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Neddylation is required for herpes simplex virus type I (HSV-1)-induced early phase interferon-beta production

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ABSTRACT

Type I interferons such as interferon-beta (IFN-β) play essential roles in the host innate immune response to herpes simplex virus type I (HSV-1) infection. The transcription of type I interferon genes is controlled by nuclear factor-κB (NF-κB) and interferon regulatory factor (IRF) family members including IRF3. NF-κB activation depends on the phosphorylation of inhibitor of κB (IκB), which triggers its ubiqitination and degradation. It has been reported that neddylation inhibition by a pharmacological agent MLN4924 potently suppresses lipopolysaccharide (LPS)-induced proinflammatory cytokine production with the accumulation of phosphorylated IκBα. However, the role of neddylation in type I interferon expression remains unknown. Here, we report that neddylation inhibition with MLN4924 or upon UBA3 deficiency led to accumulation of phosphorylated IκBα, impaired IκBα degradation, and impaired NF-κB nuclear translocation in the early phase of HSV-1 infection even though phosphorylation and nuclear translocation of IRF3 were not affected. The blockade of NF-κB nuclear translocation by neddylation inhibition becomes less efficient at the later time points of HSV-1 infection. Consequently, HSV-1-induced early phase IFN-β production significantly decreased upon MLN4924 treatment and UBA3 deficiency. NF-κB inhibitor JSH-23 mimicked the effects of neddylation inhibition in the early phase of HSV-1 infection. Moreover, the effects of neddylation inhibition on HSV-1-induced early phase IFN-β production diminished in the presence of NF-κB inhibitor JSH-23. Thus, neddylation contributes to HSV-1-induced early phase IFN-β production through, at least partially, promoting NF-κB activation.

No MeSH data available.


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NF-κB inhibitor JSH-23 inhibits HSV-1-induced IFN-β production. (a,b) BMMs were pretreated with various doses of JSH-23 for 30 min, followed by HSV-1 infection. 4 h later, IFN-β concentration in the supernatants was measured with ELISA (a) and cell viability was measured with ATPlite assay. LR, luminescence reading (b). (c) After BMMs were pretreated with 5 μM JSH-23 or DMSO of equal volume for 30 min, cells were infected with HSV-1 for 1 h. The subcellular localization of p65 subunit of NF-κB (red) was revealed by indirect immunofluorescence staining with a p65-specific antibody. Nuclei were counterstained for DNA by DAPI (blue). *p < 0.05, **p < 0.01.
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fig7: NF-κB inhibitor JSH-23 inhibits HSV-1-induced IFN-β production. (a,b) BMMs were pretreated with various doses of JSH-23 for 30 min, followed by HSV-1 infection. 4 h later, IFN-β concentration in the supernatants was measured with ELISA (a) and cell viability was measured with ATPlite assay. LR, luminescence reading (b). (c) After BMMs were pretreated with 5 μM JSH-23 or DMSO of equal volume for 30 min, cells were infected with HSV-1 for 1 h. The subcellular localization of p65 subunit of NF-κB (red) was revealed by indirect immunofluorescence staining with a p65-specific antibody. Nuclei were counterstained for DNA by DAPI (blue). *p < 0.05, **p < 0.01.

Mentions: To confirm the key role of NF-κB in the effects of neddylation inhibition on HSV-1-induced early phase IFN-β production, we pretreated BMMs with 5 μM JSH-23 before BMMs were subjected to HSV-1 infection for 4 h in the presence of 0.1 μM MLN4924 or upon UBA3 deficiency. ELISA revealed that JSH-23 alone showed more profound inhibition on HSV-1-induced early phase IFN-β production than MLN4924 pretreatment or UBA3 deficiency (Figure 7a,b). Because neddylation inhibition and JSH-23 pretreatment blocked NF-κB nuclear translocation in the early phase of HSV-1 infection with similar efficiency, the more profound inhibitory effect of JSH-23 on HSV-1-induced early phase IFN-β production might be attributed to its additional ability to block NF-κB transcriptional activity. The effects of neddylation inhibition on HSV-1-induced early phase IFN-β production diminishes in the presence of NF-κB inhibitor JSH-23 (Figure 7a,b). Thus, neddylation contributes to HSV-1-induced early phase IFN-β production through, at least partially, promoting NF-κB activation.


Neddylation is required for herpes simplex virus type I (HSV-1)-induced early phase interferon-beta production
NF-κB inhibitor JSH-23 inhibits HSV-1-induced IFN-β production. (a,b) BMMs were pretreated with various doses of JSH-23 for 30 min, followed by HSV-1 infection. 4 h later, IFN-β concentration in the supernatants was measured with ELISA (a) and cell viability was measured with ATPlite assay. LR, luminescence reading (b). (c) After BMMs were pretreated with 5 μM JSH-23 or DMSO of equal volume for 30 min, cells were infected with HSV-1 for 1 h. The subcellular localization of p65 subunit of NF-κB (red) was revealed by indirect immunofluorescence staining with a p65-specific antibody. Nuclei were counterstained for DNA by DAPI (blue). *p < 0.05, **p < 0.01.
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fig7: NF-κB inhibitor JSH-23 inhibits HSV-1-induced IFN-β production. (a,b) BMMs were pretreated with various doses of JSH-23 for 30 min, followed by HSV-1 infection. 4 h later, IFN-β concentration in the supernatants was measured with ELISA (a) and cell viability was measured with ATPlite assay. LR, luminescence reading (b). (c) After BMMs were pretreated with 5 μM JSH-23 or DMSO of equal volume for 30 min, cells were infected with HSV-1 for 1 h. The subcellular localization of p65 subunit of NF-κB (red) was revealed by indirect immunofluorescence staining with a p65-specific antibody. Nuclei were counterstained for DNA by DAPI (blue). *p < 0.05, **p < 0.01.
Mentions: To confirm the key role of NF-κB in the effects of neddylation inhibition on HSV-1-induced early phase IFN-β production, we pretreated BMMs with 5 μM JSH-23 before BMMs were subjected to HSV-1 infection for 4 h in the presence of 0.1 μM MLN4924 or upon UBA3 deficiency. ELISA revealed that JSH-23 alone showed more profound inhibition on HSV-1-induced early phase IFN-β production than MLN4924 pretreatment or UBA3 deficiency (Figure 7a,b). Because neddylation inhibition and JSH-23 pretreatment blocked NF-κB nuclear translocation in the early phase of HSV-1 infection with similar efficiency, the more profound inhibitory effect of JSH-23 on HSV-1-induced early phase IFN-β production might be attributed to its additional ability to block NF-κB transcriptional activity. The effects of neddylation inhibition on HSV-1-induced early phase IFN-β production diminishes in the presence of NF-κB inhibitor JSH-23 (Figure 7a,b). Thus, neddylation contributes to HSV-1-induced early phase IFN-β production through, at least partially, promoting NF-κB activation.

View Article: PubMed Central - PubMed

ABSTRACT

Type I interferons such as interferon-beta (IFN-&beta;) play essential roles in the host innate immune response to herpes simplex virus type I (HSV-1) infection. The transcription of type I interferon genes is controlled by nuclear factor-&kappa;B (NF-&kappa;B) and interferon regulatory factor (IRF) family members including IRF3. NF-&kappa;B activation depends on the phosphorylation of inhibitor of &kappa;B (I&kappa;B), which triggers its ubiqitination and degradation. It has been reported that neddylation inhibition by a pharmacological agent MLN4924 potently suppresses lipopolysaccharide (LPS)-induced proinflammatory cytokine production with the accumulation of phosphorylated I&kappa;B&alpha;. However, the role of neddylation in type I interferon expression remains unknown. Here, we report that neddylation inhibition with MLN4924 or upon UBA3 deficiency led to accumulation of phosphorylated I&kappa;B&alpha;, impaired I&kappa;B&alpha; degradation, and impaired NF-&kappa;B nuclear translocation in the early phase of HSV-1 infection even though phosphorylation and nuclear translocation of IRF3 were not affected. The blockade of NF-&kappa;B nuclear translocation by neddylation inhibition becomes less efficient at the later time points of HSV-1 infection. Consequently, HSV-1-induced early phase IFN-&beta; production significantly decreased upon MLN4924 treatment and UBA3 deficiency. NF-&kappa;B inhibitor JSH-23 mimicked the effects of neddylation inhibition in the early phase of HSV-1 infection. Moreover, the effects of neddylation inhibition on HSV-1-induced early phase IFN-&beta; production diminished in the presence of NF-&kappa;B inhibitor JSH-23. Thus, neddylation contributes to HSV-1-induced early phase IFN-&beta; production through, at least partially, promoting NF-&kappa;B activation.

No MeSH data available.


Related in: MedlinePlus