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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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Survival of noxR1-transformed  M. smegmatis in wild-type and genetically altered macrophages. Periodate-elicited peritoneal macrophages from (A) wild-type C57BL/ 6x129/SvEv, (B) iNOS−/− C57BL/6x129/ SvEv, (C) wild-type C57BL/6, or (D)  phox91−/− C57BL/6 mice were pretreated  or not with mouse IFN-γ and infected with  M. smegmatis transformed with either  pOLYG (black bars) or pOLYG–NO14 (gray  bars). At indicated times, macrophages were  lysed and surviving bacterial CFU were determined. Values are means ± SE for triplicate macrophage cultures as a percentage of  the starting CFU, defined as the numbers of  CFUs recoverable from the cells after the 30  min uptake period; the latter averaged 2 ×  105/well, or ∼1/macrophage. (Insets) Nitrite accumulation (nmol/well) in the same  cultures of macrophages infected with M.  smegmatis–pOLYG (squares) or pOLY– NO14 (circles). Values are means ± SE; error  bars fall within the symbols. One of six similar experiments (three with IFN-γ and  three without).
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Figure 7: Survival of noxR1-transformed M. smegmatis in wild-type and genetically altered macrophages. Periodate-elicited peritoneal macrophages from (A) wild-type C57BL/ 6x129/SvEv, (B) iNOS−/− C57BL/6x129/ SvEv, (C) wild-type C57BL/6, or (D) phox91−/− C57BL/6 mice were pretreated or not with mouse IFN-γ and infected with M. smegmatis transformed with either pOLYG (black bars) or pOLYG–NO14 (gray bars). At indicated times, macrophages were lysed and surviving bacterial CFU were determined. Values are means ± SE for triplicate macrophage cultures as a percentage of the starting CFU, defined as the numbers of CFUs recoverable from the cells after the 30 min uptake period; the latter averaged 2 × 105/well, or ∼1/macrophage. (Insets) Nitrite accumulation (nmol/well) in the same cultures of macrophages infected with M. smegmatis–pOLYG (squares) or pOLY– NO14 (circles). Values are means ± SE; error bars fall within the symbols. One of six similar experiments (three with IFN-γ and three without).

Mentions: To monitor macrophage production of NO, we measured nitrite in the culture supernatants as an accumulating oxidation product. The Griess reaction was performed as previously described (33). M. smegmatis itself produced no detectable nitrite under the conditions of these experiments, as evidenced in cultures with iNOS−/− macrophages (see Fig. 7 B, inset).


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)

Survival of noxR1-transformed  M. smegmatis in wild-type and genetically altered macrophages. Periodate-elicited peritoneal macrophages from (A) wild-type C57BL/ 6x129/SvEv, (B) iNOS−/− C57BL/6x129/ SvEv, (C) wild-type C57BL/6, or (D)  phox91−/− C57BL/6 mice were pretreated  or not with mouse IFN-γ and infected with  M. smegmatis transformed with either  pOLYG (black bars) or pOLYG–NO14 (gray  bars). At indicated times, macrophages were  lysed and surviving bacterial CFU were determined. Values are means ± SE for triplicate macrophage cultures as a percentage of  the starting CFU, defined as the numbers of  CFUs recoverable from the cells after the 30  min uptake period; the latter averaged 2 ×  105/well, or ∼1/macrophage. (Insets) Nitrite accumulation (nmol/well) in the same  cultures of macrophages infected with M.  smegmatis–pOLYG (squares) or pOLY– NO14 (circles). Values are means ± SE; error  bars fall within the symbols. One of six similar experiments (three with IFN-γ and  three without).
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Related In: Results  -  Collection

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Figure 7: Survival of noxR1-transformed M. smegmatis in wild-type and genetically altered macrophages. Periodate-elicited peritoneal macrophages from (A) wild-type C57BL/ 6x129/SvEv, (B) iNOS−/− C57BL/6x129/ SvEv, (C) wild-type C57BL/6, or (D) phox91−/− C57BL/6 mice were pretreated or not with mouse IFN-γ and infected with M. smegmatis transformed with either pOLYG (black bars) or pOLYG–NO14 (gray bars). At indicated times, macrophages were lysed and surviving bacterial CFU were determined. Values are means ± SE for triplicate macrophage cultures as a percentage of the starting CFU, defined as the numbers of CFUs recoverable from the cells after the 30 min uptake period; the latter averaged 2 × 105/well, or ∼1/macrophage. (Insets) Nitrite accumulation (nmol/well) in the same cultures of macrophages infected with M. smegmatis–pOLYG (squares) or pOLY– NO14 (circles). Values are means ± SE; error bars fall within the symbols. One of six similar experiments (three with IFN-γ and three without).
Mentions: To monitor macrophage production of NO, we measured nitrite in the culture supernatants as an accumulating oxidation product. The Griess reaction was performed as previously described (33). M. smegmatis itself produced no detectable nitrite under the conditions of these experiments, as evidenced in cultures with iNOS−/− macrophages (see Fig. 7 B, inset).

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