HCV activates oxidative stress defense system and suppresses

polyamine biosynthesis

Smirnova OlgaSmirnova Olga

Engelhardt Institute of Molecular BiologyRussian Academy of Sciences

HCV induces oxidative stress: mechanisms and HCV induces oxidative stress: mechanisms and consequencesconsequences

1.1. HCV-induced oxidative stress can contribute to:HCV-induced oxidative stress can contribute to:• Liver fibrosisLiver fibrosis• ““Non-Vogelstein type” cancerogenesisNon-Vogelstein type” cancerogenesis• Interferon Interferon resistance in chronic hepatitis C patients resistance in chronic hepatitis C patients• Insulin resistance Insulin resistance • Alteration of iron homeostasis Alteration of iron homeostasis viavia inhibition of hepcidin expression inhibition of hepcidin expression

2.2. HCV causes oxidative stress by multiple mechanisms:HCV causes oxidative stress by multiple mechanisms:• Core and NS5A proteins alter calcium homeostasis resulting in Core and NS5A proteins alter calcium homeostasis resulting in

accumulation of Reactive Oxygen Species (ROS)accumulation of Reactive Oxygen Species (ROS)• E1, E2, and NS4B proteins induce ER-stress which can also lead to E1, E2, and NS4B proteins induce ER-stress which can also lead to

activation of ER oxidoreductases and to accumulation of reactive oxygen activation of ER oxidoreductases and to accumulation of reactive oxygen species (ROS)species (ROS)

• HCV proteins activate NADPH oxidase 4 (Nox4) expressionHCV proteins activate NADPH oxidase 4 (Nox4) expression

ROS [ROS]* Nonhazardousproductsantioxidants

Phase II detoxyfying

enzymes

ARE Phase II enzymes

Nrf2 Keap1

kinases

р

РRE SSATSpermine/spermidineN1-acetyltransferase

Nrf2р

NS5ACapsidprotein

Nrf2

NS4В PERKER

stress

pcDNA3.1(+)

pGL3-promoter luciferase luciferase

pP-ARE pP-PRE

inhibitors

Regulation of antioxidant defense systemRegulation of antioxidant defense system

ARE (Antioxidant Response Element)

PRE(Polyamine Response Element)

HCV proteins induce oxidative stress HCV proteins induce oxidative stress

HCV proteins activate ARE-dependent protein HCV proteins activate ARE-dependent protein expressionexpression

Up-regulation of HO-1 and Nqo1 gene expression by HCV proteins

Impact of HCV proteins on ARE-dependent luciferase expression

HCV proteins activate ARE-dependent transcription by HCV proteins activate ARE-dependent transcription by ROS-dependent and –independent mechanismsROS-dependent and –independent mechanisms

PDTC (pyrrolidine dithiocarbamate) acts as ROS scavenger

HCV proteins induce ARE-luciferase expression via PKC in a HCV proteins induce ARE-luciferase expression via PKC in a ROS-dependent, and by PI3K and CK2 in a ROS-ROS-dependent, and by PI3K and CK2 in a ROS-

independent manner.independent manner.

Wo (Wortmannin) – PI3K inhibitor Ro (Ro31-8220) – PKC inhibitorDRB – CK2 inhibitor

Effect of protein kinase inhibitors and PDTC on HO-1/Nqo1

gene expression, and Nrf2 localization

ConclusionsConclusions

► HCV proteins induced strong up-regulation of antioxidant defense system that HCV proteins induced strong up-regulation of antioxidant defense system that might mitigate harmful effects of HCV-induced oxidative stress during acute might mitigate harmful effects of HCV-induced oxidative stress during acute stage of hepatitis Cstage of hepatitis C

► Core, E1, E2, NS4B, and NS5A activate Nrf2/ARE pathway through three Core, E1, E2, NS4B, and NS5A activate Nrf2/ARE pathway through three independent mechanismsindependent mechanisms

► All five proteins induced accumulation of reactive oxygen species (ROS) which All five proteins induced accumulation of reactive oxygen species (ROS) which activated protein kinaseactivated protein kinase CC

► Core and NS5A proteins induced ROS-independent activation of casein kinase Core and NS5A proteins induced ROS-independent activation of casein kinase 2 and phosphoinositide-3 kinase2 and phosphoinositide-3 kinase

► The effects of NS4B, E1, and E2 were presumably mediated The effects of NS4B, E1, and E2 were presumably mediated via via ROS-ROS-independent PERK pathwayindependent PERK pathway

AcknowledgementsAcknowledgements

Sergey Kochetkov lab (EIMB)Sergey Kochetkov lab (EIMB) Karolinska InstitutetKarolinska Institutet ((SwedenSweden))

Alex IvanovAlex Ivanov Maria Isaguliants Maria Isaguliants Olga IvanovaOlga Ivanova Alexey KhomutovAlexey Khomutov

A.I. Virtanen Institute (Finland)A.I. Virtanen Institute (Finland) Tuomo KeinänenTuomo Keinänen

Mervi HyvonenMervi Hyvonen

The project was supported byThe project was supported by::► Russian Foundation for Basic ResearchRussian Foundation for Basic Research► Grant of President of Russian FederationGrant of President of Russian Federation

THANK YOU FOR YOUR ATTENTIONTHANK YOU FOR YOUR ATTENTION