SCE1, the SUMO-Conjugating Enzyme in Plants That Interacts with NIb, the RNA-Dependent RNA Polymerase of Turnip Mosaic Virus, Is Required for Viral Infection

Xiong, Ruyi; Wang, Aiming

doi:10.1128/jvi.02828-12

Cited by 79 publications

(87 citation statements)

References 53 publications

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“…Two days post infiltration, the infiltrated leaf area was observed under a confocal microscope. The fluorescent signal from NIb-CFP was found mainly in the nucleus (data not shown), consistent with our observation in previous publications (Wei et al, 2010a;Xiong and Wang, 2013). In the cytoplasm, the NIb signal was found in the large irregularly shaped structure and co-localized with the fluorescence from B2-YN/VP35-YC (Fig.…”

Section: Basic Strategysupporting

confidence: 94%

Visualizing double-stranded RNA distribution and dynamics in living cells by dsRNA binding-dependent fluorescence complementation

et al. 2015

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Double-stranded RNA (dsRNA) is an important type of RNA that plays essential roles in diverse cellular processes in eukaryotic organisms and a hallmark in infections by positive-sense RNA viruses. Currently, no in vivo technology has been developed for visualizing dsRNA in living cells. Here, we report a dsRNA binding-dependent fluorescence complementation (dRBFC) assay that can be used to efficiently monitor dsRNA distribution and dynamics in vivo. The system consists of two dsRNA-binding proteins, which are fused to the N- and C-terminal halves of the yellow fluorescent protein (YFP). Binding of the two fusion proteins to a common dsRNA brings the split YFP halves in close proximity, leading to the reconstitution of the fluorescence-competent structure and restoration of fluorescence. Using this technique, we were able to visualize the distribution and trafficking of the replicative RNA intermediates of positive-sense RNA viruses in living cells.

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Section: Basic Strategysupporting

confidence: 94%

Visualizing double-stranded RNA distribution and dynamics in living cells by dsRNA binding-dependent fluorescence complementation

et al. 2015

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“…Only EV71 3C protease is SUMOylated, while coxsackievirus B5 is involved in the host cell SUMOylation system in the family Picornaviridae (38,62,70). Current understanding of viral polymerase SUMOylation is limited to RdRp of Turnip mosaic virus (71), DNA polymerase subunit UL44 of HCMV (25), nonstructural protein 5 (NS5) of dengue virus (72), and polymerase basic protein 1 (PB1) of influenza virus (30). In the currently study, we reveal that the 3D protein of EV71, an RNA-dependent RNA polymerase, is SUMOylated.…”

Section: Discussionmentioning

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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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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“…A Y2H assay was performed as described previously (87). Yeast expression plasmids were introduced into yeast strain AH109 (Clontech) by lithium acetate-mediated transformation.…”

Section: Methodsmentioning

confidence: 99%

The Potyvirus Silencing Suppressor Protein VPg Mediates Degradation of SGS3 via Ubiquitination and Autophagy Pathways

Cheng

Wang

2017

J Virol

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RNA silencing is an innate antiviral immunity response of plants and animals. To counteract this host immune response, viruses have evolved an effective strategy to protect themselves by the expression of viral suppressors of RNA silencing (VSRs). Most potyviruses encode two VSRs, helper component-proteinase (HCPro) and viral genome-linked protein (VPg). The molecular biology of the former has been well characterized, whereas how VPg exerts its function in the suppression of RNA silencing is yet to be understood. In this study, we show that infection by Turnip mosaic virus (TuMV) causes reduced levels of suppressor of gene silencing 3 (SGS3), a key component of the RNA silencing pathway that functions in doublestranded RNA synthesis for virus-derived small interfering RNA (vsiRNA) production. We also demonstrate that among 11 TuMV-encoded viral proteins, VPg is the only one that interacts with SGS3. We furthermore present evidence that the expression of VPg alone, independent of viral infection, is sufficient to induce the degradation of SGS3 and its intimate partner RNA-dependent RNA polymerase 6 (RDR6). Moreover, we discover that the VPg-mediated degradation of SGS3 occurs via both the 20S ubiquitin-proteasome and autophagy pathways. Taken together, our data suggest a role for VPg-mediated degradation of SGS3 in suppression of silencing by VPg.IMPORTANCE Potyviruses represent the largest group of known plant viruses and cause significant losses of many agriculturally important crops in the world. In order to establish infection, potyviruses must overcome the host antiviral silencing response. A viral protein called VPg has been shown to play a role in this process, but how it works is unclear. In this paper, we found that the VPg protein of Turnip mosaic virus (TuMV), which is a potyvirus, interacts with a host protein named SGS3, a key protein in the RNA silencing pathway. Moreover, this interaction leads to the degradation of SGS3 and its interacting and functional partner RDR6, which is another essential component of the RNA silencing pathway. We also identified the cellular pathways that are recruited for the VPg-mediated degradation of SGS3. Therefore, this work reveals a possible mechanism by which VPg sabotages host antiviral RNA silencing to promote virus infection. KEYWORDS RDR6, RNA silencing, SGS3, VPg, VSR, autophagy, potyvirus, ubiquitination T o establish systemic infection, an invading virus must remodel and recruit host elements and overcome resistance responses through complex molecular virushost interactions (1-3). One such essential antiviral response is RNA silencing (4-6), a cellular mechanism conserved from lower eukaryotes to mammals to regulate endogenous gene expression at the transcriptional and posttranscriptional levels (7, 8). To activate antiviral RNA silencing, the viral double-stranded RNAs (dsRNAs), which may

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SCE1, the SUMO-Conjugating Enzyme in Plants That Interacts with NIb, the RNA-Dependent RNA Polymerase of Turnip Mosaic Virus, Is Required for Viral Infection

Cited by 79 publications

References 53 publications

Visualizing double-stranded RNA distribution and dynamics in living cells by dsRNA binding-dependent fluorescence complementation

Visualizing double-stranded RNA distribution and dynamics in living cells by dsRNA binding-dependent fluorescence complementation

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

The Potyvirus Silencing Suppressor Protein VPg Mediates Degradation of SGS3 via Ubiquitination and Autophagy Pathways

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