Role of N Terminus-Truncated NS1 Proteins of Influenza A Virus in Inhibiting IRF3 Activation

Abstract

The NS1 protein encoded by influenza A virus antagonizes the interferon response through various mechanisms, including blocking cellular mRNA maturation by binding the cellular CPSF30 3= end processing factor and/or suppressing the activation of interferon regulatory factor 3 (IRF3). In the present study, we identified two truncated NS1 proteins that are translated from internal AUGs at positions 235 and 241 of the NS1 open reading frame. We analyzed the cellular localization and function of the N-truncated NS1 proteins encoded by two influenza A virus strains, Udorn/72/H3N2 (Ud) and Puerto Rico/8/34/H1N1 (PR8). The NS1 protein of PR8, but not Ud, inhibits the activation of IRF3, whereas the NS1 protein of Ud, but not PR8, binds CPSF30. The truncated PR8 NS1 proteins are localized in the cytoplasm, whereas the full-length PR8 NS1 protein is localized in the nucleus. The infection of cells with a PR8 virus expressing an NS1 protein containing mutations of the two in-frame AUGs results in both the absence of truncated NS1 proteins and the reduced inhibition of activation of IRF3 and beta interferon (IFN-␤) transcription. The expression of the truncated PR8 NS1 protein by itself enhances the inhibition of the activation of IRF3 and IFN-␤ transcription in Ud virus-infected cells. These results demonstrate that truncated PR8 NS1 proteins contribute to the inhibition of activation of this innate immune response. In contrast, the N-truncated NS1 proteins of the Ud strain, like the full-length NS1 protein, are localized in the nucleus, and mutation of the two in-frame AUGs has no effect on the activation of IRF3 and IFN-␤ transcription. IMPORTANCEInfluenza A virus causes pandemics and annual epidemics in the human population. The viral NS1 protein plays a critical role in suppressing type I interferon expression. In the present study, we identified two novel truncated NS1 proteins that are translated from the second and third in-frame AUG codons in the NS1 open reading frame. The N-terminally truncated NS1 encoded by the H1N1 PR8 strain of influenza virus that suppresses IRF3 activation is localized primarily in the cytoplasm. We demonstrate that this truncated NS1 protein by itself enhances this suppression, demonstrating that some strains of influenza A virus express truncated forms of the NS1 protein that function in the inhibition of cytoplasmic antiviral events.

Influenza Research Database: An integrated bioinformatics resource for influenza virus research

“…As new variant proteins are identified by the influenza research community and reported in the scientific literature (26), the IRD team evaluates the strength of evidence for the presence and importance of these novel proteins, determines the sequence signals that can be used to predict their expression, and adds the specific prediction algorithm to our variant protein prediction infrastructure, making these sequence annotations uniquely available in IRD for user query and downstream analysis.…”

Nuclear TRIM25 Specifically Targets Influenza Virus Ribonucleoproteins to Block the Onset of RNA Chain Elongation

“…One function of the NS1 protein is to bind the TRIM25 E3 ligase, inhibiting its multimerization and thus its ligase activity, an action that was postulated to block activation of the RIG-I pathway (Gack et al, 2009). However, not all human-circulating influenza A virus strains block RIG-I-mediated activation of IRF3 and interferon-β transcription (Kuo et al, 2016; 2010), although the NS1 proteins of all tested strains bind TRIM25 (Gack et al, 2009; Kuo et al, 2010; Rajsbaum et al, 2012). Notably, strong activation of IRF3 and interferon-β transcription still occurs in cells infected with viruses expressing NS1 proteins of human-circulating H3N2 and H2N2 viruses (Kuo et al, 2010; 2016).…”

“…However, not all human-circulating influenza A virus strains block RIG-I-mediated activation of IRF3 and interferon-β transcription (Kuo et al, 2016; 2010), although the NS1 proteins of all tested strains bind TRIM25 (Gack et al, 2009; Kuo et al, 2010; Rajsbaum et al, 2012). Notably, strong activation of IRF3 and interferon-β transcription still occurs in cells infected with viruses expressing NS1 proteins of human-circulating H3N2 and H2N2 viruses (Kuo et al, 2010; 2016). Consequently, binding of TRIM25 by the NS1 protein does not directly correlate to the suppression of interferon-β induction in influenza A virus-infected cells.…”

The C-Terminal Effector Domain of Non-Structural Protein 1 of Influenza A Virus Blocks IFN-β Production by Targeting TNF Receptor-Associated Factor 3

“…The data are presented as the means ± SD derived from three repeat experiments. *P < 0.05 or **P < 0.01, ***P < 0.001 (Student's t-test).Qian et al NS1 Inhibits IFN-β through TRAF3 Frontiers in Immunology | www.frontiersin.org July 2017 | Volume 8 | Article 779translated from internal AUGs at positions 235 and 241 of the NS1 open reading frame, were found to inhibit the activation of IRF3 and IFN-β transcription(41). This observation is supported by our findings that the influenza A virus NS1 ED plays an important role in the ability of the NS1 protein to inhibit IFN-β pathway.…”