RACK1 mediates rewiring of intracellular networks induced by hepatitis C virus infection

Jae Seung Lee; Keisuke Tabata; Woan-Ing Twu; Md Shafiqur Rahman; Hee Sun Kim; Jin Bae Yu; Min Hyeok Jee; Ralf Bartenschlager; Sung Key Jang

doi:10.1371/journal.ppat.1008021

RACK1 mediates rewiring of intracellular networks induced by hepatitis C virus infection

PLoS Pathog. 2019 Sep 16;15(9):e1008021. doi: 10.1371/journal.ppat.1008021. eCollection 2019 Sep.

Authors

Affiliations

¹ Division of Integrative Bioscience & Biotechnology, POSTECH Biotech Center, POSTECH, Nam-gu, Pohang-si, Gyeongsangbuk-do, Rep. of KOREA.
² Department of Infectious Diseases, Molecular Virology, Heidelberg University, Heidelberg, Germany.
³ Department of Life Sciences, POSTECH Biotech Center, POSTECH, Nam-gu, Pohang-si, Gyeongsangbuk-do, Rep. of KOREA.
⁴ Division Virus-Associated Carcinogenesis, German Cancer Research Center, Heidelberg, Germany.

Abstract

Hepatitis C virus (HCV) is a positive-strand RNA virus replicating in a membranous replication organelle composed primarily of double-membrane vesicles (DMVs) having morphological resemblance to autophagosomes. To define the mechanism of DMV formation and the possible link to autophagy, we conducted a yeast two-hybrid screening revealing 32 cellular proteins potentially interacting with HCV proteins. Among these was the Receptor for Activated Protein C Kinase 1 (RACK1), a scaffolding protein involved in many cellular processes, including autophagy. Depletion of RACK1 strongly inhibits HCV RNA replication without affecting HCV internal ribosome entry site (IRES) activity. RACK1 is required for the rewiring of subcellular membranous structures and for the induction of autophagy. RACK1 binds to HCV nonstructural protein 5A (NS5A), which induces DMV formation. NS5A interacts with ATG14L in a RACK1 dependent manner, and with the ATG14L-Beclin1-Vps34-Vps15 complex that is required for autophagosome formation. Both RACK1 and ATG14L are required for HCV DMV formation and viral RNA replication. These results indicate that NS5A participates in the formation of the HCV replication organelle through interactions with RACK1 and ATG14L.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adaptor Proteins, Vesicular Transport / metabolism
Autophagosomes / metabolism
Autophagosomes / virology
Autophagy
Autophagy-Related Proteins / metabolism
Cell Line
Hepacivirus / genetics
Hepacivirus / pathogenicity
Hepacivirus / physiology
Hepatitis C / metabolism*
Hepatitis C / pathology
Hepatitis C / virology*
Hepatocytes / metabolism
Hepatocytes / pathology
Hepatocytes / virology
Host Microbial Interactions / physiology
Humans
Metabolic Networks and Pathways
Neoplasm Proteins / metabolism*
Protein Binding
Protein Interaction Domains and Motifs
RNA, Viral / biosynthesis
Receptors for Activated C Kinase / metabolism*
Two-Hybrid System Techniques
Viral Nonstructural Proteins / chemistry
Viral Nonstructural Proteins / metabolism
Virus Replication

Substances

ATG14 protein, human
Adaptor Proteins, Vesicular Transport
Autophagy-Related Proteins
Neoplasm Proteins
RACK1 protein, human
RNA, Viral
Receptors for Activated C Kinase
Viral Nonstructural Proteins
NS-5 protein, hepatitis C virus

Grants and funding

This work was supported by the National Research Foundation of Korea (NRF) Grant funded by the Korean Government (MSIP) (NO. NRF-2017R1A5A1015366), and the Bio & Medical Technology Development Program of the National Research Foundation (NRF) funded by the Ministry of Science and ICT (NO. NRF-2017M3A9F6029755). Funding for the open access charge was provided by the National Research Foundation of Korea (NRF). R.B. is supported by the Deutsche Forschungsgemeinschaft, TRR179, TP11. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.