Herc5 Attenuates Influenza A Virus by Catalyzing ISGylation of Viral NS1 Protein

Tang, Yujie; Zhong, Gongxun; Zhu, Lianhui; Liu, Xing; Shan, Yufei; Feng, Huapeng; Bu, Zhigao; Chen, Hualan; Wang, Chen

doi:10.4049/jimmunol.0903588

Cited by 132 publications

(140 citation statements)

References 62 publications

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“…Then, culture media was replaced by serum-free DMEM, and SeV, vesicular stomatitis virus (VSV), influenza A virus (IAV), or Newcastle disease virus (NDV)-GFP was added to the media at a multiplicity of infection of 0.2-1 according to specific experiments. After 1 h, the medium was removed, and the cells were fed with DMEM containing 10% FBS (32,33). Human H1N1 IAV strain IAV-PR8/34 was kindly provided by Dr. Ze Chen (Wuhan Institute of Virology, Chinese Academy of Sciences).…”

Section: Virus Manipulationmentioning

confidence: 99%

IFN-Induced TPR Protein IFIT3 Potentiates Antiviral Signaling by Bridging MAVS and TBK1

Liu

Chen

Wei

et al. 2011

The Journal of Immunology

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153

140

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Intracellular RNA viruses are sensed by receptors retinoic acid-inducible gene I/MDA5, which trigger formation of the mitochondrial antiviral signaling (MAVS) complex on mitochondria. Consequently, this leads to the activation of TNFR-associated factor family member-associated NF-κB activator-binding kinase 1 (TBK1) and phosphorylation of IFN regulatory factor 3 (IRF3). It remains to be elucidated how MAVS activates TBK1/IRF3. In this study, we report that IFN-induced protein with tetratricopeptide repeats 3 (IFIT3) is significantly induced upon RNA virus infection. Ectopic expression or knockdown of IFIT3 could, respectively, enhance or impair IRF3-mediated gene expression. Mechanistically, the tetratrico-peptide repeat motif (E164/E165) of IFIT3 interacts with the N terminus (K38) of TBK1, thus bridging TBK1 to MAVS on the mitochondrion. Disruption of this interaction markedly attenuates the activation of TBK1 and IRF3. Furthermore, host antiviral responses are significantly boosted or crippled in the presence or absence of IFIT3. Collectively, our study characterizes IFIT3 as an important modulator in innate immunity, revealing a new function of the IFIT family proteins (IFN-induced protein with tetratricopeptide repeats).

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Section: Virus Manipulationmentioning

confidence: 99%

IFN-Induced TPR Protein IFIT3 Potentiates Antiviral Signaling by Bridging MAVS and TBK1

Liu

Chen

Wei

et al. 2011

The Journal of Immunology

Self Cite

153

140

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“…The initial report identifying NS1 as an ISGylated protein indicated that ISGylation prevented the interaction of NS1 with importin-␣, a karyopherin essential for its nuclear import (30). A subsequent report described three additional roles for NS1 ISGylation: disrupting NS1 dimerization, preventing its interaction with PKR, and exhibiting an overall negative effect on NS1's ability to neutralize the IFN system (31).…”

mentioning

confidence: 99%

SUMOylation Affects the Interferon Blocking Activity of the Influenza A Nonstructural Protein NS1 without Affecting Its Stability or Cellular Localization

Santos

Pal

Chacon

et al. 2013

J Virol

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Our pioneering studies on the interplay between the small ubiquitin-like modifier (SUMO) and influenza A virus identified the nonstructural protein NS1 as the first known SUMO target of influenza virus and one of the most abundantly SUMOylated influenza virus proteins. Here, we further characterize the role of SUMOylation for the A/Puerto Rico/8/1934 (PR8) NS1 protein, demonstrating that NS1 is SUMOylated not only by SUMO1 but also by SUMO2/3 and mapping the main SUMOylation sites in NS1 to residues K219 and K70. Furthermore, by using SUMOylatable and non-SUMOylatable forms of NS1 and an NS1-specific artificial SUMO ligase (ASL) that increases NS1 SUMOylation ϳ4-fold, we demonstrate that SUMOylation does not affect the stability or cellular localization of PR8 NS1. However, NS1's ability to be SUMOylated appears to affect virus multiplication, as indicated by the delayed growth of a virus expressing the non-SUMOylatable form of NS1 in the interferon (IFN)-competent MDCK cell line. Remarkably, while a non-SUMOylatable form of NS1 exhibited a substantially diminished ability to neutralize IFN production, increasing NS1 SUMOylation beyond its normal levels also exerted a negative effect on its IFN-blocking function. This observation indicates the existence of an optimal level of NS1 SUMOylation that allows NS1 to achieve maximal activity and suggests that the limited amount of SUMOylation normally observed for most SUMO targets may correspond to an optimal level that maximizes the contribution of SUMOylation to protein function. Finally, protein cross-linking data suggest that SUMOylation may affect NS1 function by regulating the abundance of NS1 dimers and trimers in the cell.

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“…The NS1 protein undergoes three posttranslational modifications: phosphorylation by cellular kinases (4,12,13), ISG15 modification by the interferon (IFN)-induced ISG15 conjugation system (17,21), and modification by SUMO (11,20). It has been reported that these modifications affect the NS1 protein and the phenotype of the virus (4,17,21).…”

mentioning

confidence: 99%

“…It has been reported that these modifications affect the NS1 protein and the phenotype of the virus (4,17,21). Phosphorylation of threonines (Ts) of the NS1 protein was observed many years ago (12,13).…”

mentioning

confidence: 99%

Roles of the Phosphorylation of Specific Serines and Threonines in the NS1 Protein of Human Influenza A Viruses

Hsiang

Zhou

Krug

2012

J Virol

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We demonstrate that phosphorylation of the NS1 protein of a human influenza A virus occurs not only at the threonine (T) at position 215 but also at serines (Ss), specifically at positions 42 and 48. By generating recombinant influenza A/Udorn/72 (Ud) viruses that encode mutant NS1 proteins, we determined the roles of these phosphorylations in virus replication. At position 215 only a T-to-A substitution attenuated replication, whereas other substitutions (T to E to mimic constitutive phosphorylation, T to N, and T to P, the amino acid in avian influenza A virus NS1 proteins) had no effect. We conclude that attenuation resulting from the T-to-A substitution at position 215 is attributable to a deleterious structural change in the NS1 protein that is not caused by other amino acid substitutions and that phosphorylation of T215 does not affect virus replication. At position 48 neither an S-to-A substitution nor an S-to-D substitution that mimics constitutive phosphorylation affected virus replication. In contrast, at position 42, an S-to-D, but not an S-to-A, substitution caused attenuation. The S-to-D substitution eliminates detectable double-stranded RNA binding by the NS1 protein, accounting for attenuation of virus replication. We show that protein kinase C ␣ (PKC␣) catalyzes S42 phosphorylation. Consequently, the only phosphorylation of the NS1 protein of this human influenza A virus that regulates its replication is S42 phosphorylation catalyzed by PKC␣. In contrast, phosphorylation of Ts or Ss in the NS1 protein of the 2009 H1N1 pandemic virus was not detected, indicating that NS1 phosphorylation probably does not play any role in the replication of this virus. Influenza A viruses cause a highly contagious respiratory disease in humans, resulting in annual epidemics and periodic worldwide pandemics. The smallest of the eight negative-sense viral gene segments encodes the NS1 protein, a nonstructural protein that plays multiple crucial roles in virus replication (5). Most NS1 proteins are 230 to 237 amino acids long, although shorter forms have also been found. The NS1 protein comprises two functional domains: the N-terminal (amino acids 1 to 73) RNA-binding domain (RBD) and the C-terminal (amino acids 74 to 230/237) effector domain (ED), which binds several cellular proteins.The NS1 protein undergoes three posttranslational modifications: phosphorylation by cellular kinases (4, 12, 13), ISG15 modification by the interferon (IFN)-induced ISG15 conjugation system (17, 21), and modification by SUMO (11,20). It has been reported that these modifications affect the NS1 protein and the phenotype of the virus (4, 17, 21). Phosphorylation of threonines (Ts) of the NS1 protein was observed many years ago (12, 13). It was reported that T at position 215 (T215) of the NS1 protein of the human H3N2 influenza A/Udorn/72 virus (Ud) is phosphorylated and that this phosphorylation is important for virus replication (4). The basis for the latter conclusion was that a recombinant Ud virrus encoding an NS1 protein with a T-to...

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Herc5 Attenuates Influenza A Virus by Catalyzing ISGylation of Viral NS1 Protein

Cited by 132 publications

References 62 publications

IFN-Induced TPR Protein IFIT3 Potentiates Antiviral Signaling by Bridging MAVS and TBK1

IFN-Induced TPR Protein IFIT3 Potentiates Antiviral Signaling by Bridging MAVS and TBK1

SUMOylation Affects the Interferon Blocking Activity of the Influenza A Nonstructural Protein NS1 without Affecting Its Stability or Cellular Localization

Roles of the Phosphorylation of Specific Serines and Threonines in the NS1 Protein of Human Influenza A Viruses

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