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STUB1 is targeted by the SUMO-interacting motif of EBNA1 to maintain Epstein-Barr Virus latency.

ABSTRACT: Latent Epstein-Barr virus (EBV) infection is strongly associated with several malignancies, including B-cell lymphomas and epithelial tumors. EBNA1 is a key antigen expressed in all EBV-associated tumors during latency that is required for maintenance of the EBV episome DNA and the regulation of viral gene transcription. However, the mechanism utilized by EBV to maintain latent infection at the levels of posttranslational regulation remains largely unclear. Here, we report that EBNA1 contains two SUMO-interacting motifs (SIM2 and SIM3), and mutation of SIM2, but not SIM3, dramatically disrupts the EBNA1 dimerization, while SIM3 contributes to the polySUMO2 modification of EBNA1 at lysine 477 in vitro. Proteomic and immunoprecipitation analyses further reveal that the SIM3 motif is required for the EBNA1-mediated inhibitory effects on SUMO2-modified STUB1, SUMO2-mediated degradation of USP7, and SUMO1-modified KAP1. Deletion of the EBNASIM motif leads to functional loss of both EBNA1-mediated viral episome maintenance and lytic gene silencing. Importantly, hypoxic stress induces the SUMO2 modification of EBNA1, and in turn the dissociation of EBNA1 with STUB1, KAP1 and USP7 to increase the SUMO1 modification of both STUB1 and KAP1 for reactivation of lytic replication. Therefore, the EBNA1SIM motif plays an essential role in EBV latency and is a potential therapeutic target against EBV-associated cancers.

SUBMITTER: Wang Y

PROVIDER: S-EPMC7105294 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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STUB1 is targeted by the SUMO-interacting motif of EBNA1 to maintain Epstein-Barr Virus latency.

Wang Yuyan Y Du Shujuan S Zhu Caixia C Wang Chong C Yu Nuoya N Lin Ziqi Z Gan Jin J Guo Yi Y Huang Xinxin X He Yuping Y Robertson Erle E Qu Di D Wei Fang F Cai Qiliang Q

PLoS pathogens 20200316 3

Latent Epstein-Barr virus (EBV) infection is strongly associated with several malignancies, including B-cell lymphomas and epithelial tumors. EBNA1 is a key antigen expressed in all EBV-associated tumors during latency that is required for maintenance of the EBV episome DNA and the regulation of viral gene transcription. However, the mechanism utilized by EBV to maintain latent infection at the levels of posttranslational regulation remains largely unclear. Here, we report that EBNA1 contains tw ...[more]

PMID: 32176739

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Project description:The EBNA1 protein of Epstein-Barr virus (EBV) plays multiple roles in EBV latent infection, including altering cellular pathways relevant for cancer. Here we used microRNA (miRNA) cloning coupled with high-throughput sequencing to identify the effects of EBNA1 on cellular miRNAs in two nasopharyngeal carcinoma cell lines. EBNA1 affected a small percentage of cellular miRNAs in both cell lines, in particular, upregulating multiple let-7 family miRNAs, including let-7a. The effects of EBNA1 on let-7a were verified by demonstrating that EBNA1 silencing in multiple EBV-positive carcinomas downregulated let-7a. Accordingly, the let-7a target, Dicer, was found to be partially downregulated by EBNA1 expression (at the mRNA and protein levels) and upregulated by EBNA1 silencing in EBV-positive cells. Reporter assays based on the Dicer 3' untranslated region with and without let-7a target sites indicated that the effects of EBNA1 on Dicer were mediated by let-7a. EBNA1 was also found to induce the expression of let-7a primary RNAs in a manner dependent on the EBNA1 transcriptional activation region, suggesting that EBNA1 induces let-7a by transactivating the expression of its primary transcripts. Consistent with previous reports that Dicer promotes EBV reactivation, we found that a let-7a mimic inhibited EBV reactivation to the lytic cycle, while a let-7 sponge increased reactivation. The results provide a mechanism by which EBNA1 could promote EBV latency by inducing let-7 miRNAs.The EBNA1 protein of Epstein-Barr virus (EBV) contributes in multiple ways to the latent mode of EBV infection that leads to lifelong infection. In this study, we identify a mechanism by which EBNA1 helps to maintain EBV infection in a latent state. This involves induction of a family of microRNAs (let-7 miRNAs) that in turn decreases the level of the cellular protein Dicer. We demonstrate that let-7 miRNAs inhibit the reactivation of latent EBV, providing an explanation for our previous observation that EBNA1 promotes latency. In addition, since decreased levels of Dicer have been associated with metastatic potential, EBNA1 may increase metastases by downregulating Dicer.

Project description:Epstein Barr virus (EBV) is a potentially oncogenic gammaherpesvirus that establishes a chronic, latent infection in memory B cells. The EBV genome persists in infected host cells as a chromatinized episome and is subject to chromatin-mediated regulation. Binding of the host insulator protein CTCF to the EBV genome has an established role in maintaining viral latency type. CTCF is posttranslationally modified by the host enzyme PARP1. PARP1, or poly(ADP-ribose) polymerase 1, catalyzes the transfer of a poly(ADP-ribose) (PAR) moiety from NAD+ onto acceptor proteins, including itself, histone proteins, and CTCF. PARylation of CTCF by PARP1 can affect CTCF's insulator activity, DNA binding capacity, and ability to form chromatin loops. Both PARP1 and CTCF have been implicated in the regulation of EBV latency and lytic reactivation. Thus, we predicted that pharmacological inhibition with PARP1 inhibitors would affect EBV latency type through a chromatin-specific mechanism. Here, we show that PARP1 and CTCF colocalize at specific sites throughout the EBV genome and provide evidence to suggest that PARP1 acts to stabilize CTCF binding and maintain the open chromatin landscape at the active Cp promoter during type III latency. Further, PARP1 activity is important in maintaining latency type-specific viral gene expression. The data presented here provide a rationale for the use of PARP inhibitors in the treatment of EBV-associated cancers exhibiting type III latency and ultimately could contribute to an EBV-specific treatment strategy for AIDS-related or posttransplant lymphomas.IMPORTANCE EBV is a human gammaherpesvirus that infects more than 95% of individuals worldwide. Upon infection, EBV circularizes as an episome and establishes a chronic, latent infection in B cells. In doing so, the virus utilizes host cell machinery to regulate and maintain the viral genome. In otherwise healthy individuals, EBV infection is typically nonpathological; however, latent infection is potentially oncogenic and is responsible for 1% of human cancers. During latent infection, EBV expresses specific sets of proteins according to the given latency type, each of which is associated with specific types of cancers. For example, type III latency, in which the virus expresses its full repertoire of latent proteins, is characteristic of AIDS-associated and posttransplant lymphomas associated with EBV infection. Understanding how viral latency type is regulated at the chromatin level may reveal potential targets for EBV-specific pharmacological intervention in EBV-associated cancers.

Dataset Information

STUB1 is targeted by the SUMO-interacting motif of EBNA1 to maintain Epstein-Barr Virus latency.

Publications

STUB1 is targeted by the SUMO-interacting motif of EBNA1 to maintain Epstein-Barr Virus latency.

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