Gammaherpesvirus Research Articles

Herpesvirus surveillance in stranded striped dolphins (Stenella coeruleoalba) and bottlenose dolphins (Tursiops truncatus) from Italy with emphasis on neuropathological characterization.

Herpesvirus (HV) is widely distributed among cetacean populations, with the highest prevalence reported in the Mediterranean Sea. In this study, a comprehensive analysis was conducted, including epidemiological, phylogenetic, and pathological aspects, with particular emphasis on neuropathology, to better understand the impact of HV in these animals. Our results show a higher presence of HV in males compared to females, with males exhibiting a greater number of positive tissues. Additionally, adults were more frequently affected by HV infection than juveniles, with no infections detected in calves or neonates. The affected species were striped (Stenella coeruleoalba) and bottlenose dolphins (Tursiops truncatus). The highest positivity rates were observed in the genital system, cerebrum, and skin tissues. Phylogenetic analysis indicated a higher occurrence of Gammaherpesvirus (GHV) sequences but increased genetic diversity within Alphaherpesvirus (AHV). Key neuropathological features included astro-microgliosis (n = 4) and meningitis with minimal to mild perivascular cuffing (n = 2). The presence of concurrent infections with other pathogens, particularly cetacean morbillivirus (CeMV), underscores the complex nature of infectious diseases in cetaceans. However, the presence of lesions at the Central Nervous System (CNS) with molecular positivity for GHV, excluding the involvement of other potential neurotropic agents, would confirm the potential of this HV subfamily to induce neurological damage. Pathological examination identified lesions in other organs that could potentially be associated with HV, characterized by lymphoid depletion and tissue inflammation. These findings enhance our understanding of HV in odontocetes and highlight the need for ongoing research into the factors driving these infections and their broader implications.

The Epstein–Barr virus small capsid protein BFRF3 disrupts the NF‐кB signaling pathway by inhibiting p65 activity

AbstractEpstein–Barr virus (EBV), a common gamma herpesvirus, establishes a life‐long latent infection in the host to defend against innate immune recognition, which is closely related to a variety of malignant tumors, but its specific mechanism is unclear. BFRF3, an EBV‐encoded small capsid protein, is mainly involved in the assembly of the viral capsid structure and the maintenance of its stability. Here, we showed that BFRF3 can inhibit TNF‐α‐mediated NF‐кB promoter activation. Moreover, BFRF3 downregulates NF‐кB‐mediated promoter activation and transcription of inflammatory cytokines, including IL‐6 and IL‐8. Dual‐luciferase reporter assay demonstrated that BFRF3 restrains NF‐кB promoter activity at or below the p65 level, and coimmunoprecipitation analysis revealed that BFRF3 not only interacts with p65 but also binds to its critical truncated Rel homology domain (RHD) and transcriptional activation domain (TAD). However, BFRF3 does not affect the dimerization of p65‐p50, but overexpression of BFRF3 reduces the nuclear accumulation of p65, and the phosphorylation of p65 (Ser536) is repressed during BFRF3 transfection and EBV lytic infection, which promotes the proliferation of EBV. Overall, our study suggested that BFRF3 may play a crucial role in antiviral immunity to defend against EBV infection by inhibiting NF‐κB activity.

Understanding Metabolic Pathway Rewiring by Oncogenic Gamma Herpesvirus.

Gamma herpesviruses, including Epstein-Barr virus (EBV) and Kaposi's sarcoma-associated herpesvirus (KSHV), are key contributors to the development of various cancers through their ability to manipulate host cellular pathways. This review explores the intricate ways these viruses rewire host metabolic pathways to sustain viral persistence and promote tumorigenesis. We look into how EBV and KSHV induce glycolytic reprogramming, alter mitochondrial function, and remodel nucleotide and amino acid metabolism, highlighting the crucial role of lipid metabolism in these oncogenic processes. By understanding these metabolic alterations, which confer proliferative and survival advantages to the virus-infected cells, we can identify potential therapeutic targets and develop innovative treatment strategies for gamma herpesvirus-associated malignancies. Ultimately, this review underscores the critical role of metabolic reprogramming in gamma herpesvirus oncogenesis and its implications for precision medicine in combating virus-driven cancers.

Equine gamma herpesvirus presence and viral load are not associated with equine glandular gastric disease

Equine gamma herpesvirus presence and viral load are not associated with equine glandular gastric disease

Conserved cysteine residues in Kaposi's sarcoma herpesvirus ORF34 are necessary for viral production and viral pre-initiation complex formation.

Kaposi's sarcoma herpesvirus (KSHV) ORF34 plays a significant role as a component of the viral pre-initiation complex (vPIC), which is indispensable for late gene expression across beta and gamma herpesviruses. Although the key role of ORF34 within the vPIC and its function as a hub protein have been recognized, further clarification regarding its specific contribution to vPIC functionality and interactions with other components is required. This study employed a deep-learning algorithm-assisted structural model of ORF34, revealing highly conserved amino acid residues across human beta- and gamma-herpesviruses localized in structured domains. Thus, we engineered ORF34 alanine-scanning mutants by substituting conserved residues with alanine. These mutants were evaluated for their ability to interact with other vPIC factors and restore viral production in cells harboring the ORF34-deficient KSHV-BAC. Our experimental results highlight the crucial role of the 4 cysteine residues conserved in ORF34: a tetrahedral arrangement consisting of a pair of C-Xn-C consensus motifs. This suggests the potential incorporation of metal cations in interacting with ORF24 and ORF66 vPIC components, facilitating late gene transcription, and promoting overall virus production by capturing metal cations. In summary, our findings underline the essential role of conserved cysteines in KSHV ORF34 for effective vPIC assembly and viral replication, thereby enhancing our understanding of the complex interplay between the vPIC components.

Insights into the panorama of multiple DNA viruses in municipal wastewater and recycled sludge in Tianjin, China

Environmental viruses in wastewater and sludge are widely recognized for their roles in waterborne diseases. However, previous studies mainly focused on RNA viruses, and little is known about the diversity of DNA viral communities and their driving factors in municipal wastewater treatment environments. Herein, we conducted a pilot study to explore DNA virus profiles in municipal wastewater and recycled sludge by metagenomics method, and track their temporal changes in northern China. Results showed that 467 viral species were co-shared among all the samples. We identified six families of human viruses with a prevalence of 0.1%, which were rare but relatively stable in wastewater and sludge for six months. Adenoviridae, Parvoviridae, and Herpersviridae were the most dominant human viral families in municipal wastewater and recycled sludge. A time series of samples revealed that the dynamic changes of human DNA viruses were stable based on qPCR results, particularly for high-risk fecal-oral transmission viruses of adenovirus, bocavirus, polyomavirus, human gamma herpesvirus, human papillomavirus, and hepatitis B virus. Concentrations of Adenovirus (5.39–7.48 log10 copies/L) and bocavirus (4.36–7.48 log10 copies/L) were observed to be the highest in these samples compared to other viruses. Our findings demonstrated the DNA viruses’ high prevalence and persistence in municipal wastewater treatment environments, highlighting the value of enhancing public health responses based on wastewater-based epidemiology.

Equine gamma herpesvirus presence and viral load are not associated with equine glandular gastric disease.

To investigate the role of equine herpesvirus-2 (EHV-2) and equine herpesvirus-5 (EHV-5) in equine glandular gastric disease (EGGD) by visualizing and quantifying these gamma herpesviruses in EGGD-affected and normal glandular gastric mucosa of horses. A secondary objective was to describe the histopathological abnormalities in the equine gastric glandular mucosa in horses with EGGD. 29 horses (n = 21 postmortem and 8 gastroscopy) categorized as normal (11), EGGD (12), or both EGGD and equine squamous gastric disease (6). Glandular gastric mucosal samples were collected from horses by gastroscopy or postmortem. Histopathology and in situ hybridization targeting EHV-2 and EHV-5 were performed on grossly normal and abnormal glandular gastric mucosa. The number of in situ hybridization-positive cells per millimeter squared of tissue was calculated. Evaluators were blinded to groups. Glandular gastric tissues from horses without EGGD had higher viral loads in the mucosa than normal or abnormal tissues from EGGD horses. There was no difference in viral loads for EHV-2 or EHV-5 between grossly or endoscopically normal to abnormal gastric tissues within horses with EGGD. Lymphocytic plasmacytic gastritis was the most common histopathological abnormality, with only 3 horses having mucosal disruption (glandular ulcer or erosion). Equine gamma herpesviruses are unlikely to play a role in the pathophysiology of EGGD. EGGD is frequently inflammatory with occasional mucosal disruption (ulcer or erosion).

CORRELATION OF EPSTEIN-BARR VIRUS WITH BREAST CANCER: A CASE CONTROL STUDY

Background and aims: Epstein-Barr virus (EBV) is a gamma herpes virus and one of the most common infections globally. It has been linked to several health problems, including breast cancer, which is a leading cause of death among females worldwide. In Yemen, breast cancer is one of the top reported cancers in women. This study aimed to investigate the correlation between EBV and breast cancer among Yemeni women in Sana'a city. Methods: The study is a case-control study that included newly diagnosed, untreated breast cancer patients as cases and healthy women attending the National Cancer Control Foundation as controls. A total of 150 participants were tested for EBV (VCA) IgG using ELISA. Results: The association between EBV and BC in the case and control groups was found by the investigation. 48 women (64.0%) who tested positive for EBV IgG were included in the control group, whereas 71 women (94.7%) among the patients had the antibody identified. A substantial correlation between EBV and BC was found by statistical analysis; a chi-square value of 21.510 and a p-value of less than 0.001 supported this conclusion. With a 95% confidence interval, the odds ratio was 10.0 (3.3-30.4). Moreover, women above the age of thirty showed a higher risk than women under that age (p=0.006). In addition, compared to the control group, divorced and bereaved women had a significantly higher chance of contracting EBV (p= 0.002). Conclusions: According to the study's findings, there may be a link between EBV infection and a higher risk of breast cancer. To further understand the underlying mechanisms and potential strategies to reduce this risk, more research is required. Peer Review History: Received 1 December 2023; Revised 19 January 2024; Accepted 27 February; Available online 15 March 2024 Academic Editor: Dr. Muhammad Zahid Iqbal, AIMST University, Malaysia, drmmziqbal@gmail.com Average Peer review marks at initial stage: 6.0/10 Average Peer review marks at publication stage: 7.0/10 Reviewers: Dr. Andrzej Szymański, Poznan University of Technology, Poland, andrzej.szymanski@put.poznan.pl Dr. Fátima Morales Marín, University of Murcia, Spain, fatima.morales@um.es

Ovine Herpesvirus 2 Glycoprotein B Complementation Restores Infectivity to a Bovine Herpesvirus 4 gB-Null Mutant.

Ovine herpesvirus 2 (OvHV-2) and bovine herpesvirus 4 (BoHV-4) are gamma herpesviruses that belong to the genera Macavirus and Rhadinovirus, respectively. As with all herpesviruses, both OvHV-2 and BoHV-4 express glycoprotein B (gB), which plays an essential role in the infection of host cells. In that context, it has been demonstrated that a BoHV-4 gB-null mutant is unable to infect host cells. In this study, we used homologous recombination to insert OvHV-2 ORF 8, encoding gB, into the BoHV-4 gB-null mutant genome, creating a chimeric BoHV-4 virus carrying and expressing OvHV-2 gB (BoHV-4∆gB/OvHV-2-gB) that was infectious and able to replicate in vitro. We then evaluated BoHV-4∆gB/OvHV-2-gB as a potential vaccine candidate for sheep-associated malignant catarrhal fever (SA-MCF), a fatal disease of ungulates caused by OvHV-2. Using rabbits as a laboratory model for MCF, we assessed the safety, immunogenicity, and efficacy of BoHV-4∆gB/OvHV-2-gB in an immunization/challenge trial. The results showed that while BoHV-4∆gB/OvHV-2-gB was safe and induced OvHV-2 gB-specific humoral immune responses, immunization conferred only 28.5% protection upon challenge with OvHV-2. Therefore, future studies should focus on alternative strategies to express OvHV-2 proteins to develop an effective vaccine against SA-MCF.

Guinea pig herpes like virus is a gamma herpesvirus

Guinea Pig Herpes-Like Virus (GPHLV) is a virus isolated from leukemic guinea pigs with herpes virus-like morphology described by Hsiung and Kaplow in 1969. GPHLV transformed embryonic cells from Syrian hamsters or rats, which were tumorigenic in adult animals. Herein, we present the genomic sequence of GPHLV strain LK40 as a reference for future molecular analysis. GPHLV has a broad host tropism and replicates efficiently in Guinea pig, Cat, and Green African Monkey-derived cell lines. GPHLV has a GC content of 35.45%. The genome is predicted to encode at least 75 open-reading frames (ORFs) with 84% (63 ORFs) sharing homology to human Kaposi Sarcoma Associated Herpes Virus (KSHV). Importantly, GPHLV encodes homologues of the KSHV oncogenes, vBCL2 (ORF16), vPK (ORF36), viral cyclin (v-cyclin, ORF72), the latency associated nuclear antigen (LANA, ORF73), and vGPCR (ORF74). GPHLV is a Rhadinovirus of Cavia porcellus, and we propose the formal name of Caviid gamma herpesvirus 1 (CaGHV-1). GPHLV can be a novel small animal model of Rhadinovirus pathogenesis with broad host tropism.

Loop-mediated isothermal amplification linked a nanoparticles-based biosensor for detecting Epstein-Barr virus

Epstein-Barr virus (EBV) is a ubiquitous gamma herpesvirus that maintains a lifelong latent association with B lymphocytes. Here, a rapid and reliable diagnosis platform for detecting EBV infection, employing loop-mediated isothermal amplification (LAMP) combined with a gold nanoparticles–based lateral flow biosensors (AuNPs-LFB) (termed LAMP Amplification Mediated AuNPs-LFB Detection, LAMAD), was developed in the current study. A set of specific LAMP primers targeting the Epstein-Barr nuclear antigen (EBNA) leader protein (EBNA-LP) gene was designed and synthesized. Subsequently, these templates extracted from various pathogens and whole blood samples were used to optimize and evaluate the EBV-LAMAD assay. As a result, the limit of detection (LoD) of the EBV-LAMAD assay was 45 copies/reaction. The EBV-LAMAD assay can detect all representative EBV pathogens used in the study, and of note, no cross-reactions were observed with other non-EBV organisms. Moreover, the whole workflow of the EBV-LAMAD assay can be completed within 70 min, including rapid EBV template preparation, EBV-LAMP amplification, and AuNPs-LFB-mediated detection. Taken together, the EBV-LAMAD assay targeting the EBNA-LP gene is a rapid, simplified, sensitive, reliable, and easy-to-use detection protocol that can be used as a competitive potential diagnostic/screening tool for EBV infection in clinical settings, especially in basic laboratories in resource-limited regions.Key points• A novel, simplified, and easy-to-use AuNPs-LFB biosensor was designed and prepared.• LAMP combined with an AuNPs-LFB targeting the novel EBNA-LP gene was established.• EBV-LAMAD is a rapid, sensitive, and reliable detection protocol for EBV infection.Graphical

Kaposi's sarcoma herpesvirus exploits the DNA damage response to circularize its genome.

To establish lifelong, latent infection, herpesviruses circularize their linear, double-stranded, DNA genomes through an unknown mechanism. Kaposi's sarcoma (KS) herpesvirus (KSHV), a gamma herpesvirus, is tightly linked with KS, primary effusion lymphoma, and multicentric Castleman's disease. KSHV persists in latently infected cells as a multi-copy, extrachromosomal episome. Here, we show the KSHV genome rapidly circularizes following infection, and viral protein expression is unnecessary for this process. The DNA damage response (DDR) kinases, ATM and DNA-PKcs, each exert roles, and absence of both severely compromises circularization and latency. These deficiencies were rescued by expression of ATM and DNA-PKcs, but not catalytically inactive mutants. In contrast, γH2AX did not function in KSHV circularization. The linear viral genomic ends resemble a DNA double strand break, and non-homologous DNA end joining (NHEJ) and homologous recombination (HR) reporters indicate both NHEJ and HR contribute to KSHV circularization. Last, we show, similar to KSHV, ATM and DNA-PKcs have roles in circularization of the alpha herpesvirus, herpes simplex virus-1 (HSV-1), while γH2AX does not. Therefore, the DDR mediates KSHV and HSV-1 circularization. This strategy may serve as a general herpesvirus mechanism to initiate latency, and its disruption may provide new opportunities for prevention of herpesvirus disease.

Cerebral meningitis due to tuberculoma and epstein barr: presentation of a clinical case

Neuroinfection is an inflammatory process that affects the meninges or brain parenchyma; it has various etiologies, including viral and non-viral, including autoimmune, bacterial and fungal; patients with this pathology represent a challenge for doctors; the severity varies, from benign, self-limiting to life-threatening. The Epstein-Barr virus (EBV) is a double-stranded DNA gamma herpesvirus that presents a latent infection and lytic replication; it can diffuse into the central nervous system and alter the integrity of the blood-brain barrier, being associated with neurocognitive impairment, neuronal damage and inflammation. In relation to tuberculosis, it became the second infectious disease that caused the most deaths in the world after COVID-19; tuberculous meningitis is considered the most severe form of extrapulmonary tuberculosis (TB) with a mortality of 70% in low-income countries. Below is the case of a 35-year-old man with a history of adrenal insufficiency and hyperthyroidism, who was receiving corticosteroids; he went to the emergency service with a 5-month history of clinical symptoms characterized by holocranial headache, focal retrograde amnesia, periods of altered state of consciousness, dizziness, nausea that led to vomiting on several occasions, with sudden loss of consciousness. , accompanied by involuntary tonic-clonic movements and hearing loss; after the diagnostic screening, EBV and TB are identified; receives specific treatment with good clinical evolution

Ivermectin inhibits replication of the malignant catarrhal fever virus alcelaphine herpesvirus 1

Malignant catarrhal fever is a lymphoproliferative disease of cattle and other ungulates that is caused by genetically and antigenically related gamma herpesviruses of the genus Macavirus. Infection of the natural host species is efficient and asymptomatic but spread to susceptible hosts is often fatal with clinical signs including fever, depression, nasal and ocular discharge. There is no recognised treatment for MCF but a vaccine for one MCF virus, alcelaphine herpesvirus 1 (AlHV-1), has been described. In this paper we describe the inhibition of AlHV-1 replication and propagation by the anthelminthic drug ivermectin. Concentrations of 10 μM or greater led to significant reductions in both copy number and viable titre of virus tested in culture medium, with little replication detected at over 20 μM ivermectin. In the absence of alternative treatments, further testing of ivermectin as a candidate antiviral treatment for MCF may therefore be justified.

Neuropilin-1 identifies a subset of highly activated CD8+ T cells during parasitic and viral infections.

Neuropilin-1 (Nrp-1) expression on CD8+ T cells has been identified in tumor-infiltrating lymphocytes and in persistent murine gamma-herpes virus infections, where it interferes with the development of long-lived memory T cell responses. In parasitic and acute viral infections, the role of Nrp-1 expression on CD8+ T cells remains unclear. Here, we demonstrate a strong induction of Nrp-1 expression on CD8+ T cells in Plasmodium berghei ANKA (PbA)-infected mice that correlated with neurological deficits of experimental cerebral malaria (ECM). Likewise, the frequency of Nrp-1+CD8+ T cells was significantly elevated and correlated with liver damage in the acute phase of lymphocytic choriomeningitis virus (LCMV) infection. Transcriptomic and flow cytometric analyses revealed a highly activated phenotype of Nrp-1+CD8+ T cells from infected mice. Correspondingly, in vitro experiments showed rapid induction of Nrp-1 expression on CD8+ T cells after stimulation in conjunction with increased expression of activation-associated molecules. Strikingly, T cell-specific Nrp-1 ablation resulted in reduced numbers of activated T cells in the brain of PbA-infected mice as well as in spleen and liver of LCMV-infected mice and alleviated the severity of ECM and LCMV-induced liver pathology. Mechanistically, we identified reduced blood-brain barrier leakage associated with reduced parasite sequestration in the brain of PbA-infected mice with T cell-specific Nrp-1 deficiency. In conclusion, Nrp-1 expression on CD8+ T cells represents a very early activation marker that exacerbates deleterious CD8+ T cell responses during both, parasitic PbA and acute LCMV infections.

Highly Active Cytotoxic CD4 + T Cells Targeting EBV Oncoproteins with Enhanced In Vivo Anti-Tumor and Self-Renewal Properties

Background: Epstein-Barr virus (EBV), a human gamma herpesvirus primarily of B-cell tropism that infects more than 90% of humans during childhood, establishing a lifelong latent persistence after the initial immune clearance. Alterations in the immune surveillance in subjects with impaired cellular immunity can lead to viral reactivation and uncontrolled B cell lymphoproliferation manifesting as post-transplant lymphoproliferative disease (PTLD) and malignant transformation into B-cell lymphomas and some epithelial tumors. Adoptive transfer of virus-specific cytotoxic T cells targeting viral oncoproteins of EBV (EBV-CTLs) may lead to rapid reconstitution of T cell immune surveillance and may efficiently control PTLD and some EBV-driven lymphomas. However, success of this approach has been hampered by suboptimal/variable potency of donor-derived EBV-CTLs and possibly their terminally differentiated/exhausted phenotype associated with limited in vivo persistence and activity, especially when targeting elusive Latency 1-associated diseases. We hypothesized that that EBV-specific CD4 + T cells differentiated using novel pro-inflammatory conditions might efficiently target viral oncoproteins concurrently retaining highly desirable stem-cell like properties required for successful eradication of tumor cells in vivo. Methods: In this study, we developed a method for generating novel and highly potent CD4 + cytotoxic T cells (CD4 +-CTLs) simultaneously targeting EBNA1, LMP1, and LMP2A. Peripheral blood mononuclear cells (PBMCs) were isolated from healthy donors, and either total T cells or CD4 +T cells were purified. These cells were then stimulated with PBMCs loaded with overlapping peptide libraries spanning LMP1, LMP2A, and EBNA1 antigens, either under neutral conditions (IL2 and IL-7) or in the presence of pro-inflammatory (type 1/17) cytokines ex vivo for 12-14 days. The phenotypes, reactivity, and cytotoxic capability of these CD4 +-CTLs against autologous EBV-Lymphoblastoid cell lines (LCLs) were assessed in vitro and in vivo upon adoptive transfer into tumor-bearing NSG mice. Results: In comparison to EBV-CTLs generated from unselected PBMCs (standard conditions), purified CD4 + expanded in the presence of pro-inflammatory cytokines exhibited significantly higher frequency of antigen specific T cells with a marked skewing towards EBNA1 antigen, and displaying marked polyfunctionality, concurrently secreting tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), and inducible granzyme B (GzmB), while maintaining interleukin-2 (IL-2) secretion capacity. Antigen-reactive CD4 +-CTLs induced in pro-inflammatory conditions retained a significant population with central memory (T CM) phenotype (CCR7 hi, CD27 hi), while those expanded in neutral conditions predominantly displayed an effector memory (T EM) phenotype. These CD4 +-CTLs demonstrated enhanced cytotoxicity against autologous EBV-LCLs and superior proliferative capacity in vitro upon secondary stimulation, whereas standard EBV-CTLs proliferated poorly. In the LCL-injected NSG mice, CD4 +-CTLs exhibited superior self-renewal and long-term persistence, resulting in a significant reduction/eradication of established LCL burden when compared to standard EBV-CTLs. Conclusion: Ex vivo priming with EBV antigens in the presence of pro-inflammatory cytokines reliably induces EBV-reactive CD4 +-CTLs with markedly enhanced antigen-specificity, polyfunctionality, cytotoxic potential, preserved stemness, proliferative capacity, and superior in vivo persistence and anti-tumor activity . These findings hold significant promise for developing improved adoptive immunotherapies to target EBV-associated diseases, especially of Latency-I origin and may be generalized as a strategy for targeting other viral oncoproteins expressed by human cancers. Figure 1: EBV-CD4 + CTLs generated under pro-inflammatory conditions display superior reactivity, persistence, and anti-tumor activity in vivo: (A.) The percentage of CD4 +Th cells primed with LMP1/2 and EBNA1 and expanded under indicated conditions producing TNF-α, IFN-γ, IL-2 and GZMB upon antigenic restimulation (n=29). (B.) Frequency of EBV-LCLs (CD23 +) and CD3 + cells detected in spleens on Day 26 post adoptive transfer of standard and pro-inflammatory EBV-CD4 + CTLs(n=4 per group).

Comprehensive Analysis of the Tegument Proteins Involved in Capsid Transport and Virion Morphogenesis of Alpha, Beta and Gamma Herpesviruses.

Herpesviruses are enveloped and have an amorphous protein layer surrounding the capsid, which is termed the tegument. Tegument proteins perform critical functions throughout the viral life cycle. This review provides a comprehensive and comparative analysis of the roles of specific tegument proteins in capsid transport and virion morphogenesis of selected, well-studied prototypes of each of the three subfamilies of Herpesviridae i.e., human herpesvirus-1/herpes simplex virus-1 (Alphaherpesvirinae), human herpesvirus-5/cytomegalovirus (Betaherpesvirinae) and human herpesvirus -8/Kaposi's sarcomavirus (Gammaherpesvirinae). Most of the current knowledge is based on alpha herpesviruses, in particular HSV-1. While some tegument proteins are released into the cytoplasm after virus entry, several tegument proteins remain associated with the capsid and are responsible for transport to and docking at the nucleus. After replication and capsid formation, the capsid is enveloped at the nuclear membrane, which is referred to as primary envelopment, followed by de-envelopment and release into the cytoplasm. This requires involvement of at least three tegument proteins. Subsequently, multiple interactions between tegument proteins and capsid proteins, other tegument proteins and glycoproteins are required for assembly of the virus particles and envelopment at the Golgi, with certain tegument proteins acting as the central hub for these interactions. Some redundancy in these interactions ensures appropriate morphogenesis.

Helicase-independent function of RIG-I against murine gammaherpesvirus 68 via blocking the nuclear translocation of viral proteins

Innate immunity is the first line of defense against viral pathogens. Retinoic Acid-Inducible Gene 1 (RIG-I) is a pattern recognition receptor that recognizes virus-associated double-stranded RNA and initiates the interferon responses. Besides signal transduction, RIG-I exerts direct antiviral functions to displace viral proteins on dsRNA via its Helicase activity. Nevertheless, this effector-like activity of RIG-I against herpesviruses remains largely unexplored. It has been previously reported that herpesviruses deamidate RIG-I, resulting in the abolishment of its Helicase activity and signal transduction. In this study, we discovered that RIG-I possessed signaling-independent antiviral activities against murine gamma herpesviruses 68 (γHV68, murid herpesvirus 4). Importantly, a Helicase-dead mutant of RIG-I (K270A) demonstrated comparable inhibition on herpesviruses lytic replication, indicating that this antiviral activity is Helicase-independent. Mechanistically, RIG-I bound the Replication and Transcription Activator (RTA) and diminished its nuclear localization to repress viral transcription. We further demonstrated that RIG-I blocked the nuclear translocation of ORF21 (Thymidine Kinase), ORF75c (vGAT), both of which form a nuclear complex with RTA and RNA polymerase II (Pol II) to facilitate viral transcription. Moreover, RIG-I retained ORF59 (DNA processivity factor) in the cytoplasm to repress viral DNA replication. Altogether, we illuminated a previously unidentified, Helicase-independent effector-like function of RIG-I against γHV68, representing an exquisite host strategy to counteract viral manipulations on innate immune signaling. ImportanceRetinoic acid-inducible gene I (RIG-I), a member of DExD/H box RNA helicase family, functions as a key pattern recognition receptor (PRR) responsible for the detection of intracellular double-stranded RNA (dsRNA) from virus-infected cells and induction of type I interferon (IFN) responses. Nevertheless, our understanding of the helicase-independent effector-like activity of RIG-I against virus infection, especially herpesvirus infection, remains largely unknown. Herein, by deploying murine gamma herpesviruses 68 (γHV68) as a model system, we demonstrated that RIG-I possessed an interferon and helicase-independent antiviral activity against γHV68 via blocking the nuclear trafficking of viral proteins, which concomitantly repressed the viral early transcription and genome replication thereof. Our work illuminates a previously unidentified antiviral strategy of RIG-I against herpesvirus infection.

Divergent structures of Mammalian and gammaherpesvirus uracil DNA glycosylases confer distinct DNA binding and substrate activity

Uracil DNA glycosylase (UNG) removes mutagenic uracil base from DNA to initiate base excision repair (BER). The result is an abasic site (AP site) that is further processed by the high-fidelity BER pathway to complete repair and maintain genome integrity. The gammaherpesviruses (GHVs), human Kaposi sarcoma herpesvirus (KSHV), Epstein-Barr virus (EBV), and murine gammaherpesvirus 68 (MHV68) encode functional UNGs that have a role in viral genome replication. Mammalian and GHVs UNG share overall structure and sequence similarity except for a divergent amino-terminal domain and a leucine loop motif in the DNA binding domain that varies in sequence and length. To determine if divergent domains contribute to functional differences between GHV and mammalian UNGs, we analyzed their roles in DNA interaction and catalysis. By utilizing chimeric UNGs with swapped domains we found that the leucine loop in GHV, but not mammalian UNGs facilitates interaction with AP sites and that the amino-terminal domain modulates this interaction. We also found that the leucine loop structure contributes to differential UDGase activity on uracil in single- versus double-stranded DNA. Taken together we demonstrate that the GHV UNGs evolved divergent domains from their mammalian counterparts that contribute to differential biochemical properties from their mammalian counterparts.

αβ and γδ T-cell responses to Epstein-Barr Virus: insights in immunocompetence, immune failure and therapeutic augmentation in transplant patients.

Epstein-Barr Virus (EBV) is a human gamma herpes virus, which causes several diseases in immunocompetent (mononucleosis, chronic fatigue syndrome, gastric cancer, endemic Burkitt's lymphoma, head and neck cancer) and immunosuppressed (post-transplant lymphoproliferative disease, EBV-associated soft tissue tumors) patients. It elicits a complex humoral and cellular immune response with both innate and adaptive immune components. Substantial progress has been made in understanding the interplay of immune cells in EBV-associated diseases in recent years, and several therapeutic approaches have been developed to augment cellular immunity toward EBV for control of EBV-associated malignancy. This review will focus on recent developments in immunosuppressed transplant recipients.