View Article: PubMed Central - PubMed

Affiliation: Division of International Medicine and Infectious Disease, Department of Medicine, Cornell University Medical College, New York 10021, USA.

ABSTRACT
Among the major antimicrobial products of macrophages are reactive intermediates of the oxidation of nitrogen (RNI) and the reduction of oxygen (ROI). Selection of recombinants in acidified nitrite led to the cloning of a novel gene, noxR1, from a pathogenic clinical isolate of Mycobacterium tuberculosis. Expression of noxR1 conferred upon Escherichia coli and Mycobacterium smegmatis enhanced ability to resist RNI and ROI, whether the bacteria were exposed to exogenous compounds in medium or to endogenous products in macrophages. These studies provide the first identification of an RNI resistance mechanism in mycobacteria, point to a new mechanism for resistance to ROI, and raise the possibility that inhibition of the noxR1 pathway might enhance the ability of macrophages to control tuberculosis.

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noxR1–mediated resistance to ASN is independent of oxyR  and soxRS. Survival of the following E. coli hosts transformed with pBS  (open squares) or pNO14 (solid squares) in ASN medium: (A) JTG100  (oxyR wild type); (B) JTG101 (oxyR deficient); (C) GC4468 (soxRS wild  type); (D) DJ901 (soxRS deficient). Overnight cultures were diluted 100-fold into LB pH 5.0 containing 10 mM NaNO2 and incubated at 37°C.  Resistance was quantified by a dye reduction microplate assay (see Materials and Methods). Results are means ± SE for triplicates; some error bars  fall within the symbols.
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Figure 6: noxR1–mediated resistance to ASN is independent of oxyR and soxRS. Survival of the following E. coli hosts transformed with pBS (open squares) or pNO14 (solid squares) in ASN medium: (A) JTG100 (oxyR wild type); (B) JTG101 (oxyR deficient); (C) GC4468 (soxRS wild type); (D) DJ901 (soxRS deficient). Overnight cultures were diluted 100-fold into LB pH 5.0 containing 10 mM NaNO2 and incubated at 37°C. Resistance was quantified by a dye reduction microplate assay (see Materials and Methods). Results are means ± SE for triplicates; some error bars fall within the symbols.

Mentions: oxyR and soxRS are multigenic regulons (25), each of which is activated by and confers resistance to both ROI and RNI in E. coli (47–49). In M. tuberculosis, oxyR is disrupted and soxRS is undescribed (9, 10, 50, 51), whereas in E. coli neither regulon contains any sequence homologous to noxR1. Nonetheless, we wished to test whether noxR1 from M. tuberculosis might function in recombinant E. coli through activation of oxyR or soxRS. The genes controlled by these factors number at least 19, including those encoding superoxide dismutase, NADPH:ferredoxin oxidoreductase, fumarase, DNA repair endonuclease IV, catalase, alkylhydroperoxide reductase, and glutathione reductase. Accordingly, we transformed E. coli deficient in either of these two regulons and their corresponding wild-type strains with pNO14 or pBS vector alone. In all four hosts, noxR1 conferred resistance to ASN (Fig. 6). The degree of protection depended on the host cell type, but not on its expression of oxyR or soxRS. Thus, genes dependent upon oxyR or soxRS are dispensable for the function of noxR1, unless noxR1 can substitute for oxyR or soxRS to induce their expression.


A novel antioxidant gene from Mycobacterium tuberculosis.

Ehrt S, Shiloh MU, Ruan J, Choi M, Gunzburg S, Nathan C, Xie Q, Riley LW - J. Exp. Med. (1997)

noxR1–mediated resistance to ASN is independent of oxyR  and soxRS. Survival of the following E. coli hosts transformed with pBS  (open squares) or pNO14 (solid squares) in ASN medium: (A) JTG100  (oxyR wild type); (B) JTG101 (oxyR deficient); (C) GC4468 (soxRS wild  type); (D) DJ901 (soxRS deficient). Overnight cultures were diluted 100-fold into LB pH 5.0 containing 10 mM NaNO2 and incubated at 37°C.  Resistance was quantified by a dye reduction microplate assay (see Materials and Methods). Results are means ± SE for triplicates; some error bars  fall within the symbols.
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Related In: Results  -  Collection

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Figure 6: noxR1–mediated resistance to ASN is independent of oxyR and soxRS. Survival of the following E. coli hosts transformed with pBS (open squares) or pNO14 (solid squares) in ASN medium: (A) JTG100 (oxyR wild type); (B) JTG101 (oxyR deficient); (C) GC4468 (soxRS wild type); (D) DJ901 (soxRS deficient). Overnight cultures were diluted 100-fold into LB pH 5.0 containing 10 mM NaNO2 and incubated at 37°C. Resistance was quantified by a dye reduction microplate assay (see Materials and Methods). Results are means ± SE for triplicates; some error bars fall within the symbols.
Mentions: oxyR and soxRS are multigenic regulons (25), each of which is activated by and confers resistance to both ROI and RNI in E. coli (47–49). In M. tuberculosis, oxyR is disrupted and soxRS is undescribed (9, 10, 50, 51), whereas in E. coli neither regulon contains any sequence homologous to noxR1. Nonetheless, we wished to test whether noxR1 from M. tuberculosis might function in recombinant E. coli through activation of oxyR or soxRS. The genes controlled by these factors number at least 19, including those encoding superoxide dismutase, NADPH:ferredoxin oxidoreductase, fumarase, DNA repair endonuclease IV, catalase, alkylhydroperoxide reductase, and glutathione reductase. Accordingly, we transformed E. coli deficient in either of these two regulons and their corresponding wild-type strains with pNO14 or pBS vector alone. In all four hosts, noxR1 conferred resistance to ASN (Fig. 6). The degree of protection depended on the host cell type, but not on its expression of oxyR or soxRS. Thus, genes dependent upon oxyR or soxRS are dispensable for the function of noxR1, unless noxR1 can substitute for oxyR or soxRS to induce their expression.

Bottom Line: Among the major antimicrobial products of macrophages are reactive intermediates of the oxidation of nitrogen (RNI) and the reduction of oxygen (ROI).Expression of noxR1 conferred upon Escherichia coli and Mycobacterium smegmatis enhanced ability to resist RNI and ROI, whether the bacteria were exposed to exogenous compounds in medium or to endogenous products in macrophages.These studies provide the first identification of an RNI resistance mechanism in mycobacteria, point to a new mechanism for resistance to ROI, and raise the possibility that inhibition of the noxR1 pathway might enhance the ability of macrophages to control tuberculosis.

View Article: PubMed Central - PubMed

Affiliation: Division of International Medicine and Infectious Disease, Department of Medicine, Cornell University Medical College, New York 10021, USA.

ABSTRACT
Among the major antimicrobial products of macrophages are reactive intermediates of the oxidation of nitrogen (RNI) and the reduction of oxygen (ROI). Selection of recombinants in acidified nitrite led to the cloning of a novel gene, noxR1, from a pathogenic clinical isolate of Mycobacterium tuberculosis. Expression of noxR1 conferred upon Escherichia coli and Mycobacterium smegmatis enhanced ability to resist RNI and ROI, whether the bacteria were exposed to exogenous compounds in medium or to endogenous products in macrophages. These studies provide the first identification of an RNI resistance mechanism in mycobacteria, point to a new mechanism for resistance to ROI, and raise the possibility that inhibition of the noxR1 pathway might enhance the ability of macrophages to control tuberculosis.

Show MeSH
Related in: MedlinePlus