OTUB1 contributes to the stability and function of Influenza A virus NS2

Yu-Jyun Li; Chi-Yuan Chen; Yu-Shen Kuo; Yi-Wen Huang; Rei-Lin Kuo; Li-Kwan Chang; Jeng-How Yang; Chih-Ho Lai; Shin-Ru Shih; Ya-Fang Chiu

doi:10.1371/journal.ppat.1012279

OTUB1 contributes to the stability and function of Influenza A virus NS2

PLoS Pathog. 2024 May 30;20(5):e1012279. doi: 10.1371/journal.ppat.1012279. eCollection 2024 May.

Authors

Yu-Jyun Li^{1

2}, Chi-Yuan Chen¹, Yu-Shen Kuo^{1

2}, Yi-Wen Huang^{1

2}, Rei-Lin Kuo³, Li-Kwan Chang⁴, Jeng-How Yang⁵, Chih-Ho Lai^{1

2}, Shin-Ru Shih³, Ya-Fang Chiu^{1

2

3

5

6}

Affiliations

¹ Department of Microbiology and Immunology, Chang Gung University, Taoyuan, Taiwan.
² Graduate Institute of Biomedical Sciences, Chang Gung University, Taoyuan, Taiwan.
³ Research Center for Emerging Viral Infections, Chang Gung University, Taoyuan, Taiwan.
⁴ Department of Biochemical Science and Technology, College of Life Science, National Taiwan University, Taipei, Taiwan.
⁵ Division of Infectious Diseases, Department of Medicine, Chang Gung Memorial Hospital, New Taipei, Taiwan.
⁶ Department of Laboratory Medicine, Linkou Chang Gung Memorial Hospital, Taoyuan, Taiwan.

Abstract

The influenza A virus (IAV) consists of 8 single-stranded, negative-sense viral RNA (vRNA) segments. After infection, vRNA is transcribed, replicated, and wrapped by viral nucleoprotein (NP) to form viral ribonucleoprotein (vRNP). The transcription, replication, and nuclear export of the viral genome are regulated by the IAV protein, NS2, which is translated from spliced mRNA transcribed from viral NS vRNA. This splicing is inefficient, explaining why NS2 is present in low abundance after IAV infection. The levels of NS2 and its subsequent accumulation are thought to influence viral RNA replication and vRNP nuclear export. Here we show that NS2 is ubiquitinated at the K64 and K88 residues by K48-linked and K63-linked polyubiquitin (polyUb) chains, leading to the degradation of NS2 by the proteasome. Additionally, we show that a host deubiquitinase, OTUB1, can remove polyUb chains conjugated to NS2, thereby stabilizing NS2. Accordingly, knock down of OTUB1 by siRNA reduces the nuclear export of vRNP, and reduces the overall production of IAV. These results collectively demonstrate that the levels of NS2 in IAV-infected cells are regulated by a ubiquitination-deubiquitination system involving OTUB1 that is necessary for optimal IAV replication.

Copyright: © 2024 Li et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

MeSH terms

Animals
Cell Line
Chlorocebus aethiops
Cysteine Endopeptidases* / genetics
Cysteine Endopeptidases* / metabolism
Deubiquitinating Enzymes / metabolism
Dogs
HEK293 Cells
Humans
Influenza A virus* / metabolism
Influenza, Human / metabolism
Influenza, Human / virology
RNA, Viral / genetics
RNA, Viral / metabolism
Ubiquitination
Vero Cells
Viral Nonstructural Proteins* / genetics
Viral Nonstructural Proteins* / metabolism
Virus Replication* / physiology

Substances

Cysteine Endopeptidases
Deubiquitinating Enzymes
NS2 protein, influenza virus A
OTUB1 protein, human
RNA, Viral
Viral Nonstructural Proteins

Grants and funding

This work was financially supported by the Ministry of Science and Technology (MOST; now the National Science and Technology Council, NSTC, URL: https://www.nstc.gov.tw/?l=en), Taiwan (MOST 109-2320-B-182-028-MY3, to Y.F.C; NSTC 112-2320-B-182-033, to Y.F.C); the Chang Gung Medical Research Program (URL: https://www1.cgmh.org.tw/intr/intr2/c3s000/research/e_index.html; CMRPD1M0871, to Y.F.C); Chang Gung Memorial Hospital, Linkou (URL: https://www.cgmh.org.tw/eng/about/system/4; BMRPF14, to Y.F.C); and the Research Center for Emerging Viral Infections (URL: https://rcevi.cgu.edu.tw/index.php?Lang=en; to S.R.S.) from The Featured Areas Research Center Program within the framework of the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.