PLoS One. 2014 May 1;9(5):e95627. doi: 10.1371/journal.pone.0095627.

Hepatitis C virus core protein inhibits interferon production by a human plasmacytoid dendritic cell line and dysregulates interferon regulatory factor-7 and signal transducer and activator of transcription (STAT) 1 protein expression.

Amy EL Stone1,2, Angela Mitchell1,2, Jessica Brownell3, Daniel J Miklin2, Lucy Golden-Mason1,2, Stephen J Polyak3,4, Michael J Gale Jr5. and Hugo R Rosen1,2,6

1 Integrated Department in Immunology, University of Colorado Denver and National Jewish Health, Denver, Colorado, United States of America; Division of Gastroenterology & Hepatology, Hepatitis C Center, Department of Medicine, University of Colorado Denver, Aurora, Colorado, United States of America.
2 Department of Global Health, University of Washington, Seattle, Washington, United States of America.
3 Division of Gastroenterology & Hepatology, Hepatitis C Center, Department of Medicine, University of Colorado Denver, Aurora, Colorado, United States of America.
4 Department of Global Health, University of Washington, Seattle, Washington, United States of America; Department of Laboratory Medicine, University of Washington, Seattle, Washington, United States of America.
5 Department of Immunology, University of Washington, Seattle, Washington, United States of America.
6 Integrated Department in Immunology, University of Colorado Denver and National Jewish Health, Denver, Colorado, United States of America; Division of Gastroenterology & Hepatology, Hepatitis C Center, Department of Medicine, University of Colorado Denver, Aurora, Colorado, United States of America; Denver Veteran’s Affairs Medical Center, Denver, Colorado, United States of America.

 

Abstract

Plasmacytoid Dendritic Cells (pDCs) represent a key immune cell population in the defense against viruses. pDCs detect viral pathogen associated molecular patterns (PAMPs) through pattern recognition receptors (PRR). PRR/PAMP interactions trigger signaling events that induce interferon (IFN) production to initiate local and systemic responses. pDCs produce Type I and Type III (IFNL) IFNs in response to HCV RNA. Extracellular HCV core protein (Core) is found in the circulation in chronic infection. This study defined how Core modulates PRR signaling in pDCs. Type I and III IFN expression and production following exposure to recombinant Core or β-galactosiade was assessed in human GEN2.2 cells, a pDC cell line. Core suppressed type I and III IFN production in response to TLR agonists and the HCV PAMP agonist of RIG-I. Core suppression of IFN induction was linked with decreased IRF-7 protein levels and increased non-phosphorylated STAT1 protein. Circulating Core protein interferes with PRR signaling by pDCs to suppress IFN production. Strategies to define and target Core effects on pDCs may serve to enhance IFN production and antiviral actions against HCV.

PMID: 24788809

 

SUPPLEMENT:

Hepatitis C Virus (HCV) is a major public health problem infecting approximately 200 million people globally with chronic infection developing in the majority of patients. Untreated infection will lead to liver disease including fibrosis, scarring, liver cancer and, eventually, liver failure. Current therapies work to cure the virus about 70% of the time, but these therapies are difficult for the patient to tolerate and can be prohibitively expensive. Further, these therapies target the virus which encourages the virus to change in order to escape the treatment. New treatments that target the built-in host mechanisms that fight the virus may lead to better, safer treatments.

Plasmacytoid dendritic cells (pDCs) are a rare population of white blood cells whose job is to detect and respond to viruses. They control viruses by making a molecule called Interferon. Previous work from our lab showed that pDCs can recognize HCV and control virus through interferon production1. After that project, we were left with the question “If pDCs can control virus through interferon, why do patients develop chronic infection?” The work here answers that question. We hypothesized that a soluble factor made by the virus called Core (the nucleocapsid protein) could prevent pDCs from making enough interferon to control the virus.

We first examined the effect of adding Core to pDCs before stimulating the cells with synthetic mimics of viruses. We found that Core could prevent interferon production that would normally be produced when the pDCs came into contact with the viral mimics. We showed that Core could inhibit interferon production at both the transcriptional level (mRNA) and the translational level (protein). We also showed that Core does not kill our cells or prevent them from dividing as normal.

We next investigated the status of the transcription factors that control interferon production when Core was present. We found that Core did not affect one of the transcription factors, Interferon Regulatory Factor (IRF) 3, but did affect the other transcription factor, IRF7. Further, we showed that the activation status of those transcription factors did not change when Core was present.

We then looked downstream of interferon to ask if Core could stop the actions of interferon after the interferon was made. We found an increase in the total amount of the downstream transcription factor STAT1 but no changes in the ability of STAT1 to become activated. Other common innate immunity transcription factors were not affected by the presence of Core.

This work allowed us to conclude that HCV uses an immune evasion technique that effectively stops interferon production by pDCs through the production of its soluble factor Core. In summary, this study shows that Core directly inhibits production of interferon within human pDCs and dysregulates the transcription factors IRF-7 and STAT1, possibly contributing to the persistent HCV infection (Figure 1).

The impact of this work is two-fold. First, knowing that Core can turn off antiviral actions of the immune system, we can target this molecule for future therapeutic approaches. One way to do this would be to add an antibody that recognizes Core, potentially increasing the effectiveness of the current drugs as well as boost the patient’s immune system, leading to better outcomes. It is important that in resource-poor areas, direct-acting antivirals may not be available and hence additional therapeutic approaches should be considered. The work presented in this paper provides rationale to explore adding a blocking antibody to current therapies in the clinic. Second, this work furthers our understanding of HCV infection and immunity by demonstrating a new immune evasion technique that HCV uses stop the host immune system. The studies presented in this publication included responses to viral mimics as well as responses to HCV components, thus extending these observations to other infectious diseases. In total, this work provides a step forward in treating HCV and thus preventing its long-term complications such as endstage liver disease.

REFERENCES

1 Stone, A. E. et al. Hepatitis C virus pathogen associated molecular pattern (PAMP) triggers production of lambda-interferons by human plasmacytoid dendritic cells. PLoS Pathog 9, e1003316, doi:10.1371/journal.ppat.1003316 (2013).

Supported by NIH/NCATS Colorado CTSI Grant Number TL1 TR000155. Contents are the authors’ sole responsibility and do not necessarily represent official NIH views.

Figure-Biomed-frontiers
Figure 1: Paradigm Model of HCV Core acting on pDCs
(A) pDCs respond to stimulation by viruses to produce interferons. However, in the presence of Core (B), there is increased STAT1 but decreased interferon production. The decreased interferon results in decreased IRF-7 which inhibits future interferon production.

 

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