Potentiation of drug toxicity through virus latency reversal promotes preferential elimination of HIV infected cells

Eliminating latently infected cells is a highly challenging, indispensable step towards the overall cure for HIV/AIDS. We recognized that the unique HIV protease cut site (Phe-Pro) can be reconstructed using a potent toxin, monomethyl auristatin F (MMAF), which features Phe at its C-terminus. We hypothesized that this presents opportunities to design prodrugs that are specifically activated by the HIV protease. To investigate this, a series of MMAF derivatives was synthesized and evaluated in cell culture using latently HIV-infected cells. Cytotoxicity of compounds was enhanced upon latency reversal by up to 11-fold. In a mixed cell population, nanomolar concentrations of the lead compound depleted predominantly the HIV-infected cells and in doing so markedly enriched the pool with the uninfected cells. Despite expectation, mechanism of action of the synthesized toxins was not as HIV protease-specific prodrugs, but likely through the synergy of toxicities between the toxin and the reactivated virus.

Study On the Mechanism of LMP2A Maintaining Epstein-barr Virus Latency Infection Through Interaction With CXCR4

Epstein-Barr virus (EBV) belongs to the γ-herpesvirus subfamily and is the first human tumor virus to be discovered. The global adult infection rate exceeds 90%. EBV can participate in the regulation of multiple genes and multiple signal pathways through its latent genes. Many studies have reported that CXCR4 is involved in the development of gastric cancer, but there are few studies on the specific mechanism of its role in EBV-associated gastric cancer (EBVaGC). In this study, we explored the mechanism by which EBV-encoded products maintain EBV latent infection through interaction with CXCR4, and the role of CXCR4 in EBV positive cells. The results show that there is a positive feedback between the EBV-encoded products and CXCR4, and LMP2A can activate CXCR4 through the NF-κB pathway. In addition, CXCR4 can be fed back to LMP2A and EBNA1 through the ERK signaling pathway. At the same time, CXCR4 can promote the proliferation and migration of EBV-positive cells, reduce the expression of the immediate early protein BZLF1, and play an important role in maintaining the incubation period of EBV infection. These findings are conducive to the further targeted therapy of EBVaGC.

Molecular Basis of Epstein–Barr Virus Latency Establishment and Lytic Reactivation

Epstein–Barr virus (EBV) is a causative agent of infectious mononucleosis and several types of cancer. Like other herpesviruses, it establishes an asymptomatic, life-long latent infection, with occasional reactivation and shedding of progeny viruses. During latency, EBV expresses a small number of viral genes, and exists as an episome in the host–cell nucleus. Expression patterns of latency genes are dependent on the cell type, time after infection, and milieu of the cell (e.g., germinal center or peripheral blood). Upon lytic induction, expression of the viral immediate-early genes, BZLF1 and BRLF1, are induced, followed by early gene expression, viral DNA replication, late gene expression, and maturation and egress of progeny virions. Furthermore, EBV reactivation involves more than just progeny production. The EBV life cycle is regulated by signal transduction, transcription factors, promoter sequences, epigenetics, and the 3D structure of the genome. In this article, the molecular basis of EBV latency establishment and reactivation is summarized.

Recent Issues in Varicella-Zoster Virus Latency

Varicella-zoster virus (VZV) is a human herpes virus which causes varicella (chicken pox) as a primary infection, and, following a variable period of latency in neurons in the peripheral ganglia, may reactivate to cause herpes zoster (shingles) as well as a variety of neurological syndromes. In this overview we consider some recent issues in alphaherpesvirus latency with special focus on VZV ganglionic latency. A key question is the nature and extent of viral gene transcription during viral latency. While it is known that this is highly restricted, it is only recently that the very high degree of that restriction has been clarified, with both VZV gene 63-encoded transcripts and discovery of a novel VZV transcript (VLT) that maps antisense to the viral transactivator gene 61. It has also emerged in recent years that there is significant epigenetic regulation of VZV gene transcription, and the mechanisms underlying this are complex and being unraveled. The last few years has also seen an increased interest in the immunological aspects of VZV latency and reactivation, in particular from the perspective of inborn errors of host immunity that predispose to different VZV reactivation syndromes.

Implication of the cellular factor CTCF in the regulation of Bovine Leukemia Virus latency and tridimensional chromatin organization

ABSTRACTBovine Leukemia Virus (BLV)-induced tumoral development is a multifactorial phenomenon which remains largely unelucidated. Here, we highlighted the critical role of the cellular CCCTC-binding factor (CTCF) both in the regulation of BLV transcriptional activities and in the deregulation of the tridimensional (3D) chromatin architecture surrounding the BLV integration site. We demonstrated the in vivo recruitment of CTCF to three conserved CTCF binding motifs along the BLV provirus. Next, we showed a critical role for CTCF in delimitating the epigenetic landscape along the BLV provirus as well as to repress the 5’Long Terminal Repeat (LTR) promoter activity, thereby contributing to viral latency, while favoring the 3’LTR promoter activity. Finally, we demonstrated that BLV integration deregulated host cellular 3D chromatin organization through the formation of abnormal viral/host chromatin loops. Altogether, our results highlight CTCF as a new critical effector of BLV transcriptional regulation and BLV-induced physiopathology.

​Dermal Response to Experimental Orfvirus (ORFV) Infection in Goats, Mice and Rabbit

Background: During a study on the outbreak of orf in goats, it was intended to study the disease transmissibility in different hosts from field samples and ascertain the infective potential of the agent in laboratory animals compared to goats. Methods: Cutaneous clinical materials from orf virus (ORFV) infected goats was used to experimentally infect naive goats, rabbits and mice and ascertain its infective potential and transmissibility in different hosts. The processed inoculum was applied topically to mimic a natural transmission through injured skin. Regular skin biopsies were taken that revealed characteristic macroscopic and microscopic lesions typical of orf. Result: Virus inoculum applied on abraded skin in goats successfully established the lesions of orf. A parallel inoculation in rabbit and mice could not successfully reproduce the disease in these unnatural hosts beyond a subtle vesicular stage on 3 dpi with subsequent healing by 7 dpi. The lesions in goats regressed spontaneously by 28 days post-infection (dpi). Intracytoplasmic inclusions were associated only in the vesicular stage. Immunopathological progression was observed by immunoperoxidase staining of CD4+ and CD8+ T-cells which were found to appear by day 5 in the dermis and became more abundant and distributed by day 8, but subsequently reduced in number by 15 dpi. CD4+ cells were found to be more numerous and widespread. Viral antigen in tissues could be demonstrated by 4 dpi by immunohistological methods that increased in signal intensity progressively and disappear by 28 dpi. Similarly, viral nucleic acid in the skin could be detected on day 8 dpi but not on 28 dpi by PCR. The present experiment depicts the ease of disease transmissibility through traumatized skin in the primary hosts, but establishment in unnatural hosts may not be readily achieved. The infection was self-limiting with possibly no virus latency as indicated by immunofluorescence and PCR studies.

Different expression pattern of human cytomegalovirus-encoded microRNAs in circulation from virus latency to reactivation

Background: Human cytomegalovirus (HCMV) is a beta-hersvirinae that has a high latent infection rate worldwide and can cause serious consequences in immunocompromised patients when reactivation; however, the mechanism of how HCMV convert from latent to reactivation has rarely been investigated. In the present study, we aimed to perform a comprehensive analysis of the HCMV-encoded microRNA (miRNA) profile in serum of patients upon HCMV reactivation from latency and to further evaluate its clinical significance for the disease monitoring and preventing usefulness.Methods: Serum samples from 59 viremia patients and 60 age-gender matched controls were enrolled in this study for screening and validation of different expression of HCMV miRNAs. Serum concentrations of 22 known HCMV miRNAs were determined by a hydrolysis probe-based stem-loop quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay. HCMV DNA was measured by quantitative real-time PCR (qPCR) with the whole blood sample. Serum HCMV IgG and IgM were assessed using enzyme linked immunosorbent assay (ELISA). Another 47 samples from 5 patients at different time points were collected to evaluate the monitoring effectiveness and disease prediction ability of differential expression HCMV-miRNAs during the antiviral treatment. Results: The RT-qPCR analysis revealed that the serum levels of 16 of the 22 examined HCMV miRNAs were elevated in HCMV viremia patients compared with controls, and a profile of 8 HCMV miRNAs including hcmv-miR-US25-2-3p, hcmv-miR-US4-5p, hcmv-miR-US25-2-5p, hcmv-miR-US25-1-3p, hcmv-miR-US25-1, hcmv-miR-UL36, hcmv-miR-UL148D, hcmv-miR-US29-3p were markedly elevated (fold change > 2, P < 0.01). Receiver operating characteristic curve (ROC) analysis were performed on the selected HCMV-miRNAs in all of the patients and controls that enrolled in this study, and which ranged from 0.72 to 0.80 in the autoimmune patients. In addition, hcmv-miR-US25-1-3p levels were significantly correlated with HCMV DNA load (r = 0.349，P = 0.007), and were obviously higher in the reactivation set than the latency set in the autoimmune patients, which could be a predictor for the monitoring of the antiviral treatment.Conclusions: HCMV miRNAs profile showed markedly shift-switch from latency to reactivation in circulation from HCMV infected patients and hcmv-miR-US25-1-3p may be served as a predictor for the switch upon reactivation from latency in patients suffered with autoimmune diseases.

Different expression pattern of human cytomegalovirus-encoded microRNAs in circulation from virus latency to reactivation

Background Human cytomegalovirus (HCMV) is a beta-hersvirinae that has a high latent infection rate worldwide and can cause serious consequences in immunocompromised patients when reactivation; however, the mechanism of how HCMV convert from latent to reactivation has rarely been investigated. In the present study, we aimed to perform a comprehensive analysis of the HCMV-encoded microRNA (miRNA) profile in serum of patients upon HCMV reactivation from latency and to further evaluate its clinical significance for the disease monitoring and preventing usefulness. Methods Serum samples from 59 viremia patients and 60 age-gender matched controls were enrolled in this study for screening and validation of different expression of HCMV miRNAs. Serum concentrations of 22 known HCMV miRNAs were determined by a hydrolysis probe-based stem-loop quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay. HCMV DNA was measured by quantitative real-time PCR (qPCR) with the whole blood sample. Serum HCMV IgG and IgM were assessed using enzyme linked immunosorbent assay (ELISA). Another 47 samples from 5 patients at different time points were collected to evaluate the monitoring effectiveness and disease prediction ability of differential expression HCMV-miRNAs during the antiviral treatment. Results The RT-qPCR analysis revealed that the serum levels of 16 of the 22 examined HCMV miRNAs were elevated in HCMV viremia patients compared with controls, and a profile of 8 HCMV miRNAs including hcmv-miR-US25-2-3p, hcmv-miR-US4-5p, hcmv-miR-US25-2-5p, hcmv-miR-US25-1-3p, hcmv-miR-US25-1, hcmv-miR-UL36, hcmv-miR-UL148D, hcmv-miR-US29-3p were markedly elevated (fold change > 2, P < 0.01). Receiver operating characteristic curve (ROC) analysis were performed on the selected HCMV-miRNAs in all of the patients and controls that enrolled in this study, and which ranged from 0.72 to 0.80 in the autoimmune patients. In addition, hcmv-miR-US25-1-3p levels were significantly correlated with HCMV DNA load (r = 0.349, P = 0.007), and were obviously higher in the reactivation set than the latency set in the autoimmune patients, which could be a predictor for the monitoring of the antiviral treatment. Conclusions HCMV miRNAs profile showed markedly shift-switch from latency to reactivation in circulation from HCMV infected patients and hcmv-miR-US25-1-3p may be served as a predictor for the switch upon reactivation from latency in patients suffered with autoimmune diseases.