J Bacteriol Virol.  2018 Sep;48(3):93-101. 10.4167/jbv.2018.48.3.93.

IKKγ Facilitates the Activation of NF-κB by Hepatitis C Virus Core Protein

• Kang BY
• Lee SY
• Kim JI
• Choi HJ
• Joo WH
• Kim DW

• KMID: 2421304
• DOI: http://doi.org/10.4167/jbv.2018.48.3.93

Abstract

Hepatitis C virus (HCV) is a major cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. HCV core protein has been shown to modulate various cellular signaling pathways including the nuclear factor κB (NF-κB) pathway which is associated with inflammation, cell proliferation and apoptosis. However, there have been conflicting reports about the effect of HCV core protein on NF-κB pathway, and the mechanism by which the core protein affects NF-κB activity remains nuclear. In this study, the functional interaction of HCV core protein and IκB kinase γ (IKKγ) was investigated using the expression plasmids of core and the components of IKK complex. The data revealed that HCV core protein activates NF-κB. Also, HCV core protein up-regulated the phosphorylation and degradation of IκBα. The activating effect of HCV core protein on NF-κB was synergistically elevated by IKKγ. It was noticed that the N-terminal IKKβ binding site, C-terminal leucine zipper, and zinc finger domains of IKKγ are not necessary for its synergistic effect. HCV core protein and IKKγ appeared to activate NF-κB by up-regulating the IKKβ activity resulting in the degradation of IκBα. As expected, HCV core protein induced the expression of NF-κB-targeted pro-inflammatory genes such as iNOS, IL-1β and IL-6 in the transcription level. These results suggest that HCV core protein induces NF-κB through the interaction with IKKγ and may play a critical role in the development of inflammation and related liver diseases.

Keyword

Hepatitis C virus; Core protein; NF-κB; IKKγ; Inflammation

MeSH Terms

Apoptosis
Binding Sites
Carcinoma, Hepatocellular
Cell Proliferation
Hepacivirus*
Hepatitis C*
Hepatitis*
Hepatitis, Chronic
Inflammation
Interleukin-6
Leucine Zippers
Liver Cirrhosis
Liver Diseases
Phosphorylation
Phosphotransferases
Plasmids
Zinc Fingers
Interleukin-6
Phosphotransferases

Figure

• 

  Figure 1. Activation of NF-κB by HCV core and IKKγ. (A) COS-7 cells were co-transfected with pNL-κB-RE reporter and each expression plasmid of IKKβ, NIK, HCV core, or IKKγ. Luciferase assays were performed 45 hr after transfection. (mean ± SD, n=3, ∗ p<0.05, ∗∗ p<0.001). (B) COS-7 cells were co-transfected with indicated expression plasmids. Cells were harvested at 45 hr post-transfection and the cytoplasmic extracts were analyzed by Western blotting.

• 

  Figure 2. The structure of wild-type and N-terminal, C-terminal deletion mutants of mouse IKKγ. IKKβ binding domain at N-terminal and leucine zipper domain (LZ), zinc finger domain (ZF) at C-terminal are shown. The numbers indicate the number of amino acids.

• 

  Figure 3. Activation of NF-κB by HCV core and IKKγ mutants. (A) COS-7 cells were co-transfected with pNL-κB-RE reporter and each expression plasmid of IKKβ, NIK, HCV core or IKKγ mutant. Luciferase assay was performed at 45 hr post-transfection. (mean ± SD, n=3, ∗ p<0.05, ∗∗ p<0.001). (B) COS-7 cells were co-transfected with expression plasmids as indicated. Cells were harvested at 45 hr post-transfection and the cytoplasmic extracts were analyzed by western blotting.

• 

  Figure 4. Effect of HCV core and IKKγ on the activation of NF-κB. COS-7 cells were co-transfected with pNL-κB-RE reporter and each expression plasmid of IKKβ, IKKβ-EE, HCV core, wild-type IKKγ, IKKγ-Δ N, or Δ C. Luciferase assay was performed at 45 hr post-transfection (mean ± SD, n=3, ∗ p<0.05, ∗∗ p<0.001).

• 

  Figure 5. Effect of HCV core on the expression of inflammatory mediators. Raw 264.7 cells were transfected with empty vector (C) or HCV core-expression plasmid (core). Cells were harvested at 45 hr post-transfection and analyzed by RT-PCR. GAPDH was used as the internal control.

• 

  Figure 6. Effect of HCV core on the phosphorylation of signaling molecules in the proinflammatory pathway. Raw 264.7 cells were transfected with empty vector (C) or HCV core expression plasmid. The whole cell extracts were prepared at 45 hr post-transfection and analyzed by western blotting.

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