Severe fever with thrombocytopenia syndrome virus (SFTSV) and Rift Valley fever virus (RVFV) are two arthropod-borne phleboviruses in the Bunyaviridae family, which cause severe illness in humans and animals. Glycoprotein N (Gn) is one of the envelope proteins on the virus surface and is a major antigenic component. Despite its importance for virus entry and fusion, the molecular features of the phleboviruse Gn were unknown. Here, we present the crystal structures of the Gn head domain from both SFTSV and RVFV, which display a similar compact triangular shape overall, while the three subdomains (domains I, II, and III) making up the Gn head display different arrangements. Ten cysteines in the Gn stem region are conserved among phleboviruses, four of which are responsible for Gn dimerization, as revealed in this study, and they are highly conserved for all members in Bunyaviridae. Therefore, we propose an anchoring mode on the viral surface. The complex structure of the SFTSV Gn head and human neutralizing antibody MAb 4-5 reveals that helices α6 in subdomain III is the key component for neutralization. Importantly, the structure indicates that domain III is an ideal region recognized by specific neutralizing antibodies, while domain II is probably recognized by broadly neutralizing antibodies. Collectively, Gn is a desirable vaccine target, and our data provide a molecular basis for the rational design of vaccines against the diseases caused by phleboviruses and a model for bunyavirus Gn embedding on the viral surface.bunyavirus | SFTSV | glycoprotein | neutralizing antibody | RVFV

EBV infection in tumor cells is generally restricted to the latent forms of viral infection. Switching the latent form of viral infection into the lytic form may induce tumor cell death. We have previously reported that certain chemotherapy agents can increase the amount of lytic viral gene expression in EBV-positive tumor cells. In this report, we have explored the potential utility of valproic acid (VPA), an anti-seizure drug that also has strong histone deacetylase inhibitory activity, for activating lytic viral gene expression in EBVpositive tumors. Although VPA treatment alone induced only a modest increase in the level of lytic viral gene expression, it strongly enhanced the ability of chemotherapeutic agents to induce lytic EBV gene expression in EBV-positive epithelial and lymphoid cells in vitro. Furthermore, VPA enhanced cell killing in vitro by chemotherapeutic agents in lymphoblastoid cells and gastric cells (AGS) containing wild-type EBV. In contrast, VPA did not enhance the cytotoxicity of chemotherapy in lymphoblastoid cells containing a lytic-defective (BZLF1-knockout) form of EBV or in EBV-negative AGS cells. Finally, we found that the combination of VPA and chemotherapy was significantly more effective in inhibiting EBVdriven lymphoproliferative disease in severe combined immunodeficient mice than chemotherapy alone. These results suggest that VPA could potentiate the efficacy of chemotherapy for EBV-positive tumors in patients.

Epstein-Barr virus (EBV) establishes a latent form of infection in memory B cells, while antibody-secreting plasma cells often harbor the lytic form of infection. The switch between latent and lytic EBV infection is mediated by the two viral immediate-early proteins BZLF1 (Z) and BRLF1 (R), which are not expressed in latently infected B cells. Here we demonstrate that a cellular transcription factor that plays an essential role in plasma cell differentiation, X-box-binding protein 1 (XBP-1), also activates the transcription of the two EBV immediate-early gene promoters. In reporter gene assays, XBP-1 alone was sufficient to activate the R promoter, whereas the combination of XBP-1 and protein kinase D (PKD) was required for efficient activation of the Z promoter. Most importantly, the expression of XBP-1 and activated PKD was sufficient to induce lytic viral gene expression in EBV-positive nasopharyngeal carcinoma cells and lymphoblastoid cells, while an XBP-1 small interfering RNA inhibited constitutive lytic EBV gene expression in lymphoblastoid cells. These results suggest that the plasma cell differentiation factor XBP-1, in combination with activated PKD, can mediate the reactivation of EBV, thereby allowing the viral life cycle to be intimately linked to plasma cell differentiation. Epstein-Barr virus (EBV)is the causative agent of infectious mononucleosis and is associated with B-cell lymphomas, nasopharyngeal carcinomas, gastric carcinomas, and other malignancies (26,45). EBV causes lytic infection in normal oral epithelial cells (32, 51) while usually establishing one of the latent forms of infection in circulating memory B cells. In contrast, tonsillar B cells that express antigens specific for plasma markers commonly harbor the lytic form of EBV infection, which results in the production of infectious viral particles (10,29,30).The switch from latent to lytic EBV infection is mediated by the immediate-early (IE) protein BZLF1 (Z) and the immediate-early/early protein BRLF1 (R) (1,16,57). Z and R are transcription factors that activate each other's transcription and together are sufficient to activate the entire lytic viral gene expression cascade (17,49). In latently infected cells, the promoters driving Z and R expression (Zp and Rp) are inactive. Therefore, the activation of Zp and Rp by cellular transcription factors is the crucial initial step required for lytic viral gene expression. B-cell receptor engagement activates lytic EBV gene expression in some B-cell lines in vitro and activates both EBV IE promoters in reporter gene assays (23). Although several different individual cellular transcription factors can activate one or both of the two EBV IE promoters in reporter gene assays (23), to date these factors have not been shown to be sufficient for the efficient reactivation of lytic viral gene expression from the endogenous viral genome in latently infected cells.While there is a strong correlation between plasma cell differentiation and lytic EBV gene expression in human tonsils, it is not pr...

Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important viral pathogens in the swine industry. Emerging evidence indicates that the host microRNAs (miRNAs) are involved in host-pathogen interactions. However, whether host miRNAs can target PRRSV and be used to inhibit PRRSV infection has not been reported. Recently, microRNA 181 (miR-181) has been identified as a positive regulator of immune response, and here we report that miR-181 can directly impair PRRSV infection. Our results showed that delivered miR-181 mimics can strongly inhibit PRRSV replication in vitro through specifically binding to a highly (over 96%) conserved region in the downstream of open reading frame 4 (ORF4) of the viral genomic RNA. The inhibition of PRRSV replication was specific and dose dependent. In PRRSV-infected Marc-145 cells, the viral mRNAs could compete with miR-181-targeted sequence in luciferase vector to interact with miR-181 and result in less inhibition of luciferase activity, further demonstrating the specific interactions between miR-181 and PRRSV RNAs. As expected, miR-181 and other potential PRRSV-targeting miRNAs (such as miR-206) are expressed much more abundantly in minimally permissive cells or tissues than in highly permissive cells or tissues. Importantly, highly pathogenic PRRSV (HP-PRRSV) strain-infected pigs treated with miR-181 mimics showed substantially decreased viral loads in blood and relief from PRRSV-induced fever compared to negative-control (NC)-treated controls. These results indicate the important role of host miRNAs in modulating PRRSV infection and viral pathogenesis and also support the idea that host miRNAs could be useful for RNA interference (RNAi)-mediated antiviral therapeutic strategies. P orcine reproductive and respiratory syndrome virus (PRRSV) is one of the most economically important viral pathogens in pigs, leading to significant economic losses in the swine industry worldwide. PRRS is characterized by severe reproductive failure in sows and respiratory syndromes and persistent infection in young pigs. Atypical PRRS is characterized by high fever, high morbidity, and high mortality in pigs of all ages and emerged in China in 2006. The causative agent was confirmed to be a highly pathogenic PRRSV (HP-PRRSV) with a discontinuous deletion of 30 amino acids in nonstructural protein 2 (NSP2) (1). PRRSV is classified within the family Arteriviridae, order Nidovirales. It has a singlestranded, 5=-capped positive-sense RNA genome of approximately 15.4 kb, containing at least 9 opening reading frames (ORFs) (1, 2). MicroRNAs (miRNAs), which can be produced by hosts or viruses, are a class of small noncoding RNAs that can inhibit gene expression through base-pairing interactions between the loaded miRNA and its mRNA target. Recent studies have reported that cellular miRNAs can target viral RNAs during infections, resulting in inhibition of virus replication as a new antiviral defense (3-8) or a new pathway to alter the virus life cycle (9-11), whereas virusderive...

All positive-stranded RNA viruses with genomes >∼7 kb encode helicases, which generally are poorly characterized. The core of the nidovirus superfamily 1 helicase (HEL1) is associated with a unique N-terminal zinc-binding domain (ZBD) that was previously implicated in helicase regulation, genome replication and subgenomic mRNA synthesis. The high-resolution structure of the arterivirus helicase (nsp10), alone and in complex with a polynucleotide substrate, now provides first insights into the structural basis for nidovirus helicase function. A previously uncharacterized domain 1B connects HEL1 domains 1A and 2A to a long linker of ZBD, which further consists of a novel RING-like module and treble-clef zinc finger, together coordinating three Zn atoms. On substrate binding, major conformational changes were evident outside the HEL1 domains, notably in domain 1B. Structural characterization, mutagenesis and biochemistry revealed that helicase activity depends on the extensive relay of interactions between the ZBD and HEL1 domains. The arterivirus helicase structurally resembles the cellular Upf1 helicase, suggesting that nidoviruses may also use their helicases for post-transcriptional quality control of their large RNA genomes.