Publications
164 results found
Farrell PJ, 2023, EBV and MS: the evidence is growing stronger, Cell, Vol: 186, Pages: 5675-5676, ISSN: 0092-8674
Autoimmune cross-reaction of specific antibodies to Epstein-Barr virus (EBV) has been associated with development of multiple sclerosis (MS). In this issue of Cell, Vietzen et al. identify additional immunological regulatory mechanisms that protect against autoimmunity in healthy people but are reduced in MS cases. The results confirm the link between EBV and MS.
Wang Y, Ungerleider N, Hoffman BA, et al., 2022, A polymorphism in the Epstein-Barr virus EBER2 noncoding RNA drives in vivo expansion of latently infected B cells, mBio, Vol: 13, ISSN: 2150-7511
The oncogenic gammaherpesviruses, including human Epstein-Barr virus (EBV), human Kaposi’s sarcoma-associated herpesvirus (KSHV), and murine gammaherpesvirus 68 (MHV68, γHV68, MuHV-4), are associated with numerous malignancies, including B cell lymphomas and nasopharyngeal carcinoma. These viruses employ numerous molecular strategies to colonize the host, including the expression of noncoding RNAs (ncRNAs). As the first viral ncRNAs identified, EBV-encoded RNA 1 and 2 (EBER1 and EBER2, respectively) have been investigated extensively for decades; however, their specific in vivo functions remain largely unknown. In work here, we used chimeric MHV68 viruses in an in vivo complementation system to test whether EBV EBER2 contributes to acute and/or chronic phases of infection. Expression of EBER2 derived from EBV strain B95-8 resulted in a significant expansion of latently infected B cells in vivo, which was accompanied by a decrease in virus-infected plasma cells. EBV strains typically carry one of two variants of EBER2, which differ primarily by a 5-nucleotide core polymorphism identified initially in the EBV strain M81. Strikingly, mutation of the 5 nucleotides that define this core polymorphism resulted in the loss of the infected B cell expansion and restored plasma cell infection. This work reveals that the B95-8 variant of EBER2 promotes the expansion of the latently infected B cell pool in vivo and may do so in part through inhibition of terminal differentiation. These findings provide new insight into mechanisms by which viral ncRNAs promote in vivo colonization and further and provide further evidence of the inherent tumorigenic risks associated with gammaherpesvirus manipulation of B cell differentiation.
Wongwiwat W, Fournier B, Bassano I, et al., 2022, Epstein-Barr virus genome deletions in Epstein-Barr virus-positive T/NK cell lymphoproliferative diseases, Journal of Virology, Vol: 96, Pages: 1-15, ISSN: 0022-538X
The main target cells for Epstein-Barr virus (EBV) infection and persistence are B lymphocytes, although T and NK cells can also become infected. In this paper we characterise the EBV present in 21 pediatric and adult patients treated in France for a range of diseases that involve infection of T or NK cells. Of these 21 cases, 5 pediatric patients (21%) and 11 adult patients (52%) are of Caucasian origin. In about 30% of the cases, some of the EBV genomes contain a large deletion. The deletions are different in every patient but tend to cluster near the BART region of the viral genome. Detailed investigation of a family, in which several members have persistent T or NK cell infection by EBV, indicates that the virus genome deletions arise or are selected independently in each individual patient. Genome sequence polymorphisms in the EBV in these T or NK cell diseases reflect the geographic origin of the patient, not a distinct type of EBV (the 21 cases studied included examples of both type 1 and type 2 EBV infection). Using virus produced from type 1 or type 2 EBV genomes cloned in bacterial artificial chromosome (BAC) vectors, we demonstrate infection of T cells in cord blood from healthy donors. Our results are consistent with transient infection of some T cells being part of normal asymptomatic infection by EBV in young children.
Farrell PJ, White RE, 2022, Do Epstein-Barr virus mutations and natural genome sequence variations contribute to disease?, Biomolecules, Vol: 12, ISSN: 2218-273X
Most of the world’s population is infected by the Epstein–Barr virus (EBV), but the incidence of the diseases associated with EBV infection differs greatly in different parts of the world. Many factors may determine those differences, but variation in the virus genome is likely to be a contributing factor for some of the diseases. Here, we describe the main forms of EBV genome sequence variation, and the mechanisms by which variations in the virus genome are likely to contribute to disease. EBV genome deletions or polymorphisms can also provide useful markers for monitoring disease. If some EBV strains prove to be more pathogenic than others, this suggests the possible value of immunising people against infection by those pathogenic strains
Farrell PJ, 2021, Iron metabolism as a novel therapeutic target, Blood, Vol: 138, Pages: 2155-2156, ISSN: 0006-4971
Venturini C, Houldcroft CJ, Lazareva A, et al., 2021, Epstein-Barr virus (EBV) deletions as biomarkers of response to treatment of chronic active EBV, British Journal of Haematology, Vol: 195, Pages: 249-255, ISSN: 0007-1048
Epstein–Barr virus (EBV) infects >95% of the population worldwide.1 A small number of patients develop life-threatening persistence of high-level EBV replication following an infectious mononucleosis syndrome, often associated with splenomegaly and hepatitis.2 Chronic active EBV (CAEBV) disease is characterised by infiltration of tissues by EBV+ T, natural killer (NK) or less frequently B cells and can progress into lymphoproliferative disease. Clonal expansion of EBV-infected T or NK cells is well described.3To date, CAEBV has been mostly described in Asian or South/Central American patients.4 Frequent deletions in the EBV genome have been found in samples from Japanese CAEBV patients (35%) and other EBV-driven neoplasms.5-7 However, it is unclear if these deletions are specific for Asian EBV strains, whether they are present in viral genomes in different sites and how they evolve overtime.In the present study, we sequenced EBV from serial blood samples obtained from three UK patients with CAEBV disease. The results were compared with sequences from saliva of patients with CAEBV and blood and tissue from other benign and malignant EBV-related conditions.
Su C, Lu F, Soldan SS, et al., 2021, EBNA2 driven enhancer switching at the CIITA-DEXI locus suppresses HLA class II gene expression during EBV infection of B-lymphocytes, PLoS Pathogens, Vol: 17, ISSN: 1553-7366
Viruses suppress immune recognition through diverse mechanisms. Epstein-Barr Virus (EBV) establishes latent infection in memory B-lymphocytes and B-cell malignancies where it impacts B-cell immune function. We show here that EBV primary infection of naïve B-cells results in a robust down-regulation of HLA genes. We found that the viral encoded transcriptional regulatory factor EBNA2 bound to multiple regulatory regions in the HLA locus. Conditional expression of EBNA2 correlated with the down regulation of HLA class II transcription. EBNA2 down-regulation of HLA transcription was found to be dependent on CIITA, the major transcriptional activator of HLA class II gene transcription. We identified a major EBNA2 binding site downstream of the CIITA gene and upstream of DEXI, a dexamethasone inducible gene that is oriented head-to-head with CIITA gene transcripts. CRISPR/Cas9 deletion of the EBNA2 site upstream of DEXI attenuated CIITA transcriptional repression. EBNA2 caused an increase in DEXI transcription and a graded change in histone modifications with activation mark H3K27ac near the DEXI locus, and a loss of activation marks at the CIITA locus. A prominent CTCF binding site between CIITA and DEXI enhancers was mutated and further diminished the effects of EBNA2 on CIITA. Analysis of HiC data indicate that DEXI and CIITA enhancers are situated in different chromosome topological associated domains (TADs). These findings suggest that EBNA2 down regulates HLA-II genes through the down regulation of CIITA, and that this down regulation is an indirect consequence of EBNA2 enhancer formation at a neighboring TAD. We propose that enhancer competition between these neighboring chromosome domains represents a novel mechanism for gene regulation demonstrated by EBNA2.
Lippert T, Marzec P, Idilli A, et al., 2021, Oncogenic herpesvirus KSHV triggers hallmarks of alternative lengthening of telomeres, Nature Communications, Vol: 12, ISSN: 2041-1723
To achieve replicative immortality, cancer cells must activate telomere maintenance mechanisms to prevent telomere shortening. ~85% of cancers circumvent telomeric attrition by re-expressing telomerase, while the remaining ~15% of cancers induce alternative lengthening of telomeres (ALT), which relies on break-induced replication (BIR) and telomere recombination. Although ALT tumours were first reported over 20 years ago, the mechanism of ALT induction remains unclear and no study to date has described a cell-based model that permits the induction of ALT. Here, we demonstrate that infection with Kaposi’s sarcoma herpesvirus (KSHV) induces sustained acquisition of ALT-like features in previously non-ALT cell lines. KSHV-infected cells acquire hallmarks of ALT activity that are also observed in KSHV-associated tumour biopsies. Down-regulating BIR impairs KSHV latency, suggesting that KSHV co-opts ALT for viral functionality. This study uncovers KSHV infection as a means to study telomere maintenance by ALT and reveals features of ALT in KSHV-associated tumours.
Zha S, Chau H-F, Chau WY, et al., 2021, Dual-targeting peptide-guided approach for precision delivery and cancer monitoring by using a safe upconversion nanoplatform, Advanced Science, Vol: 8, Pages: 1-15, ISSN: 2198-3844
Using Epstein‐Barr virus (EBV)‐induced cancer cells and HeLa cells as a comparative study model, a novel and safe dual‐EBV‐oncoproteins‐targeting pH‐responsive peptide engineering, coating, and guiding approach to achieve precision targeting and treatment strategy against EBV‐associated cancers is introduced. Individual functional peptide sequences that specifically bind to two overexpressed EBV‐specific oncoproteins, EBNA1 (a latent cellular protein) and LMP1 (a transmembrane protein), are engineered in three different ways and incorporated with a pH‐sensitive tumor microenvironment (TME)‐cleavable linker onto the upconversion nanoparticles (UCNP) NaGdF4:Yb3+, Er3+@NaGdF4 (UCNP‐Pn, n = 5, 6, and 7). A synergistic combination of the transmembrane LMP1 targeting ability and the pH responsiveness of UCNP‐Pn is found to give specific cancer differentiation with higher cellular uptake and accumulation in EBV‐infected cells, thus a lower dose is needed and the side effects and health risks from treatment would be greatly reduced. It also gives responsive UC signal enhancement upon targeted dual‐protein binding and shows efficacious EBV cancer inhibition in vitro and in vivo. This is the first example of simultaneous imaging and inhibition of two EBV latent proteins, and serves as a blueprint for next‐generation peptide‐guided precision delivery nanosystem for the safe monitoring and treatment against one specific cancer.
Romero-Masters JC, Huebner SM, Ohashi M, et al., 2020, B cells infected with Type 2 Epstein-Barr virus (EBV) have increased NFATc1/NFATc2 activity and enhanced lytic gene expression in comparison to Type 1 EBV infection, PLoS Pathogens, Vol: 16, ISSN: 1553-7366
Humans are infected with two distinct strains (Type 1 (T1) and Type 2 (T2)) of Epstein-Barr virus (EBV) that differ substantially in their EBNA2 and EBNA 3A/B/C latency genes and the ability to transform B cells in vitro. While most T1 EBV strains contain the “prototype” form of the BZLF1 immediate-early promoter (“Zp-P”), all T2 strains contain the “Zp-V3” variant, which contains an NFAT binding motif and is activated much more strongly by B-cell receptor signalling. Whether B cells infected with T2 EBV are more lytic than cells infected with T1 EBV is unknown. Here we show that B cells infected with T2 EBV strains (AG876 and BL5) have much more lytic protein expression compared to B cells infected with T1 EBV strains (M81, Akata, and Mutu) in both a cord blood-humanized (CBH) mouse model and EBV-transformed lymphoblastoid cell lines (LCLs). Although T2 LCLs grow more slowly than T1 LCLs, both EBV types induce B-cell lymphomas in CBH mice. T1 EBV strains (M81 and Akata) containing Zp-V3 are less lytic than T2 EBV strains, suggesting that Zp-V3 is not sufficient to confer a lytic phenotype. Instead, we find that T2 LCLs express much higher levels of activated NFATc1 and NFATc2, and that cyclosporine (an NFAT inhibitor) and knockdown of NFATc2 attenuate constitutive lytic infection in T2 LCLs. Both NFATc1 and NFATc2 induce lytic EBV gene expression when combined with activated CAMKIV (which is activated by calcium signaling and activates MEF2D) in Burkitt Akata cells. Together, these results suggest that B cells infected with T2 EBV are more lytic due to increased activity of the cellular NFATc1/c2 transcription factors in addition to the universal presence of the Zp-V3 form of BZLF1 promoter.
Ponnusamy R, Khatri R, Correia PB, et al., 2019, Increased association between Epstein-Barr virus EBNA2 from type 2 strains and the transcriptional repressor BS69 restricts EBNA2 activity, PLoS Pathogens, Vol: 15, ISSN: 1553-7366
Natural variation separates Epstein-Barr virus (EBV) into type 1 and type 2 strains. Type 2 EBV is less transforming in vitro due to sequence differences in the EBV transcription factor EBNA2. This correlates with reduced activation of the EBV oncogene LMP1 and some cell genes. Transcriptional activation by type 1 EBNA2 can be suppressed through the binding of two PXLXP motifs in its transactivation domain (TAD) to the dimeric coiled-coil MYND domain (CC-MYND) of the BS69 repressor protein (ZMYND11). We identified a third conserved PXLXP motif in type 2 EBNA2. We found that type 2 EBNA2 peptides containing this motif bound BS69CC-MYND efficiently and that the type 2 EBNA2TAD bound an additional BS69CC-MYND molecule. Full-length type 2 EBNA2 also bound BS69 more efficiently in pull-down assays. Molecular weight analysis and low-resolution structures obtained using small-angle X-ray scattering showed that three BS69CC-MYND dimers bound two molecules of type 2 EBNA2TAD, in line with the dimeric state of full-length EBNA2 in vivo. Importantly, mutation of the third BS69 binding motif in type 2 EBNA2 improved B-cell growth maintenance and the transcriptional activation of the LMP1 and CXCR7 genes. Our data indicate that increased association with BS69 restricts the function of type 2 EBNA2 as a transcriptional activator and driver of B cell growth and may contribute to reduced B-cell transformation by type 2 EBV.
Alizon S, Bravo IG, Farrell PJ, et al., 2019, Towards a multi-level and a multi-disciplinary approach to DNA oncovirus virulence, PHILOSOPHICAL TRANSACTIONS OF THE ROYAL SOCIETY B-BIOLOGICAL SCIENCES, Vol: 374, ISSN: 0962-8436
Bridges R, Correia S, Wegner F, et al., 2019, Essential role of inverted repeat in Epstein–Barr virus IR-1 in B cell transformation; geographical variation of the viral genome, Philosophical Transactions B: Biological Sciences, Vol: 374, ISSN: 0962-8436
Many regions of the Epstein‐Barr virus (EBV) genome, repeated and unique sequences, contribute to the geographic variation observed between strains. Here we use a large alignment of curated EBV genome sequences to identify major sites of variation in the genome of type 1 EBV strains; the CAO deletion in latent membrane protein 1 (LMP1) is the most frequent major indel present in the unique regions of EBV strains from various parts of the world. Principal component analysis was used to identify patterns of sequence variation and nucleotide positions in the sequences which can distinguish EBV from some different geographic regions. Viral genome sequence variation also affects interpretation of genetic content; known genes, origins of replication and gene expression control regions explain most of the viral genome but there are still a few sections of unknown function. One of these EBV genome regions contains a large inverted repeat sequence (invR) within the IR‐1 major internal repeat array. We deleted this invR sequence and showed that this abolished the ability of the virus to transform human B cells into lymphoblastoid cell lines.
Farrell P, Paschos K, 2019, Requirement for PRC1 subunit BMI1 in host gene activation by Epstein-Barr virus potein EBNA3C, Nucleic Acids Research, Vol: 47, Pages: 2807-2821, ISSN: 0305-1048
Epstein–Barr virus proteins EBNA3A, EBNA3B and EBNA3C control hundreds of host genes after infection. Changes in epigenetic marks around EBNA3-regulated genes suggest that they exert transcriptional control in collaboration with epigenetic factors. The roles of polycomb repressive complex (PRC)2 subunit SUZ12 and of PRC1 subunit BMI1 were assessed for their importance in EBNA3-mediated repression and activation. ChIP-seq experiments for SUZ12 and BMI1 were performed to determine their global localization on chromatin and analysis offered further insight into polycomb protein distribution in differentiated cells. Their localization was compared to that of each EBNA3 to resolve longstanding questions about the EBNA3–polycomb relationship. SUZ12 did not co-localize with any EBNA3, whereas EBNA3C co-localized significantly and co-immunoprecipitated with BMI1. In cells expressing a conditional EBNA3C, BMI1 was sequestered to EBNA3C-binding sites after EBNA3C activation. When SUZ12 or BMI1 was knocked down in the same cells, SUZ12 did not contribute to EBNA3C-mediated regulation. Surprisingly, after BMI1 knockdown, EBNA3C repressed equally efficiently but host gene activation by EBNA3C was impaired. This overturns previous assumptions about BMI1/PRC1 functions during EBNA3C-mediated regulation, for the first time identifies directly a host factor involved in EBNA3-mediated activation and provides a new insight into how PRC1 can be involved in gene activation.
Correia S, Bridges R, Wegner F, et al., 2018, Sequence variation of Epstein-Barr virus: viral types, geography, codon usage and diseases, Journal of Virology, Vol: 92, ISSN: 1098-5514
138 new Epstein-Barr virus (EBV) genome sequences have been determined. 125 of these and 116 from previous reports were combined to produce a multiple sequence alignment of 241 EBV genomes, which we have used to analyze variation within the viral genome. The type 1/type2 classification of EBV remains the major form of variation and is defined mostly by EBNA2 and EBNA3, but the type 2 SNPs at the EBNA3 locus extend into the adjacent gp350 and gp42 genes, whose products mediate infection of B cells by EBV. A small insertion within the BART miRNA region of the genome was present in 21 EBV strains. EBV from saliva of USA patients with chronic active EBV infection aligned with the wild type EBV genome, with no evidence of WZhet rearrangements. The V3 polymorphism in the Zp promoter for BZLF1 was found to be frequent in nasopharyngeal carcinoma cases both from Hong Kong and Indonesia. Codon usage was found to differ between latent and lytic cycle EBV genes and the main forms of variation of the EBNA1 protein have been identified.IMPORTANCE Epstein-Barr virus causes most cases of infectious mononucleosis and post-transplant lymphoproliferative disease. It contributes to several types of cancer including Hodgkin's lymphoma, Burkitt's lymphoma, diffuse large B cell lymphoma, nasopharyngeal carcinoma and gastric carcinoma. EBV genome variation is important because some of the diseases associated with EBV have very different incidences in different populations and geographic regions - differences in the EBV genome might contribute to these diseases. Some specific EBV genome alterations that appear to be significant in EBV associated cancers are already known and current efforts to make an EBV vaccine and antiviral drugs should also take account of sequence differences in the proteins used as targets.
Farrell PJ, 2018, Epstein‐Barr virus and cancer, Annual Review of Pathology: Mechanisms of Disease, ISSN: 1553-4014
Epstein‐Barr virus (EBV) contributes to about 1.5% of all cases of human cancer worldwide and viral genes are expressed in the malignant cells. EBV also very efficiently causes proliferation of infected human B lymphocytes. The functions of the viral proteins and small RNAs that may contribute to the EBV associated cancers are becoming increasingly clear and a broader understanding of sequence variation of the virus genome has helped to interpret their roles. The improved understanding of the mechanisms of these cancers means that the EBV associated cancers offer great opportunities for early diagnosis of treatable stages of disease and immunotherapy to target EBV infected cells or overcome immune evasion. There is also scope for prevention of disease by immunisation and for development of therapeutic agents that target EBV gene products expressed in the cancers.
Bristol J, Djavadian R, Albright E, et al., 2018, A cancer-associated Epstein-Barr virus BZLF1 promoter variant enhances lytic infection, PLoS Pathogens, Vol: 14, ISSN: 1553-7366
Latent Epstein-Barr virus (EBV) infection contributes to both B-cell and epithelial-cell malignancies. However, whether lytic EBV infection also contributes to tumors is unclear, although the association between malaria infection and Burkitt lymphomas (BLs) may involve excessive lytic EBV replication. A particular variant of the viral promoter (Zp) that controls lytic EBV reactivation is over-represented, relative to its frequency in non-malignant tissue, in EBV-positive nasopharyngeal carcinomas and AIDS-related lymphomas. To date, no functional differences between the prototype Zp (Zp-P) and the cancer-associated variant (Zp-V3) have been identified. Here we show that a single nucleotide difference between the Zp-V3 and Zp-P promoters creates a binding site for the cellular transcription factor, NFATc1, in the Zp-V3 (but not Zp-P) variant, and greatly enhances Zp activity and lytic viral reactivation in response to NFATc1-inducing stimuli such as B-cell receptor activation and ionomycin. Furthermore, we demonstrate that restoring this NFATc1-motif to the Zp-P variant in the context of the intact EBV B95.8 strain genome greatly enhances lytic viral reactivation in response to the NFATc1-activating agent, ionomycin, and this effect is blocked by the NFAT inhibitory agent, cyclosporine, as well as NFATc1 siRNA. We also show that the Zp-V3 variant is over-represented in EBV-positive BLs and gastric cancers, and in EBV-transformed B-cell lines derived from EBV-infected breast milk of Kenyan mothers that had malaria during pregnancy. These results demonstrate that the Zp-V3 enhances EBV lytic reactivation to physiologically-relevant stimuli, and suggest that increased lytic infection may contribute to the increased prevalence of this variant in EBV-associated malignancies.
Styles CT, Paschos K, White R, et al., 2018, The cooperative functions of the EBNA3 Proteins are central to EBV persistence and latency, Pathogens, Vol: 31, ISSN: 2076-0817
The Epstein–Barr nuclear antigen 3 (EBNA3) family of proteins, comprising EBNA3A, EBNA3B, and EBNA3C, play pivotal roles in the asymptomatic persistence and life-long latency of Epstein–Barr virus (EBV) in the worldwide human population. EBNA3-mediated transcriptional reprogramming of numerous host cell genes promotes in vitro B cell transformation and EBV persistence in vivo. Despite structural and sequence similarities, and evidence of substantial cooperative activity between the EBNA3 proteins, they perform quite different, often opposing functions. Both EBNA3A and EBNA3C are involved in the repression of important tumour suppressive pathways and are considered oncogenic. In contrast, EBNA3B exhibits tumour suppressive functions. This review focuses on how the EBNA3 proteins achieve the delicate balance required to support EBV persistence and latency, with emphasis on the contribution of the Allday laboratory to the field of EBNA3 biology.
Correia S, Palser A, Elgueta Karstegl C, et al., 2017, Natural variation of Epstein-Barr virus genes, proteins and pri-microRNA, Journal of Virology, Vol: 91, ISSN: 1098-5514
Viral gene sequences from an enlarged set of about 200 Epstein-Barr virus (EBV) strains including many primary isolates have been used to investigate variation in key viral genetic regions, particularly LMP1, Zp, gp350, EBNA1 and the BART miRNA cluster 2. Determination of type 1 and type 2 EBV in saliva samples from people from a wide range of geographic and ethnic backgrounds demonstrates a small percentage of healthy white Caucasian British people carrying predominantly type 2 EBV. Linkage of Zp and gp350 variants to type 2 EBV is likely to be due to their genes being adjacent to the EBNA3 locus, which is one of the major determinants of the type 1/type 2 distinction. A novel classification of EBNA1 DNA binding domains named QCIGP results from phylogeny analysis of their protein sequences but is not linked to the type 1/type 2 classification. The BART cluster 2 miRNA region is classified into three major variants through SNPs in the pri-miRNA outside of the mature miRNA sequences. These SNPs can result in altered levels of expression of some miRNAs from the BART variant frequently present in Chinese and Indonesian nasopharyngeal carcinoma (NPC) samples. The EBV genetic variants identified here provide a basis for future more directed analysis of association of specific EBV variation with EBV biology and EBV associated diseases.IMPORTANCE Incidence of diseases associated with EBV varies greatly in different parts of the world. Relationships between EBV genome sequence variation and health, disease, geography and ethnicity of the host may thus be important for understanding the role of EBV in diseases and for development of an effective EBV vaccine. This paper provides the most comprehensive analysis so far of variation in specific EBV genes relevant to these diseases and proposed EBV vaccines. By focussing on variation in LMP1, Zp, gp350, EBNA1 and the BART miRNA cluster 2, new relationships to the known type 1/type 2 strains are demonstrated and novel classificati
Brady G, Haas DA, Farrell PJ, et al., 2017, Molluscum Contagiosum Virus Protein MC005 Inhibits NF-kappa B Activation by Targeting NEMO-Regulated I kappa B Kinase Activation, Journal of Virology, Vol: 91, ISSN: 1098-5514
Molluscum contagiosum virus (MCV), the only known extant human-adapted poxvirus, causes a long-duration infection characterized by skin lesions that typically display an absence of inflammation despite containing high titers of live virus. Despite this curious presentation, MCV is very poorly characterized in terms of host-pathogen interactions. The absence of inflammation around MCV lesions suggests the presence of potent inhibitors of human antiviral immunity and inflammation. However, only a small number of MCV immunomodulatory genes have been characterized in detail. It is likely that many more remain to be discovered, given the density of such sequences in other poxvirus genomes. NF-κB activation occurs in response to both virus-induced pattern recognition receptor (PRR) signaling and cellular activation by virus-induced proinflammatory cytokines like tumor necrosis factor and interleukin-1. Activated NF-κB drives cytokine and interferon gene expression, leading to inflammation and virus clearance. We report that MC005, which has no orthologs in other poxvirus genomes, is a novel inhibitor of PRR- and cytokine-stimulated NF-κB activation. MC005 inhibited NF-κB proximal to the IκB kinase (IKK) complex, and unbiased affinity purification revealed that MC005 interacts with the IKK subunit NEMO (NF-κB essential modulator). MC005 binding to NEMO prevents the conformational priming of the IKK complex that occurs when NEMO binds to ubiquitin chains during pathway activation. These data reveal a novel mechanism of poxvirus inhibition of human innate immunity, validate current dynamic models of NEMO-dependent IKK complex activation, and further clarify how the human-adapted poxvirus MCV can so effectively evade antiviral immunity and suppress inflammation to persist in human skin lesions.
West MJ, Farrell PJ, 2017, Roles of RUNX in B Cell Immortalisation, RUNX Proteins in Development and Cancer, Editors: Groner, Ito, Liu, Neil, Speck, VanWijnen, Publisher: SPRINGER-VERLAG SINGAPORE PTE LTD, Pages: 283-298, ISBN: 978-981-10-3231-8
RUNX1 and RUNX3 are the main RUNX genes expressed in B lymphocytes. Both are expressed throughout B-cell development and play key roles at certain key developmental transitions. The tumour-associated Epstein-Barr virus (EBV) has potent B-cell transforming ability and manipulates RUNX3 and RUNX1 transcription through novel mechanisms to control B cell growth. In contrast to resting mature B cells where RUNX1 expression is high, in EBV-infected cells RUNX1 levels are low and RUNX3 levels are high. Downregulation of RUNX1 in these cells results from cross-regulation by RUNX3 and serves to relieve RUNX1-mediated growth repression. RUNX3 is upregulated by the EBV transcription factor (TF) EBNA2 and represses RUNX1 transcription through RUNX sites in the RUNX1 P1 promoter. Recent analysis revealed that EBNA2 activates RUNX3 transcription through an 18 kb upstream super-enhancer in a manner dependent on the EBNA2 and Notch DNA-binding partner RBP-J. This super-enhancer also directs RUNX3 activation by two further RBP-J-associated EBV TFs, EBNA3B and 3C. Counter-intuitively, EBNA2 also hijacks RBP-J to target a super-enhancer region upstream of RUNX1 to maintain some RUNX1 expression in certain cell backgrounds, although the dual functioning EBNA3B and 3C proteins limit this activation. Interestingly, the B-cell genome binding sites of EBV TFs overlap extensively with RUNX3 binding sites and show enrichment for RUNX motifs. Therefore in addition to B-cell growth manipulation through the long-range control of RUNX transcription, EBV may also use RUNX proteins as co-factors to deregulate the transcription of many B cell genes during immortalisation.
Gunnell A, Webb HM, Wood CD, et al., 2016, RUNX super-enhancer control through the Notch pathway by Epstein-Barr virus transcription factors regulates B cell growth., Nucleic Acids Research, Vol: 44, Pages: 4636-4650, ISSN: 1362-4962
In B cells infected by the cancer-associated Epstein-Barr virus (EBV), RUNX3 and RUNX1 transcription is manipulated to control cell growth. The EBV-encoded EBNA2 transcription factor (TF) activates RUNX3 transcription leading to RUNX3-mediated repression of the RUNX1 promoter and the relief of RUNX1-directed growth repression. We show that EBNA2 activates RUNX3 through a specific element within a -97 kb super-enhancer in a manner dependent on the expression of the Notch DNA-binding partner RBP-J. We also reveal that the EBV TFs EBNA3B and EBNA3C contribute to RUNX3 activation in EBV-infected cells by targeting the same element. Uncovering a counter-regulatory feed-forward step, we demonstrate EBNA2 activation of a RUNX1 super-enhancer (-139 to -250 kb) that results in low-level RUNX1 expression in cells refractory to RUNX1-mediated growth inhibition. EBNA2 activation of the RUNX1 super-enhancer is also dependent on RBP-J. Consistent with the context-dependent roles of EBNA3B and EBNA3C as activators or repressors, we find that these proteins negatively regulate the RUNX1 super-enhancer, curbing EBNA2 activation. Taken together our results reveal cell-type-specific exploitation of RUNX gene super-enhancers by multiple EBV TFs via the Notch pathway to fine tune RUNX3 and RUNX1 expression and manipulate B-cell growth.
Gregorovic G, Boulden EA, Bosshard R, et al., 2015, Epstein-Barr viruses deficient in EBER RNAs give higher LMP2 RNA expression in lymphoblastoid cell lines and efficiently establish persistent infection in humanized mice., Journal of Virology, Vol: 89, Pages: 1171-11714, ISSN: 1098-5514
Functions of EBER RNAs were tested in lymphoblastoid cell lines containing EBER mutants of Epstein-Barr Virus (EBV). Binding of EBER1 to RPL22 was confirmed. Deletion of EBER1 or EBER2 correlated with increased cytoplasmic EBV LMP2 RNA and with small effects on specific cellular miRNA levels but protein levels of LMP1 and LMP2A were not affected. Wild type and EBER deletion EBV had approximately equal ability to infect immunodeficient mice reconstituted with a human haematopoietic system.
Brady G, Haas DA, Farrell PJ, et al., 2015, Poxvirus Protein MC132 from Molluscum Contagiosum Virus Inhibits NF-kappa B Activation by Targeting p65 for Degradation, JOURNAL OF VIROLOGY, Vol: 89, Pages: 8406-8415, ISSN: 0022-538X
Palser AL, Grayson NE, White RE, et al., 2015, Genome Diversity of Epstein-Barr Virus from Multiple Tumor Types and Normal Infection, Journal of Virology, Vol: 89, Pages: 5222-5237, ISSN: 1098-5514
Epstein-Barr virus (EBV) infects most of the world’s population and is causally associated with several human cancers, but littleis known about how EBV genetic variation might influence infection or EBV-associated disease. There are currently no publishedwild-type EBV genome sequences from a healthy individual and very few genomes from EBV-associated diseases. We havesequenced 71 geographically distinct EBV strains from cell lines, multiple types of primary tumor, and blood samples and thefirst EBV genome from the saliva of a healthy carrier. We show that the established genome map of EBV accurately represents allstrains sequenced, but novel deletions are present in a few isolates. We have increased the number of type 2 EBV genomes sequencedfrom one to 12 and establish that the type 1/type 2 classification is a major feature of EBV genome variation, definedalmost exclusively by variation of EBNA2 and EBNA3 genes, but geographic variation is also present. Single nucleotide polymorphism(SNP) density varies substantially across all known open reading frames and is highest in latency-associated genes. SomeT-cell epitope sequences in EBNA3 genes show extensive variation across strains, and we identify codons under positive selection,both important considerations for the development of vaccines and T-cell therapy. We also provide new evidence for recombinationbetween strains, which provides a further mechanism for the generation of diversity. Our results provide the firstglobal view of EBV sequence variation and demonstrate an effective method for sequencing large numbers of genomes to furtherunderstand the genetics of EBV infection.
Foerster A, Maertens GN, Farrell PJ, et al., 2015, Dimerization of Matrix Protein Is Required for Budding of Respiratory Syncytial Virus, Journal of Virology, Vol: 89, Pages: 4624-4635, ISSN: 1098-5514
Respiratory syncytial virus (RSV) infects epithelial cells of the respiratory tract and is a major cause of bronchiolitis and pneumonia in children and the elderly. The virus assembles and buds through the plasma membrane, forming elongated membrane filaments, but details of how this happens remain obscure. Oligomerization of the matrix protein (M) is a key step in the process of assembly and infectious virus production. In addition, it was suggested to affect the conformation of the fusion protein, the major current target for RSV antivirals, in the mature virus. The structure and assembly of M are thus key parameters in the RSV antiviral development strategy. The structure of RSV M was previously published as a monomer. Other paramyxovirus M proteins have been shown to dimerize, and biochemical data suggest that RSV M also dimerizes. Here, using size exclusion chromatography-multiangle laser light scattering, we show that the protein is dimeric in solution. We also crystallized M in two crystal forms and show that it assembles into equivalent dimers in both lattices. Dimerization interface mutations destabilize the M dimer in vitro. To assess the biological relevance of dimerization, we used confocal imaging to show that dimerization interface mutants of M fail to assemble into viral filaments on the plasma membrane. Additionally, budding and release of virus-like particles are prevented in M mutants that fail to form filaments. Importantly, we show that M is biologically active as a dimer and that the switch from M dimers to higher-order oligomers triggers viral filament assembly and virus production.
Farrell PJ, 2015, Epstein-Barr Virus Strain Variation, EPSTEIN BARR VIRUS, VOL 1: ONE HERPES VIRUS: MANY DISEASES, Vol: 390, Pages: 45-69, ISSN: 0070-217X
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- Citations: 32
Tzellos S, Correia PB, Karstegl CE, et al., 2014, A Single Amino Acid in EBNA-2 Determines Superior B Lymphoblastoid Cell Line Growth Maintenance by Epstein-Barr Virus Type 1 EBNA-2, JOURNAL OF VIROLOGY, Vol: 88, Pages: 8743-8753, ISSN: 0022-538X
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- Citations: 35
Pereira C, Sousa H, Silva J, et al., 2014, The-1195G Allele Increases the Transcriptional Activity of Cyclooxygenase-2 Gene (COX-2) in Colon Cancer Cell Lines, MOLECULAR CARCINOGENESIS, Vol: 53, Pages: E92-E95, ISSN: 0899-1987
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- Citations: 10
Hill ER, Koganti S, Zhi J, et al., 2013, Signal Transducer and Activator of Transcription 3 Limits Epstein-Barr Virus Lytic Activation in B Lymphocytes, JOURNAL OF VIROLOGY, Vol: 87, Pages: 11438-11446, ISSN: 0022-538X
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- Citations: 37
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