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Pathogen recognition receptor signaling accelerates phosphorylation-dependent degradation of IFNAR1.

Qian J, Zheng H, Huangfu WC, Liu J, Carbone CJ, Leu NA, Baker DP, Fuchs SY - PLoS Pathog. (2011)

Bottom Line: However, at the same time, Type I IFNs elicit potent anti-proliferative and pro-apoptotic effects that could be detrimental for IFN-producing cells.This phosphorylation promotes IFNAR1 ubiquitination and accelerates the proteolytic turnover of this receptor leading to an attenuation of Type I IFN signaling and the protection of activated dendritic cells from the cytotoxic effects of autocrine or paracrine Type I IFN.In this paper we discuss a potential role of this mechanism in regulating the processes of innate immunity.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
An ability to sense pathogens by a number of specialized cell types including the dendritic cells plays a central role in host's defenses. Activation of these cells through the stimulation of the pathogen-recognition receptors induces the production of a number of cytokines including Type I interferons (IFNs) that mediate the diverse mechanisms of innate immunity. Type I IFNs interact with the Type I IFN receptor, composed of IFNAR1 and IFNAR2 chains, to mount the host defense responses. However, at the same time, Type I IFNs elicit potent anti-proliferative and pro-apoptotic effects that could be detrimental for IFN-producing cells. Here, we report that the activation of p38 kinase in response to pathogen-recognition receptors stimulation results in a series of phosphorylation events within the IFNAR1 chain of the Type I IFN receptor. This phosphorylation promotes IFNAR1 ubiquitination and accelerates the proteolytic turnover of this receptor leading to an attenuation of Type I IFN signaling and the protection of activated dendritic cells from the cytotoxic effects of autocrine or paracrine Type I IFN. In this paper we discuss a potential role of this mechanism in regulating the processes of innate immunity.

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HSV induces ligand/TYK2-independent phosphorylation and downregulation of IFNAR1.(A) Human KR-2 cells were infected with HSV (MOI 0.1) and harvested at 0, 24, 26, 28, and 30 h post-infection. Cell lysates were immunoprecipitated (IP) using the anti-IFNAR1 (“R1”) antibody. The phosphorylation of IFNAR1 at Ser535 and total levels of IFNAR1 were analyzed by immunoblotting (IB) using the indicated antibodies. (B) MEF (from wild type or S526A mice) were uninfected (red line) or infected (blue line) with HSV (MOI 0.1 for 32 h). Cells were subsequently incubated with the anti-mouse IFNAR1 antibody, biotin-conjugated goat-anti-mouse IgG and PE-streptavidine and then analyzed by FACS (BD Caliber). Gray area depicts a control reaction with an isotype antibody. (C) Human WT-5, KR-2 or U5A fibrosarcoma cells were either infected with HSV (MOI 0.1 for 22 h) or treated with human IFNβ (1000 IU/ml for 30 min). The analysis of IFNAR1 levels and phosphorylation in IFNAR1 immunoprecipitates was carried out using the indicated antibodies. The analyses of STAT1 phosphorylation and levels in whole cell lysates (WCL) are also shown. Exp., exposure. (D) Human WT-5, KR-2 or U5A fibrosarcoma cells were left intact or infected with HSV (MOI 0.1 for 30 h). Analyses of IFNAR1 levels in IFNAR1 immunoprecipitates and β-actin (as a loading control) in the supernatants of the immunoprecipitation reactions were carried out using indicated antibodies.
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ppat-1002065-g001: HSV induces ligand/TYK2-independent phosphorylation and downregulation of IFNAR1.(A) Human KR-2 cells were infected with HSV (MOI 0.1) and harvested at 0, 24, 26, 28, and 30 h post-infection. Cell lysates were immunoprecipitated (IP) using the anti-IFNAR1 (“R1”) antibody. The phosphorylation of IFNAR1 at Ser535 and total levels of IFNAR1 were analyzed by immunoblotting (IB) using the indicated antibodies. (B) MEF (from wild type or S526A mice) were uninfected (red line) or infected (blue line) with HSV (MOI 0.1 for 32 h). Cells were subsequently incubated with the anti-mouse IFNAR1 antibody, biotin-conjugated goat-anti-mouse IgG and PE-streptavidine and then analyzed by FACS (BD Caliber). Gray area depicts a control reaction with an isotype antibody. (C) Human WT-5, KR-2 or U5A fibrosarcoma cells were either infected with HSV (MOI 0.1 for 22 h) or treated with human IFNβ (1000 IU/ml for 30 min). The analysis of IFNAR1 levels and phosphorylation in IFNAR1 immunoprecipitates was carried out using the indicated antibodies. The analyses of STAT1 phosphorylation and levels in whole cell lysates (WCL) are also shown. Exp., exposure. (D) Human WT-5, KR-2 or U5A fibrosarcoma cells were left intact or infected with HSV (MOI 0.1 for 30 h). Analyses of IFNAR1 levels in IFNAR1 immunoprecipitates and β-actin (as a loading control) in the supernatants of the immunoprecipitation reactions were carried out using indicated antibodies.

Mentions: RNA-containing viruses (HCV and VSV) can downregulate IFNAR1 in human KR-2 cells that harbor a catalytically inactive TYK2 and that are deficient in IFNα-stimulated Ser535 phosphorylation of IFNAR1 and the degradation of this receptor chain [27], [33]. We sought to investigate whether a DNA-containing virus such as the herpes simplex virus (HSV) is also capable of such activity. We observed that the infection of KR-2 cells with HSV stimulates degron phosphorylation of endogenous human IFNAR1 and robustly downregulates the levels of this receptor (Figure 1A). A marked decrease in cell surface levels of murine IFNAR1 in response to HSV infection was also seen in mouse embryo fibroblasts (MEFs, Figure 1B).


Pathogen recognition receptor signaling accelerates phosphorylation-dependent degradation of IFNAR1.

Qian J, Zheng H, Huangfu WC, Liu J, Carbone CJ, Leu NA, Baker DP, Fuchs SY - PLoS Pathog. (2011)

HSV induces ligand/TYK2-independent phosphorylation and downregulation of IFNAR1.(A) Human KR-2 cells were infected with HSV (MOI 0.1) and harvested at 0, 24, 26, 28, and 30 h post-infection. Cell lysates were immunoprecipitated (IP) using the anti-IFNAR1 (“R1”) antibody. The phosphorylation of IFNAR1 at Ser535 and total levels of IFNAR1 were analyzed by immunoblotting (IB) using the indicated antibodies. (B) MEF (from wild type or S526A mice) were uninfected (red line) or infected (blue line) with HSV (MOI 0.1 for 32 h). Cells were subsequently incubated with the anti-mouse IFNAR1 antibody, biotin-conjugated goat-anti-mouse IgG and PE-streptavidine and then analyzed by FACS (BD Caliber). Gray area depicts a control reaction with an isotype antibody. (C) Human WT-5, KR-2 or U5A fibrosarcoma cells were either infected with HSV (MOI 0.1 for 22 h) or treated with human IFNβ (1000 IU/ml for 30 min). The analysis of IFNAR1 levels and phosphorylation in IFNAR1 immunoprecipitates was carried out using the indicated antibodies. The analyses of STAT1 phosphorylation and levels in whole cell lysates (WCL) are also shown. Exp., exposure. (D) Human WT-5, KR-2 or U5A fibrosarcoma cells were left intact or infected with HSV (MOI 0.1 for 30 h). Analyses of IFNAR1 levels in IFNAR1 immunoprecipitates and β-actin (as a loading control) in the supernatants of the immunoprecipitation reactions were carried out using indicated antibodies.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3111542&req=5

ppat-1002065-g001: HSV induces ligand/TYK2-independent phosphorylation and downregulation of IFNAR1.(A) Human KR-2 cells were infected with HSV (MOI 0.1) and harvested at 0, 24, 26, 28, and 30 h post-infection. Cell lysates were immunoprecipitated (IP) using the anti-IFNAR1 (“R1”) antibody. The phosphorylation of IFNAR1 at Ser535 and total levels of IFNAR1 were analyzed by immunoblotting (IB) using the indicated antibodies. (B) MEF (from wild type or S526A mice) were uninfected (red line) or infected (blue line) with HSV (MOI 0.1 for 32 h). Cells were subsequently incubated with the anti-mouse IFNAR1 antibody, biotin-conjugated goat-anti-mouse IgG and PE-streptavidine and then analyzed by FACS (BD Caliber). Gray area depicts a control reaction with an isotype antibody. (C) Human WT-5, KR-2 or U5A fibrosarcoma cells were either infected with HSV (MOI 0.1 for 22 h) or treated with human IFNβ (1000 IU/ml for 30 min). The analysis of IFNAR1 levels and phosphorylation in IFNAR1 immunoprecipitates was carried out using the indicated antibodies. The analyses of STAT1 phosphorylation and levels in whole cell lysates (WCL) are also shown. Exp., exposure. (D) Human WT-5, KR-2 or U5A fibrosarcoma cells were left intact or infected with HSV (MOI 0.1 for 30 h). Analyses of IFNAR1 levels in IFNAR1 immunoprecipitates and β-actin (as a loading control) in the supernatants of the immunoprecipitation reactions were carried out using indicated antibodies.
Mentions: RNA-containing viruses (HCV and VSV) can downregulate IFNAR1 in human KR-2 cells that harbor a catalytically inactive TYK2 and that are deficient in IFNα-stimulated Ser535 phosphorylation of IFNAR1 and the degradation of this receptor chain [27], [33]. We sought to investigate whether a DNA-containing virus such as the herpes simplex virus (HSV) is also capable of such activity. We observed that the infection of KR-2 cells with HSV stimulates degron phosphorylation of endogenous human IFNAR1 and robustly downregulates the levels of this receptor (Figure 1A). A marked decrease in cell surface levels of murine IFNAR1 in response to HSV infection was also seen in mouse embryo fibroblasts (MEFs, Figure 1B).

Bottom Line: However, at the same time, Type I IFNs elicit potent anti-proliferative and pro-apoptotic effects that could be detrimental for IFN-producing cells.This phosphorylation promotes IFNAR1 ubiquitination and accelerates the proteolytic turnover of this receptor leading to an attenuation of Type I IFN signaling and the protection of activated dendritic cells from the cytotoxic effects of autocrine or paracrine Type I IFN.In this paper we discuss a potential role of this mechanism in regulating the processes of innate immunity.

View Article: PubMed Central - PubMed

Affiliation: Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.

ABSTRACT
An ability to sense pathogens by a number of specialized cell types including the dendritic cells plays a central role in host's defenses. Activation of these cells through the stimulation of the pathogen-recognition receptors induces the production of a number of cytokines including Type I interferons (IFNs) that mediate the diverse mechanisms of innate immunity. Type I IFNs interact with the Type I IFN receptor, composed of IFNAR1 and IFNAR2 chains, to mount the host defense responses. However, at the same time, Type I IFNs elicit potent anti-proliferative and pro-apoptotic effects that could be detrimental for IFN-producing cells. Here, we report that the activation of p38 kinase in response to pathogen-recognition receptors stimulation results in a series of phosphorylation events within the IFNAR1 chain of the Type I IFN receptor. This phosphorylation promotes IFNAR1 ubiquitination and accelerates the proteolytic turnover of this receptor leading to an attenuation of Type I IFN signaling and the protection of activated dendritic cells from the cytotoxic effects of autocrine or paracrine Type I IFN. In this paper we discuss a potential role of this mechanism in regulating the processes of innate immunity.

Show MeSH
Related in: MedlinePlus