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J Biol Chem. 2016 Jul 15;291(29):15093-107. doi: 10.1074/jbc.M116.717942. Epub 2016 May 17.

Interferon β (IFN-β) Production during the Double-stranded RNA (dsRNA) Response in Hepatocytes Involves Coordinated and Feedforward Signaling through Toll-like Receptor 3 (TLR3), RNA-dependent Protein Kinase (PKR), Inducible Nitric Oxide Synthase (iNOS), and Src Protein.

Author information

1
From the Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213.
2
From the Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, the Family Planning Research Institute, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
3
From the Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, the Center for Biologic Imaging, University of Pittsburgh, Pittsburgh, Pennsylvania 15213, and.
4
the Xiangya Third Hospital and Central South University School of Medicine, Changsha, China.
5
From the Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213, billiartr@upmc.edu.

Abstract

The sensing of double-stranded RNA (dsRNA) in the liver is important for antiviral defenses but can also contribute to sterile inflammation during liver injury. Hepatocytes are often the target of viral infection and are easily injured by inflammatory insults. Here we sought to establish the pathways involved in the production of type I interferons (IFN-I) in response to extracellular poly(I:C), a dsRNA mimetic, in hepatocytes. This was of interest because hepatocytes are long-lived and, unlike most immune cells that readily die after activation with dsRNA, are not viewed as cells with robust antimicrobial capacity. We found that poly(I:C) leads to rapid up-regulation of inducible nitric oxide synthase (iNOS), double-stranded RNA-dependent protein kinase (PKR), and Src. The production of IFN-β was dependent on iNOS, PKR, and Src and partially dependent on TLR3/Trif. iNOS and Src up-regulation was partially dependent on TLR3/Trif but entirely dependent on PKR. The phosphorylation of TLR3 on tyrosine 759 was shown to increase in parallel to IFN-β production in an iNOS- and Src-dependent manner, and Src was found to directly interact with TLR3 in the endosomal compartment of poly(I:C)-treated cells. Furthermore, we identified a robust NO/cGMP/PKG-dependent feedforward pathway for the amplification of iNOS expression. These data identify iNOS/NO as an integral component of IFN-β production in response to dsRNA in hepatocytes in a pathway that involves the coordinated activities of TLR3/Trif and PKR.

KEYWORDS:

TIR domain-containing adapter-inducing interferon B (TRIF); Toll-like receptor (TLR); hepatocyte; immunology; inflammation; nitric oxide; nitric oxide synthase

PMID:
27226571
PMCID:
PMC4946926
DOI:
10.1074/jbc.M116.717942
[Indexed for MEDLINE]
Free PMC Article

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