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Hepatitis C Virus Nonstructural Protein 5A Inhibits MG132-Induced Apoptosis of Hepatocytes in Line with NF-κB-Nuclear Translocation.

Jiang X, Kanda T, Wu S, Nakamoto S, Nakamura M, Sasaki R, Haga Y, Wakita T, Shirasawa H, Yokosuka O - PLoS ONE (2015)

Bottom Line: The aim of this study was to assess the effect of HCV NS5A protein in the abrogation of apoptotic cell death induced by the proteasome inhibitor MG132.Consistent with a conferred prosurvival advantage, HCV NS5A reduced MG132-induced poly(adenosine diphosphate-ribose) polymerase cleavage.HCV NS5A expression enhances phosphorylation of IκBα, liberates NF-κB for nuclear translocation and downregulates MG132-induced apoptotic pathways in human hepatocytes.

View Article: PubMed Central - PubMed

Affiliation: Departments of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, Japan.

ABSTRACT

Background: Hepatitis C virus (HCV) infection is one of the major causes of cirrhosis and hepatocellular carcinoma. HCV nonstructural protein 5A (NS5A) is an attractive antiviral target and plays an important role in HCV replication as well as hepatocarcinogenesis. The aim of this study was to assess the effect of HCV NS5A protein in the abrogation of apoptotic cell death induced by the proteasome inhibitor MG132.

Methods: Apoptotic responses to MG132 and the expression of molecules involved in NF-κB signaling pathways in human hepatocytes were investigated with or without the expression of HCV NS5A.

Results: HCV NS5A protected HepG2 cells against MG132-induced apoptosis, in line with NF-κB-nuclear translocation. A similar NF-κB-nuclear translocation was observed in Huh7 cells infected with HCV JFH1. In agreement with this, after treatment with MG132, HCV NS5A could elevate the transcription of several NF-κB target genes such as BCL2 and BCLXL to inhibit MG132-induced apoptosis in hepatocytes. HCV HCV NS5A also enhanced phosphorylation of IκBα. Consistent with a conferred prosurvival advantage, HCV NS5A reduced MG132-induced poly(adenosine diphosphate-ribose) polymerase cleavage.

Conclusions: HCV NS5A expression enhances phosphorylation of IκBα, liberates NF-κB for nuclear translocation and downregulates MG132-induced apoptotic pathways in human hepatocytes. It is possible that the disruption of proteasome-associated apoptosis plays a role in the pathogenesis of HCV infection.

No MeSH data available.


Related in: MedlinePlus

HCV NS5A interferon-sensitivity determining region (ISDR) has no impact on MG132-induced apoptosis.A, B, HepG2 cells were transfected with 0.3 μg of each vector as indicated. After 24 hours, cells were treated with 0–10 μM MG132 for 24 hours, and apoptosis was evaluated by the APOPercentage Apoptosis Assay. A, Purple-red-stained cells were identified as apoptotic cells by light microscopy (40X). B, The number of purple-red cells is shown. *P<0.05, compared to HepG2 control cells without MG132 treatment by Student's t-test.
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pone.0131973.g003: HCV NS5A interferon-sensitivity determining region (ISDR) has no impact on MG132-induced apoptosis.A, B, HepG2 cells were transfected with 0.3 μg of each vector as indicated. After 24 hours, cells were treated with 0–10 μM MG132 for 24 hours, and apoptosis was evaluated by the APOPercentage Apoptosis Assay. A, Purple-red-stained cells were identified as apoptotic cells by light microscopy (40X). B, The number of purple-red cells is shown. *P<0.05, compared to HepG2 control cells without MG132 treatment by Student's t-test.

Mentions: HCV NS5A ISDR has an association with the treatment response in interferon-including regimens against chronic HCV infection [28]. As we reported previously [14], we made HCV NS5A expression vectors, having 0 (W1 and W2), 1 (I1) or 2 (I2), or 5 amino acid changes (M1) in the HCV NS5A2209-2248 region, compared with those of HCV-J (wild type). We examined MG132-induced apoptosis after transient transfection of each HCV NS5A expression plasmid into HepG2 cells. After 24 hours, cells were treated with MG132 for 24 hours and apoptosis was evaluated as previously described (Fig 3A and 3B). HCV NS5A containing different sequences in the ISDR region have a similar effect on MG132-induced apoptosis as compared to wild type sequence. It may be possible that NS5A ISDR interacts with host protein and blocks MG132-induced apoptosis.


Hepatitis C Virus Nonstructural Protein 5A Inhibits MG132-Induced Apoptosis of Hepatocytes in Line with NF-κB-Nuclear Translocation.

Jiang X, Kanda T, Wu S, Nakamoto S, Nakamura M, Sasaki R, Haga Y, Wakita T, Shirasawa H, Yokosuka O - PLoS ONE (2015)

HCV NS5A interferon-sensitivity determining region (ISDR) has no impact on MG132-induced apoptosis.A, B, HepG2 cells were transfected with 0.3 μg of each vector as indicated. After 24 hours, cells were treated with 0–10 μM MG132 for 24 hours, and apoptosis was evaluated by the APOPercentage Apoptosis Assay. A, Purple-red-stained cells were identified as apoptotic cells by light microscopy (40X). B, The number of purple-red cells is shown. *P<0.05, compared to HepG2 control cells without MG132 treatment by Student's t-test.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4489642&req=5

pone.0131973.g003: HCV NS5A interferon-sensitivity determining region (ISDR) has no impact on MG132-induced apoptosis.A, B, HepG2 cells were transfected with 0.3 μg of each vector as indicated. After 24 hours, cells were treated with 0–10 μM MG132 for 24 hours, and apoptosis was evaluated by the APOPercentage Apoptosis Assay. A, Purple-red-stained cells were identified as apoptotic cells by light microscopy (40X). B, The number of purple-red cells is shown. *P<0.05, compared to HepG2 control cells without MG132 treatment by Student's t-test.
Mentions: HCV NS5A ISDR has an association with the treatment response in interferon-including regimens against chronic HCV infection [28]. As we reported previously [14], we made HCV NS5A expression vectors, having 0 (W1 and W2), 1 (I1) or 2 (I2), or 5 amino acid changes (M1) in the HCV NS5A2209-2248 region, compared with those of HCV-J (wild type). We examined MG132-induced apoptosis after transient transfection of each HCV NS5A expression plasmid into HepG2 cells. After 24 hours, cells were treated with MG132 for 24 hours and apoptosis was evaluated as previously described (Fig 3A and 3B). HCV NS5A containing different sequences in the ISDR region have a similar effect on MG132-induced apoptosis as compared to wild type sequence. It may be possible that NS5A ISDR interacts with host protein and blocks MG132-induced apoptosis.

Bottom Line: The aim of this study was to assess the effect of HCV NS5A protein in the abrogation of apoptotic cell death induced by the proteasome inhibitor MG132.Consistent with a conferred prosurvival advantage, HCV NS5A reduced MG132-induced poly(adenosine diphosphate-ribose) polymerase cleavage.HCV NS5A expression enhances phosphorylation of IκBα, liberates NF-κB for nuclear translocation and downregulates MG132-induced apoptotic pathways in human hepatocytes.

View Article: PubMed Central - PubMed

Affiliation: Departments of Gastroenterology and Nephrology, Chiba University, Graduate School of Medicine, Chiba, Japan.

ABSTRACT

Background: Hepatitis C virus (HCV) infection is one of the major causes of cirrhosis and hepatocellular carcinoma. HCV nonstructural protein 5A (NS5A) is an attractive antiviral target and plays an important role in HCV replication as well as hepatocarcinogenesis. The aim of this study was to assess the effect of HCV NS5A protein in the abrogation of apoptotic cell death induced by the proteasome inhibitor MG132.

Methods: Apoptotic responses to MG132 and the expression of molecules involved in NF-κB signaling pathways in human hepatocytes were investigated with or without the expression of HCV NS5A.

Results: HCV NS5A protected HepG2 cells against MG132-induced apoptosis, in line with NF-κB-nuclear translocation. A similar NF-κB-nuclear translocation was observed in Huh7 cells infected with HCV JFH1. In agreement with this, after treatment with MG132, HCV NS5A could elevate the transcription of several NF-κB target genes such as BCL2 and BCLXL to inhibit MG132-induced apoptosis in hepatocytes. HCV HCV NS5A also enhanced phosphorylation of IκBα. Consistent with a conferred prosurvival advantage, HCV NS5A reduced MG132-induced poly(adenosine diphosphate-ribose) polymerase cleavage.

Conclusions: HCV NS5A expression enhances phosphorylation of IκBα, liberates NF-κB for nuclear translocation and downregulates MG132-induced apoptotic pathways in human hepatocytes. It is possible that the disruption of proteasome-associated apoptosis plays a role in the pathogenesis of HCV infection.

No MeSH data available.


Related in: MedlinePlus