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A novel role for Stat1 in phagosome acidification and natural host resistance to intracellular infection by Leishmania major.

Späth GF, Schlesinger P, Schreiber R, Beverley SM - PLoS Pathog. (2009)

Bottom Line: Intracellular parasites of the genus Leishmania generate severe diseases in humans, which are associated with a failure of the infected host to induce a protective interferon gamma (IFNgamma)-mediated immune response.We tested the role of the JAK/STAT1 signaling pathway in Leishmania pathogenesis by utilizing knockout mice lacking the signal transducer and activator of transcription 1 (Stat1) and derived macrophages.This novel Stat1 function may have important implications to studies of other pathogens, as the acidic phagolysosomal pH plays an important role in antigen processing and the uncoating process of many viruses.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Microbiology, Washington University Medical School, St. Louis, Missouri, USA.

ABSTRACT
Intracellular parasites of the genus Leishmania generate severe diseases in humans, which are associated with a failure of the infected host to induce a protective interferon gamma (IFNgamma)-mediated immune response. We tested the role of the JAK/STAT1 signaling pathway in Leishmania pathogenesis by utilizing knockout mice lacking the signal transducer and activator of transcription 1 (Stat1) and derived macrophages. Unexpectedly, infection of Stat1-deficient macrophages in vitro with promastigotes from Leishmania major and attenuated LPG1 knockout mutants (lpg(-)) specifically lacking lipophosphoglycan (LPG) resulted in a twofold increased intracellular growth, which was independent of IFNgamma and associated with a substantial increase in phagosomal pH. Phagosomes in Stat1-/- macrophages showed normal maturation as judged by the accumulation of the lysosomal marker protein rab7, and provided normal vATPase activity, but were defective in the anion conductive pathway required for full vesicular acidification. Our results suggest a role of acidic pH in the control of intracellular Leishmania growth early during infection and identify for the first time an unexpected role of Stat1 in natural anti-microbial resistance independent from its function as IFNgamma-induced signal transducer. This novel Stat1 function may have important implications to studies of other pathogens, as the acidic phagolysosomal pH plays an important role in antigen processing and the uncoating process of many viruses.

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Phago-lysosomal fusion does not affect Leishmania major survival.(A) Phago-lysosomal fusion was quantified by labeling of parasitophorous vacuoles with FITC-dextran, previously loaded into the lysosomal compartment (Methods). (B) PEM infection was performed as described in legend of Figure 1B. Experiment was done in duplicate and mean values are shown.
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ppat-1000381-g007: Phago-lysosomal fusion does not affect Leishmania major survival.(A) Phago-lysosomal fusion was quantified by labeling of parasitophorous vacuoles with FITC-dextran, previously loaded into the lysosomal compartment (Methods). (B) PEM infection was performed as described in legend of Figure 1B. Experiment was done in duplicate and mean values are shown.

Mentions: Control and Stat1−/− PEMs previously labeled with dextran-FITC were infected synchronously with either wild-type or lpg1− Leishmania and fusogenic phagosomes were identified by florescence microscopy 3 h later as described [18]. As previously shown, wild-type parasites reside in non-fusogenic phagosomes (Figure 7A). As expected form the absence of LPG, phagosomes containing lpg1− parasites were highly fusogenic [23],[28]. Significantly, the exposure to lysosomal content in SV129 control and Stat1−/− cells had no effect on parasite survival during the first 2 days post-infection, when Leishmania-containing phagosomes are generally fully acidified (Figure 7B). In contrast, parasite numbers showed a substantial increase in Stat1−/− PEMs between day 2 and day 5 post-infection, when amastigote differentiation was completed and intracellular growth initiated. Together these data suggest that Stat1−/− PEMs show normal fusogenic properties during Leishmania infection. Additionally, the fact that LPG-deficient parasites show no difference in intracellular survival during the first 48 h in WT and Stat1−/− macrophages, despite the significant difference in their phagolysosomal pH, further supports the conclusion that killing of the LPG-deficient mutant is independent of phagosome acidification.


A novel role for Stat1 in phagosome acidification and natural host resistance to intracellular infection by Leishmania major.

Späth GF, Schlesinger P, Schreiber R, Beverley SM - PLoS Pathog. (2009)

Phago-lysosomal fusion does not affect Leishmania major survival.(A) Phago-lysosomal fusion was quantified by labeling of parasitophorous vacuoles with FITC-dextran, previously loaded into the lysosomal compartment (Methods). (B) PEM infection was performed as described in legend of Figure 1B. Experiment was done in duplicate and mean values are shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2663844&req=5

ppat-1000381-g007: Phago-lysosomal fusion does not affect Leishmania major survival.(A) Phago-lysosomal fusion was quantified by labeling of parasitophorous vacuoles with FITC-dextran, previously loaded into the lysosomal compartment (Methods). (B) PEM infection was performed as described in legend of Figure 1B. Experiment was done in duplicate and mean values are shown.
Mentions: Control and Stat1−/− PEMs previously labeled with dextran-FITC were infected synchronously with either wild-type or lpg1− Leishmania and fusogenic phagosomes were identified by florescence microscopy 3 h later as described [18]. As previously shown, wild-type parasites reside in non-fusogenic phagosomes (Figure 7A). As expected form the absence of LPG, phagosomes containing lpg1− parasites were highly fusogenic [23],[28]. Significantly, the exposure to lysosomal content in SV129 control and Stat1−/− cells had no effect on parasite survival during the first 2 days post-infection, when Leishmania-containing phagosomes are generally fully acidified (Figure 7B). In contrast, parasite numbers showed a substantial increase in Stat1−/− PEMs between day 2 and day 5 post-infection, when amastigote differentiation was completed and intracellular growth initiated. Together these data suggest that Stat1−/− PEMs show normal fusogenic properties during Leishmania infection. Additionally, the fact that LPG-deficient parasites show no difference in intracellular survival during the first 48 h in WT and Stat1−/− macrophages, despite the significant difference in their phagolysosomal pH, further supports the conclusion that killing of the LPG-deficient mutant is independent of phagosome acidification.

Bottom Line: Intracellular parasites of the genus Leishmania generate severe diseases in humans, which are associated with a failure of the infected host to induce a protective interferon gamma (IFNgamma)-mediated immune response.We tested the role of the JAK/STAT1 signaling pathway in Leishmania pathogenesis by utilizing knockout mice lacking the signal transducer and activator of transcription 1 (Stat1) and derived macrophages.This novel Stat1 function may have important implications to studies of other pathogens, as the acidic phagolysosomal pH plays an important role in antigen processing and the uncoating process of many viruses.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Microbiology, Washington University Medical School, St. Louis, Missouri, USA.

ABSTRACT
Intracellular parasites of the genus Leishmania generate severe diseases in humans, which are associated with a failure of the infected host to induce a protective interferon gamma (IFNgamma)-mediated immune response. We tested the role of the JAK/STAT1 signaling pathway in Leishmania pathogenesis by utilizing knockout mice lacking the signal transducer and activator of transcription 1 (Stat1) and derived macrophages. Unexpectedly, infection of Stat1-deficient macrophages in vitro with promastigotes from Leishmania major and attenuated LPG1 knockout mutants (lpg(-)) specifically lacking lipophosphoglycan (LPG) resulted in a twofold increased intracellular growth, which was independent of IFNgamma and associated with a substantial increase in phagosomal pH. Phagosomes in Stat1-/- macrophages showed normal maturation as judged by the accumulation of the lysosomal marker protein rab7, and provided normal vATPase activity, but were defective in the anion conductive pathway required for full vesicular acidification. Our results suggest a role of acidic pH in the control of intracellular Leishmania growth early during infection and identify for the first time an unexpected role of Stat1 in natural anti-microbial resistance independent from its function as IFNgamma-induced signal transducer. This novel Stat1 function may have important implications to studies of other pathogens, as the acidic phagolysosomal pH plays an important role in antigen processing and the uncoating process of many viruses.

Show MeSH
Related in: MedlinePlus