Demethylase JMJD6 as a New Regulator of Interferon Signaling: Effects of HCV and Ethanol Metabolism

Abstract

Background & AimsAlcohol-induced progression of hepatitis C virus (HCV) infection is related to dysfunction of innate immunity in hepatocytes. Endogenously produced interferon (IFN)α induces activation of interferon-stimulated genes (ISGs) via triggering of the Janus kinase–signal transducer and activator of transcription 1 (STAT1) pathway. This activation requires protein methyltransferase 1–regulated arginine methylation of STAT1. Here, we aimed to study whether STAT1 methylation also depended on the levels of demethylase jumonji domain-containing 6 protein (JMJD6) and whether ethanol and HCV affect JMJD6 expression in hepatocytes.MethodsHuh7.5-CYP (RLW) cells and hepatocytes were exposed to acetaldehyde-generating system (AGS) and 50 mmol/L ethanol, respectively. JMJD6 messenger RNA and protein expression were measured by real-time polymerase chain reaction and Western blot. IFNα-activated cells either overexpressing JMJD6 or with knocked-down JMJD6 expression were tested for STAT1 methylation, ISG activation, and HCV RNA. In vivo studies have been performed on C57Bl/6 mice (expressing HCV structural proteins or not) or chimeric mice with humanized livers fed control or ethanol diets.ResultsAGS exposure to cells up-regulated JMJD6 expression in RLW cells. These results were corroborated by ethanol treatment of primary hepatocytes. The promethylating agent betaine reversed the effects of AGS/ethanol. Similar results were obtained in vivo, when mice were fed control/ethanol with and without betaine supplementation. Overexpression of JMJD6 suppressed STAT1 methylation, IFNα-induced ISG activation, and increased HCV-RNA levels. In contrast, JMJD6 silencing enhanced STAT1 methylation, ISG stimulation by IFNα, and attenuated HCV-RNA expression in Huh7.5 cells.ConclusionsWe conclude that arginine methylation of STAT1 is suppressed by JMJD6. Both HCV and acetaldehyde increase JMJD6 levels, thereby impairing STAT1 methylation and innate immunity protection in hepatocytes exposed to the virus and/or alcohol.

Human hepatocytes depletion in the presence of HIV-1 infection in dual reconstituted humanized mice

“…TK-NOG (NOD.Cg-Prkdc scid Il2rg tm1Sug Tg(Alb-TK)7-2/ShiJic) mice expressing a liver-specific herpes simplex virus type 1 TK transgene under the control of an ALB promoter were provided by the Central Institute for Experimental Animals (CIEA, Japan; Drs Mamoru Ito and Hiroshi Suemizu) ( Ganesan et al, 2017b ). Mice were bred and housed in the pathogen-free animal facility at the University of Nebraska Medical Center (UNMC).…”

Alcohol Metabolism Potentiates HIV-Induced Hepatotoxicity: Contribution to End-Stage Liver Disease

“…Human immunodeficiency virus-RNA (HIV RNA), interferon-stimulated genes (ISGs) with anti-viral activities such as ISG15, 2 -5 -oligoadenylate synthetase 1 (OAS1), inflammasome markers NLR family pyrin domain containing 3 (NLRP3), caspase-1, Interleukin (IL)-1β, and IL-18, and pro-fibrotic markers collagen 1 A1 (Col1A1), transforming growth factor beta (TGFβ), and prostaglandin D receptor 2 (PTGDR2) were quantified by real-time PCR as previously described [27]. Total RNA was isolated from cells using Trizol Reagent.…”

“…These were incubated in a Model 7500 qRT-PCR thermal cycler. The relative quantity of each RNA transcript was calculated by its threshold cycle (Ct) after subtracting that of the reference cDNA (GAPDH) as described before [27]. Reverse transcriptase (RT) activity was detected in cell supernatants as described [28].…”

“…Samples were subjected to denaturing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) in polyacrylamide gels. Western blot was performed as described previously [27], blots were developed using Odyssey ® infrared imaging system (Li-Cor Bioscience, Lincoln, NE, USA), and the protein band was quantified using Li-Cor software. The B-actin was used as the loading control to normalize the proteins.…”

Jumonji: Welcome to the World of Interferon Signaling in Alcohol and HCV

“… 1 It is known that the increase in hepatotoxicity is caused in part by increased viral replication and impaired innate immune responses. 1 , 2 A novel mechanism for alcohol-accelerated HCV replication is identified in the article by Ganesan et al 3 in this issue of Cellular and Molecular Gastroenterology and Hepatology . The critical role of arginine methylation of STAT1 in interferon (IFN)-α-induced JAK-STAT signaling and expression of antiviral genes (ISGs) is well-established.…”