Pyrrolidine dithiocarbamate inhibits enterovirus 71 replication by down-regulating ubiquitin–proteasome system

Lin, Lexun; Qin, Ying; Wu, Heng; Chen, Yang; Wu, Shuo; Si, Xiaoning; Wang, Hui; Wang, Tianying; Zhong, Xiaoyan; Zhai, Xia; Tong, Lei; Pan, Bo; Zhang, Fengmin; Zhong, Zhaohua; Wang, Yan; Zhao, Wenran

doi:10.1016/j.virusres.2014.10.012

Cited by 15 publications

(13 citation statements)

References 72 publications

(85 reference statements)

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“…In such a case, ubiquitin might be hybridized with poly-SUMO chains or conjugated on the exposed internal residues of 3D due to the conformational change induced by SUMOylation. The importance of UPS during replication of both CVB3 and EV71 may rely on 3D SUMOylation (65,66). We found that the degradation of 3D by K63-linked ubiquitination was not observed and that wildtype 3D showed a longer half-life than the SUMOylation-deficient mutant K159R/150-152 SIM.…”

Section: Discussionmentioning

confidence: 78%

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SUMO Modification Stabilizes Enterovirus 71 Polymerase 3D To Facilitate Viral Replication

Liu

Zheng

Shu

et al. 2016

J Virol

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Accumulating evidence suggests that viruses hijack cellular proteins to circumvent the host immune system. Ubiquitination and SUMOylation are extensively studied posttranslational modifications (PTMs) that play critical roles in diverse biological processes. Cross talk between ubiquitination and SUMOylation of both host and viral proteins has been reported to result in distinct functional consequences. Enterovirus 71 (EV71), an RNA virus belonging to the family Picornaviridae, is a common cause of hand, foot, and mouth disease. Little is known concerning how host PTM systems interact with enteroviruses. Here, we demonstrate that the 3D protein, an RNA-dependent RNA polymerase (RdRp) of EV71, is modified by small ubiquitin-like modifier 1 (SUMO-1) both during infection and in vitro. Residues K159 and L150/D151/L152 were responsible for 3D SUMOylation as determined by bioinformatics prediction combined with site-directed mutagenesis. Also, primer-dependent polymerase assays indicated that mutation of SUMOylation sites impaired 3D polymerase activity and virus replication. Moreover, 3D is ubiquitinated in a SUMO-dependent manner, and SUMOylation is crucial for 3D stability, which may be due to the interplay between the two PTMs. Importantly, increasing the level of SUMO-1 in EV71-infected cells augmented the SUMOylation and ubiquitination levels of 3D, leading to enhanced replication of EV71. These results together suggested that SUMO and ubiquitin cooperatively regulated EV71 infection, either by SUMO-ubiquitin hybrid chains or by ubiquitin conjugating to the exposed lysine residue through SUMOylation. Our study provides new insight into how a virus utilizes cellular pathways to facilitate its replication. IMPORTANCEInfection with enterovirus 71 (EV71) often causes neurological diseases in children, and EV71 is responsible for the majority of fatalities. Based on a better understanding of interplay between virus and host cell, antiviral drugs against enteroviruses may be developed. As a dynamic cellular process of posttranslational modification, SUMOylation regulates global cellular protein localization, interaction, stability, and enzymatic activity. However, little is known concerning how SUMOylation directly influences virus replication by targeting viral polymerase. Here, we found that EV71 polymerase 3D was SUMOylated during EV71 infection and in vitro. Moreover, the SUMOylation sites were determined, and in vitro polymerase assays indicated that mutations at SUMOylation sites could impair polymerase synthesis. Importantly, 3D is ubiquitinated in a SUMOylation-dependent manner that enhances the stability of the viral polymerase. Our findings indicate that the two modifications likely cooperatively enhance virus replication. Our study may offer a new therapeutic strategy against virus replication. S mall ubiquitin-like modifier (SUMO) modification, a member of the group of posttranslational modifications (PTMs), is important for the regulation of many cellular proteins and pathways (1). SUMO is a novel...

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Section: Discussionmentioning

confidence: 78%

“…The ubiquitin-proteasome system (UPS) is required for the effective replication of both coxsackievirus B3 (CVB3) (65) and EV71 (66). Since SUMOylation can interact with ubiquitination in many viral proteins, we determined whether EV71 3D is ubiquitinated.…”

Section: D Polymerase Is Sumoylated During Ev71 Infectionmentioning

confidence: 99%

SUMO Modification Stabilizes Enterovirus 71 Polymerase 3D To Facilitate Viral Replication

Liu

Zheng

Shu

et al. 2016

J Virol

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“…The UPS plays an important role in the degradation of intracellular proteins and regulates the cellular signalling pathway [ 48 ]. Many viruses have manipulated this system to their own advantage [ 40 , 49 – 51 ]. In the present study, we found that proteasome inhibition impacts ASF infection leading to infectivity reduction, as determined by the decrease in the number of cells expressing late protein p72.…”

Section: Discussionmentioning

confidence: 99%

The ubiquitin-proteasome system is required for African swine fever replication

et al. 2017

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Several viruses manipulate the ubiquitin-proteasome system (UPS) to initiate a productive infection. Determined viral proteins are able to change the host’s ubiquitin machinery and some viruses even encode their own ubiquitinating or deubiquitinating enzymes. African swine fever virus (ASFV) encodes a gene homologous to the E2 ubiquitin conjugating (UBC) enzyme. The viral ubiquitin-conjugating enzyme (UBCv1) is expressed throughout ASFV infection and accumulates at late times post infection. UBCv is also present in the viral particle suggesting that the ubiquitin-proteasome pathway could play an important role at early ASFV infection. We determined that inhibition of the final stage of the ubiquitin-proteasome pathway blocked a post-internalization step in ASFV replication in Vero cells. Under proteasome inhibition, ASF viral genome replication, late gene expression and viral production were severely reduced. Also, ASFV enhanced proteasome activity at late times and the accumulation of polyubiquitinated proteins surrounding viral factories. Core-associated and/or viral proteins involved in DNA replication may be targets for the ubiquitin-proteasome pathway that could possibly assist virus uncoating at final core breakdown and viral DNA release. At later steps, polyubiquitinated proteins at viral factories could exert regulatory roles in cell signaling.

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“…Previous studies have implicated the involvement of ubiquitin-proteasome system (UPS) in EV71's regulation of host factors. Some of these studies noted potent inhibition of EV71 by MG132 treatment 57 58 . Interestingly, these findings were demonstrated exclusively in EV71 infection of Vero cells.…”

Section: Discussionmentioning

confidence: 99%

Human genome-wide RNAi screen reveals host factors required for enterovirus 71 replication

Phuektes

Kumar

et al. 2016

Nat Commun

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Enterovirus 71 (EV71) is a neurotropic enterovirus without antivirals or vaccine, and its host-pathogen interactions remain poorly understood. Here we use a human genome-wide RNAi screen to identify 256 host factors involved in EV71 replication in human rhabdomyosarcoma cells. Enrichment analyses reveal overrepresentation in processes like mitotic cell cycle and transcriptional regulation. We have carried out orthogonal experiments to characterize the roles of selected factors involved in cell cycle regulation and endoplasmatic reticulum-associated degradation. We demonstrate nuclear egress of CDK6 in EV71 infected cells, and identify CDK6 and AURKB as resistance factors. NGLY1, which co-localizes with EV71 replication complexes at the endoplasmatic reticulum, supports EV71 replication. We confirm importance of these factors for EV71 replication in a human neuronal cell line and for coxsackievirus A16 infection. A small molecule inhibitor of NGLY1 reduces EV71 replication. This study provides a comprehensive map of EV71 host factors and reveals potential antiviral targets.

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Pyrrolidine dithiocarbamate inhibits enterovirus 71 replication by down-regulating ubiquitin–proteasome system

Cited by 15 publications

References 72 publications

SUMO Modification Stabilizes Enterovirus 71 Polymerase 3D To Facilitate Viral Replication

SUMO Modification Stabilizes Enterovirus 71 Polymerase 3D To Facilitate Viral Replication

The ubiquitin-proteasome system is required for African swine fever replication

Human genome-wide RNAi screen reveals host factors required for enterovirus 71 replication

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