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Gammaherpesviruses establish life-long infection in over 95% of adults and are associated with several cancers, including B cell lymphomas. Using the murine gammaherpesvirus 68 (MHV68) animal model, we previously showed a pro-viral role of Interleukin-1 (IL-1) signaling that supported viral reactivation during the establishment of chronic infection. Unexpectedly, in this study we found that the proviral effects of IL-1 signaling originally observed during the establishment of chronic gammaherpesvirus infection convert to antiviral effects during the long-term stage of infection. Specifically, IL-1 signaling promoted expansion of antiviral CD8

Epstein-Barr virus (EBV) is a ubiquitous, oncogenic virus that is associated with a number of different human malignancies as well as autoimmune disorders. The expression of EBV viral proteins and non-coding RNAs contribute to EBV-mediated disease pathologies. The virus establishes life-long latency in the human host and is adept at evading host innate and adaptive immune responses. In this review, we discuss the life cycle of EBV, the various functions of EBV-encoded proteins and RNAs, the ability of the virus to activate and evade immune responses, as well as the neoplastic and autoimmune diseases that are associated with EBV infection in the human population.

The persistence of latent HIV-1 reservoirs in cells presents a formidable challenge towards a complete HIV cure. Edelfosine is an FDA-approved investigational, anti-neoplastic drug. In this study, we aimed to investigate its role as a HIV-1 Latency Reversal Agent (LRA) using latency model cell lines. Our findings demonstrated that edelfosine reactivated latent HIV-1 viruses in myeloid cells in a dose and time-dependent manner. The mechanism of reactivation by edelfosine involved the activation of NF-κB and AP1 pathways in these cells. The reactivated virus was non-infectious. Delineating the mechanism of non-infectious virus production revealed an increased stabilization of cellular APOBEC3G protein as well as its enhanced incorporation into the released viruses. Thus, our study demonstrated for the first time an additional role of edelfosine in reactivation of latent HIV-1 and production of non-infectious virus. Our results have paved the way for repurposing of edelfosine as a novel HIV-1 latency reversal agent.

Elimination of the latent HIV cell reservoir may be possible, if the molecular identity of latently infected cells were fully elucidated. We conducted comprehensive molecular profiling, at the protein and RNA levels, of primary T cells latently infected with HIV in vitro. Isobaric labelling quantitative proteomics and RNA sequencing identified 1453 proteins and 618 genes, altered in latently infected cells compared to mock-infected controls (p 0.05). Biomarker selection was based on results from integrated data analysis. Relative enrichment for latently infected cells was monitored using flow cytometric sorting and the HIV integrant assay. Antibodies against selected proteins, encoded by CEACAM1 and PLXNB2, enabled enrichment of latently infected cells from cell mixtures by 3-10 fold (5.8 average, p 0.001), comparable to levels obtained with biomarkers reported previously. Individual biomarkers are likely linked to subsets of latently infected cells, and an extended antibody panel will be required to inclusively target the latent HIV reservoir.

Since the discovery of HIV-1, progress has been made in deciphering the viral replication cycle and mechanisms of host-pathogen interactions that has facilitated the implementation of effective antiretroviral therapies (ARTs). Major barriers to HIV-1 remission/cure include the persistence of viral reservoirs (VRs) in long-lived CD4

Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic herpesvirus that exhibits two distinct phases of infection in the host- latent and lytic. The quiescent latent phase is defined by limited expression of a subset of viral proteins and microRNAs, and an absence of virus production. KSHV periodically reactivates from latency to undergo active lytic replication, leading to production of new infectious virions. This switch from the latent to the lytic phase requires the viral protein regulator of transcription activator (RTA). RTA, along with other virally encoded proteins, is aided by host factors to facilitate this transition. Herein, we highlight the key host proteins that are involved in mediating RTA activation and KSHV lytic replication and discuss the cellular processes in which they function. We will also focus on the modulation of viral reactivation by the innate immune system, and how KSHV influences key immune signaling pathways to aid its own lifecycle.

Epstein-Barr virus (EBV) infects 95% of adults worldwide, causes infectious mononucleosis, is etiologically linked to multiple sclerosis and is associated with 200 000 cases of cancer each year. EBV manipulates host epigenetic pathways to switch between a series of latency programs and to reactivate from latency in order to colonize the memory B-cell compartment for lifelong infection and to ultimately spread to new hosts. Here, we review recent advances in the understanding of epigenetic mechanisms that control EBV latency and lytic gene expression in EBV-transformed B and epithelial cells. We highlight newly appreciated roles of DNA methylation epigenetic machinery, host histone chaperones, the Hippo pathway, m6A RNA modification and nonsense mediated decay in control of the EBV lifecycle.

Latency remains a barrier to achieving a sterilizing cure to HIV infection. It is thus important to find new host factor(s) to better understand maintenance of HIV latency and be exploited to develop new and more efficient latency reversing agents (LRAs). Here we employed RNA interference screening with a latently HIV-1-infected cell-line to identify Stathmin 1 (STMN1) as a host factor required for maintaining HIV-1 latency. Depletion of STMN1 significantly enhanced HIV-1 expression in a STMN1 depletion-dependent manner and forced expression of exogenous STMN1 suppressed it. We further showed that STMN1 depletion increases HIV-1 proviral transcriptional elongation. Moreover, chromatin immunoprecipitation (ChIP)-qPCR assays revealed STMN1 accumulation on/near the HIV-1 5' LTR region compared to other regions on the HIV-1 provirus, suggesting the possible contribution of STMN1 to HIV-1 transcription. These results suggest that STMN1 is required for the maintenance of HIV-1 latency and implicates STMN1 as a novel therapeutic target to eradicate HIV-1.

Pepper crops in Israel are infected by poleroviruses, Pepper vein yellows virus 2 (PeVYV-2) and Pepper whitefly-borne vein yellows virus (PeWBVYV). Herein we characterize the transmission of PeWBVYV and the aphid-transmitted PeVYV-2, and show that PeWBVYV is specifically transmitted by MEAM1 species of the whitefly Bemisia tabaci, with a minimum latency period of 120 h, and not by the Mediterranean (MED). PeWBVYV and PeVYV-2 were detected in the hemolymph of MED and MEAM1, respectively, however, amounts of PeWBVYV in the hemolymph of MED or PeVYV-2 in MEAM1 were much lower than PeWBVYV in hemolymph of MEAM1. Moreover, we show that PeWBVYV does not interact with the GroEL protein of the symbiont Hamiltonella and thus does not account for the non-transmissibility by MED. An insect glycoprotein, C1QBP, interacting in vitro with the capsid proteins of both PeWBVYV and PeVYV-2 is reported which suggests a putative functional role in polerovirus transmission.

Infection with HIV-1 remains uncurable due to reservoirs of latently infected cells. Any potential cure for HIV will require a mechanism to identify and target these cells in vivo. We created a panel of Jurkat cell lines latently infected with the HIV DuoFlo virus to identify candidate biomarkers of latency. SWATH mass spectrometry was used to compare the membrane proteomes of one of the cell lines to parental Jurkat cells. Several candidate proteins with significantly altered expression were identified. The differential expression of several candidates was validated in multiple latently infected cell lines. Three factors (LAG-3, CD147,CD231) were altered across numerous cell lines, but the expression of most candidate biomarkers was variable. These results confirm that phenotypic differences in latently infected cells exists and identify additional novel biomarkers. The variable expression of biomarkers across different cell clones suggests universal antigen-based detection of latently infected cells may require a multiplex approach.

The CREB-regulated transcriptional co-activators (CRTCs), including CRTC1, CRTC2 and CRTC3, enhance transcription of CREB-targeted genes. In addition to regulating host gene expression in response to cAMP, CRTCs also increase the infection of several viruses. While human immunodeficiency virus type 1 (HIV-1) long terminal repeat (LTR) promoter harbors a cAMP response element and activation of the cAMP pathway promotes HIV-1 transcription, it remains unknown whether CRTCs have any effect on HIV-1 transcription and HIV-1 infection. Here, we reported that CRTC2 expression was induced by HIV-1 infection, but CRTC2 suppressed HIV-1 infection and diminished viral RNA expression. Mechanistic studies revealed that CRTC2 inhibited transcription from HIV-1 LTR and diminished RNA Pol II occupancy at the LTR independent of its association with CREB. Importantly, CRTC2 inhibits the activation of latent HIV-1. Together, these data suggest that in response to HIV-1 infection, cells increase the expression of CRTC2 which inhibits HIV-1 gene expression and may play a role in driving HIV-1 into latency.

New infections in general, and new viral infections amongst them, represent a serious challenge to an older organism. This review discusses the age-related alterations in responsiveness to infection from the standpoint of virus:host relationship and the host physiological whole-organism and specific immune response to the virus. Changes with age in the innate and adaptive immune system homeostasis and function are reviewed briefly. This is followed by a review of specific alterations and defects in the response of older organisms (chiefly mice and humans) to acute (particularly emerging and re-emerging) viral infections, with a very brief summary of the response to latent persistent infections. Finally, we provide a brief summary of the perspectives for possible interventions to enhance antiviral immunity.

Cytomegaloviruses (CMV) coevolve with their hosts and latently persist in the vast majority of adult mammals. Therefore, persistent T-cell responses to CMV antigens during virus latency offer a fascinating perspective on the evolution of the T-cell repertoire in natural settings. We addressed here the life-long interactions between CMV antigens presented on MHC-I molecules and the CD8 T-cell response. We present the mechanistic evidence from the murine model of CMV infection and put it in context of clinical laboratory results. We will highlight the remarkable parallels in T-cell responses between the two biological systems, and focus in particular on memory inflation as a result of competitive processes, both between viral antigenic peptides and between T-cell receptors on the host's cytotoxic lymphocytes.

A sterilizing or functional cure for HIV is currently precluded by resting CD4(+) T cells that harbor latent but replication-competent provirus. The “shock-and-kill” pharmacological ap-proach aims to reactivate provirus expression in the presence of antiretroviral therapy and target virus-expressing cells for elimination. However, no latency reversal agent (LRA) to date effectively clears viral reservoirs in humans, suggesting a need for new LRAs and LRA combinations. Here, we screened 216 compounds from the pan-African Natural Product Library and identified knipholone anthrone (KA) and its basic building block anthralin (dithranol) as novel LRAs that reverse viral latency at low micromolar concentrations in multiple cell lines. Neither agent's activity depends on protein kinase C; nor do they inhibit class I/II histone deacetylases. However, they are differentially modulated by oxidative stress and metal ions and induce distinct patterns of global gene expression from established LRAs. When applied in combination, both KA and anthralin synergize with LRAs representing multiple functional classes. Finally, KA induces both HIV RNA and protein in primary cells from HIV-infected donors. Taken together, we describe two novel LRAs that enhance the activities of multiple “shock-and-kill” agents, which in turn may inform ongoing LRA combination therapy efforts.

Cyprinid herpesvirus 2 (CyHV-2) is a highly contagious pathogen of goldfish (Carassius auratus) and Prussian carp (Carassius auratus gibelio) causing herpesviral hematopoietic necrosis. Our previous study revealed that CyHV-2 can persistently infect the kidney and spleen of goldfish that recovered from a primary infection. In this study, we tried to identify the cells persistently infected with the virus in surviving fish and investigated virus reactivation in the survivors injected with immunosuppressants, namely dexamethasone (Dex) and cyclosporine A (CsA). Virus DNA was detected from the monocytes that were isolated from the trunk kidney of the asymptomatic survivors, suggesting that monocytes/macrophages are major cells that may be persistently infected with CyHV-2. A significant increase of virus DNA levels was detected in the group injected with Dex at 10 and 21 days post-injection (dpi). In the fish group injected with CsA, the virus DNA level was the same as that in the control group at 10 dpi but increased in some organs at 21 dpi. Compared with Dex-injected fish at 10 dpi, the group injected with both Dex and CsA showed a greater increase in virus DNA levels. The gene expression of phagocytosis-associated genes, major histocompatibility complex (MHC) class II and p47phox, and anti-virus antibody levels increased in the CsA group due to virus reactivation in the infected cells but not in the Dex and Dex & CsA groups, indicating that Dex effectively suppressed monocyte/macrophage function and antibody production. In addition, recombinant interferon γ (IFNγ) supplementation in the kidney leukocyte culture that was isolated from survivors showed a reduction of virus DNA. CsA may inhibit T-helper 1 (Th1) cells and consequently IFNγ production, causing a synergetic effect with Dex on virus reactivation. The results suggest that the activity of monocytes/macrophages stimulated by IFNγ can relate to virus latency and reactivation in asymptomatic virus carriers.

Summary High rates of cognitive disorders in antiretroviral-treated people living with HIV have been described worldwide. The exact prevalence of such cognitive disorders is determined by the definitions used, and the presence of these cognitive disorders significantly impacts the overall wellbeing of people with HIV. With the cohort of people with HIV becoming increasingly older, and having high rates of comorbidities and concomitant medication use, rates of cognitive disorders are likely to increase. Conversely, interventions are being sought to reduce the size of the latent HIV reservoir. If successful, such interventions are likely to also reduce the HIV reservoir in the brain compartment, which could result in improvements in cognitive function and reduced rates of impairment.

Herpesviruses such as herpes simplex virus (HSV) type 1 and 2, varicella-zoster virus (VZV), and cytomegalovirus (CMV) maintain lifelong latency in the host after primary infection and can reactivate periodically either as asymptomatic viral shedding or as clinical disease. Immunosuppression, including biologic therapy, may increase frequency and severity of herpesvirus reactivation and infection. Licensed biologics are reviewed regarding their risks of potentiating HSV, VZV, and CMV reactivation and infection. Approaches to prophylaxis against HSV, VZV, and CMV infection or reactivation are discussed.

Latent HIV-1 persists indefinitely during antiretroviral therapy (ART) as an integrated silent genome in long-lived memory CD4+ T cells. In untreated infections, immune activation increases the turnover of intrinsically long-lived provirus-containing CD4+ T cells. Those are 'washed out' as a result of their activation, which when coupled to viral protein expression can facilitate local inflammation and recruitment of uninfected cells to activation sites, causing latently infected cells to compete for survival. De novo infection can counter this washout. During ART, inflammation and CD4+ T cell activation wane, resulting in reduced cell turnover and a persistent reservoir. We propose accelerating reservoir washout during ART by triggering sequential waves of polyclonal CD4+ T cell activation while simultaneously enhancing virus protein expression. Reservoir reduction as an adjunct to other therapies might achieve lifelong viral control.

The biology of primary lytic Kaposi's sarcoma-associated herpesvirus (KSHV) infection is still not well understood, which is largely attributed to the lack of cell lines permissive to robust lytic KSHV infection in vitro. Our study demonstrates that primary human dermal lymphatic microvascular endothelial cells (HDLMEC) support lytic KSHV replication following de novo infection, resulting in robust KSHV production, indicating that HDLMECs are suitable for studying the regulation of primary lytic KSHV infection. Importantly, by utilizing lytically infected HDLMECs, we show for the first time that the KSHV latent genes LANA and viral cyclin are required for lytic replication during de novo lytic infection, a function of these latent genes that has not yet been recognized. Since Kaposi's sarcoma is considered to be originated from infected lymphatic endothelial cells, HDLMECs represent a valuable in vitro cell culture model for investigating lytic KSHV infection, which has been understudied in KSHV pathogenesis.

Despite remarkable therapeutic advances in the past two decades, the elimination of human immunodeficiency virus type 1 (HIV-1) from latent reservoirs constitutes a major barrier to eradication and preventing neurological disease associated with HIV/AIDS. Invasion of the central nervous system (CNS) by HIV-1 occurs early in infection, leading to viral infection and productive persistence in brain macrophage-like cells (BMCs) including resident microglia and infiltrating macrophages. HIV-1 persistence in the brain and chronic neuroinflammation occur despite effective treatment with antiretroviral therapy (ART). This review examines the evidence from clinical studies, in vivo and in vitro models for HIV-1 CNS persistence, as well as therapeutic considerations in targeting latent CNS reservoirs.

Following the success of HIV-1 antiviral treatment that maintains undetectable levels of viral replication and lack of clinical progression, the design of an HIV-1 cure for patients became the next objective. The success of the treated individuals together with the identification of subjects that spontaneously control the clinical progression for long periods, such as long-term non-progressors (LTNPs) and particularly LTNP Elite Controllers (LTNP EC) have shed hope for the feasibility of a potential cure. Although a successful cure has not been attained yet, these patients have provided critical information on the mechanisms involved in the clinical control such as host genetic factors, as well as strong immune responses against the virus. Less attention has been paid to virological factors, particularly the association of the genetic variability and the control of viral infection. Considering all these studies, it has become clear that a combination of several host, immune and viral factors is needed to attain control of the viral replication control and the non-progressor clinical phenotype. Because this control can be reached through different combinations of factors, this group of individuals is not homogenous. As HIV-1 cure has been shown to be extremely difficult to achieve, a more feasible objective is the functional cure of the viral infection. After the analysis of multiple studies on the mechanisms of control in LTNP EC, we found subjects with various host protective factors and prolonged viral control. These subjects present a complete lack of evolution after more than 20-30 years of infection, stable levels of CD4+ cells (>400-500 cells/μl), a strong immune response, and no signs of clinical progression. We propose that individuals with these characteristics could have attained a functional cure of the HIV-1 infection.

Zostavax is a live-attenuated varicella zoster virus (VZV) vaccine recommended for use in adults >50 years of age to prevent shingles. The main risk factor for the development of shingles is age, which correlates with decreasing cell-mediated immunity. These data suggest a predominant role of T cell immunity in controlling VZV latency. However, other components of the immune system may also contribute. In this review, we will discuss how the immune system responds to Zostavax, focusing on recent studies examining innate immunity, transcriptomics, metabolomics, cellular, and humoral immunity.

Epstein-Barr virus (EBV) is a human herpesvirus that infects over 90% of the world's population that persists as a latent infection in various lymphoid and epithelial malignancies. The total number of EBV associated malignancies is estimated to exceed 200,000 new cancers per year. Current chemotherapeutic treatments of EBV-positive cancers include broad-spectrum cytotoxic drugs that ignore the EBV positive status of tumors and have limited safety and selectivity. In an effort to develop new and more efficacious molecules for inducing EBV reactivation, we have developed high-throughput screening assays to identify a class of small molecules (referred to as the C60 series) that efficiently activate the EBV lytic cycle in multiple latency types, including lymphoblastoid and nasopharyngeal carcinoma cell lines. In this paper we report our preliminary structure activity relationship studies and demonstrate reactivation of EBV in the SNU719 gastric carcinoma mouse model and the AGS-Akata gastric carcinoma mouse model.

Tuberculosis (TB) is a global epidemic with devastating consequences. Emerging evidence suggests that B-cells have the ability to modulate the immune response and understanding these roles during Mycobacterium tuberculosis (M.tb) infection can help to find new strategies to treat TB. The immune system of individuals with pulmonary TB form granulomas in the lung which controls the infection by inhibiting the M.tb growth and acts as a physical barrier. Thereafter, surviving M.tb become dormant and in most cases the host's immunity prevents TB reactivation. B-cells execute several immunological functions and are regarded as protective regulators of immune responses by antibody and cytokine production, as well as presenting antigen. Some of these B-cells, or regulatory B-cells, have been shown to express death-inducing ligands, such as Fas ligand (FasL). This expression and binding to the Fas receptor leads to apoptosis, a major immune regulation mechanism, in addition to the ability to induce T-cell tolerance. Here, I discuss the relevance of B-cells, in particular their non-humoral functions by addressing their regulatory properties during M.tb infection.

Even in the presence of a successful combination therapy stalling the progress of AIDS, developing a cure for this disease is still an open question. One of the major steps towards a cure would be to be able to eradicate latent HIV reservoirs present in patients. During the last decade, multiple findings point to the dominant role of the viral protein Tat in the establishment of latency. Here we present a mathematical study to understand the potential role of Tat inhibitors as virus-suppressing agents. For this aim, we implemented a computational model that reproduces intracellular dynamics. Simulating an HIV-infected cell and its intracellular feedback we observed that removing Tat protein from the system via inhibitors resulted in a temporary and reversible viral suppression. In contrast, we observed that compounds that interact with Tat protein and disrupt the integrated viral genome produced a more permanent viral suppression.

Hepatitis E virus is a zoonotic pathogen for which pigs have been identified as the main reservoir in industrialised countries. HEV infection dynamics in pig herds and pigs are influenced by several factors, including herd practices and possibly co-infection with immunomodulating viruses. This study therefore investigates the impact of porcine circovirus type 2 (PCV2) on HEV infection and transmission through experimental HEV/PCV2 co-infection of specific-pathogen-free pigs. No statistical difference between HEV-only and HEV/PCV2-infected animals was found for either the infectious period or the quantity of HEV shed in faeces. The HEV latency period was shorter for HEV/PCV2 co-infected pigs than for HEV-only infected pigs (11.6 versus 12.3 days). Its direct transmission rate was three times higher in cases of HEV/PCV2 co-infection than in cases of HEV-only infection (0.12 versus 0.04). On the other hand, the HEV transmission rate through environmental accumulation was lower in cases of HEV/PCV2 co-infection (4.3.10(-6) versus 1.5.10(-5) g/RNA copies/day for HEV-only infected pigs). The time prior to HEV seroconversion was 1.9 times longer in HEV/PCV2 co-infected pigs (49.4 versus 25.6 days for HEV-only infected pigs). In conclusion, our study shows that PCV2 affects HEV infection and transmission in pigs under experimental conditions.

The persistence of latent human immunodeficiency virus type 1 (HIV-1) reservoirs remains a major hurdle for HIV-1 eradication. The "shock and kill" strategy relies on the drug-mediated reversion of HIV-1 latency and the subsequent death of HIV-producing cells. Unfortunately, none of the agents currently in use possess a sufficient potency to reactivate latent virus or eliminate the latent HIV-1 reservoir in vivo. Here, we demonstrated that a promising specific bromodomain and extraterminal domain inhibitor, CPI-203, could potently reactivate latent HIV-1 in different latently infected cell lines with minimal cytotoxicity by activating the positive transcription elongation factor b signaling pathway. Notably, CPI-203 exhibited synergism in latent HIV-1 reactivation and alleviated the HIV-1-induced "cytokine storm" when used in combination with the protein kinase C (PKC) agonist prostratin. These findings highlight that CPI-203 shows promise as a novel, safe candidate for the design of targeted strategies to "shock and kill" HIV-1 and thus represents a potential functional cure.

MicroRNAs (miRNAs) encoded by latency-associated transcript are associated with both latent and acute stages of herpes simplex virus 2 (HSV-2) infection. In this study, miRNA-H4-5p and miRNA-H4-3p were ectopically expressed in HeLa cells to explore potential cellular targets of viral miRNAs and demonstrate their potential biological functions. The results showed that miRNA-H4-5p could reverse apoptosis induced by actinomycin D (Act-D) and promote cell cycle progression, but miRNA-H4-3p had no such obvious functions. Bioinformatics analysis, luciferase report assay, quantitative reverse transcription polymerase chain reaction (qRT-PCR), and Western blotting demonstrated that miRNA-H4-5p could bind to the 3′-untranslated region (UTR) of cyclin-dependent kinase inhibitor 2A (CDKN2A) and cyclin-dependent kinase-like 2 (CDKL2) to negatively regulate their expression. We verified that these two targeted genes were associated with cell apoptosis and cell cycle. Furthermore, in HeLa cells infected with HSV-2, we detected significantly reduced expression of CDKN2A and CDKL2 and demonstrated the negative regulation effect of miRNA-H4-5p on these two target genes. Our findings show that viral miRNAs play a vital role in regulating the expression of the host’s cellular genes that participate in cell apoptosis and progression to reshape the cellular environment in response to HSV-2 infection, providing further information on the roles of encoded herpesvirus miRNAs in pathogen–host interaction. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12250-019-00101-8) contains supplementary material, which is available to authorized users.

Histone deacetylase (HDAC) inhibitors (HDACis) have been widely tested in clinical trials for their ability to reverse HIV latency but have yielded only limited success. One HDACi, suberoylanilide hydroxamic acid (SAHA), exhibits off-target effects on host gene expression predicted to interfere with induction of HIV transcription. Romidepsin (RMD) has higher potency and specificity for class I HDACs implicated in maintaining HIV provirus in the latent state. More robust HIV reactivation has indeed been achieved with RMD use ex vivo than with SAHA; however, reduction of viral reservoir size has not been observed in clinical trials. Therefore, using RNA-Seq, we sought to compare the effects of SAHA and RMD on gene expression in primary CD4(+) T cells. Among the genes whose expression was modulated by both HDACi agents, we identified genes previously implicated in HIV latency. Two genes, SMARCB1 and PARP1, whose modulation by SAHA and RMD is predicted to inhibit HIV reactivation, were evaluated in the major maturation subsets of CD4(+) T cells and were consistently either up- or down-regulated by both HDACi compounds. Our results indicate that despite having different potencies and HDAC specificities, SAHA and RMD modulate an overlapping set of genes, implicated in HIV latency regulation. Some of these genes merit exploration as additional targets to improve the therapeutic outcomes of “shock and kill” strategies. The overall complexity of HDACi-induced responses among host genes with predicted stimulatory or inhibitory effects on HIV expression likely contributes to differential HDACi potencies and dictates the outcome of HIV reactivation.

G-quadruplexes (G4s) are noncanonical nucleic acid secondary structures formed by guanine-rich DNA and RNA sequences. In this review we aim to provide an overview of the biological roles of G4s in microbial genomes with emphasis on recent discoveries. G4s are enriched and conserved in the regulatory regions of microbes, including bacteria, fungi, and viruses. Importantly, G4s in hepatitis B virus (HBV) and hepatitis C virus (HCV) genomes modulate genes crucial for virus replication. Recent studies on Epstein–Barr virus (EBV) shed light on the role of G4s within the microbial transcripts as cis-acting regulatory signals that modulate translation and facilitate immune evasion. Furthermore, G4s in microbial genomes have been linked to radioresistance, antigenic variation, recombination, and latency. G4s in microbial genomes represent novel therapeutic targets for antimicrobial therapy., Highlights G4s display functional diversity among microbes. Their ability to influence molecular processes, including replication, transcription, translation, and recombination, has implications for the observed microbial phenotypes, including latency, virulence, rapid evolution, and radioresistance. Quadruplexes are increasingly being recognized as novel therapeutic targets in microbes. Several reports convincingly demonstrate antimicrobial activity of quadruplex-binding ligands against clinically challenging pathogens, including HIV-1, HCV, Ebola virus, Plasmodium falciparum, and Mycobacterium tuberculosis.

Latency-reversing agents (LRAs) are considered a potential strategy for curing cells of HIV-1 infection. Certain protein kinase C (PKC) activators have been previously reported to be LRAs because they can reverse HIV latency. In the present study, we examined the activities of a panel of benzolactam derivatives against cells latently infected with HIV. Using determination of p24 antigen in cell supernatants or altered intracellular GFP expression to measure HIV reactivation from latently infected cells along with a cytotoxicity assay, we found that some of the compounds exhibited latency-reversing activity, which was followed by enhanced release of HIV particles from the cells. One derivative, BL-V8-310, displayed activity in ACH-2 and J-Lat cells latently infected with HIV at a concentration of 10 nm or higher, which was superior to the activity of another highly active PKC activator, prostratin. These results were confirmed with peripheral blood cells from HIV-infected patients. We also found that these drugs up-regulate the expression of caspase 3 and enhance apoptosis specifically in latently HIV-infected cells. Moreover, combining BL-V8-310 with a bromodomain-containing 4 (BRD4) inhibitor, JQ1, not only enhanced HIV latency-reversing activity, but also reduced the effect on cytotoxic cytokine secretion from CD4+ T-cells induced by BL-V8-310 alone. Our results suggest that BL-V8-310 and its related benzolactam derivatives are potential LRA lead compounds that are effective in reversing HIV latency and reducing viral reservoirs in HIV-positive individuals with few adverse effects.

Following acute infection, herpes simplex virus type 1 (HSV-1) establishes life-long latency in sensory and other neurons. Recurrent ocular HSV-1 outbreaks are generally due to reactivation from latency. The HSV-1 latency-reactivation cycle is a complex virus-host relationship. The viral encoded latency-associated transcript (LAT) is abundantly expressed in latency and encodes several micro-RNAs and other small non-coding RNAs, which may regulate expression of key viral and cellular genes. Certain cellular signaling pathways, including Wnt/β-catenin and mTOR pathway, mediate certain aspect of the latency-reactivation cycle. Stress, via activation of the glucocorticoid receptor and other stress induced cellular transcription factors, are predicted to trigger reactivation from latency by stimulating viral gene expression and impairing immune responses and inflammation. These observations suggest stress and certain cellular signaling pathways play key roles in regulating the latency-reactivation cycle and recurrent ocular disease.

The HIV-1 reservoir is composed of cells harboring latent proviruses that have the potential to contribute to viremia upon antiretroviral treatment (ART) interruption. While this reservoir is known to be maintained by clonal expansion of infected cells, the contribution of these cell clones to residual viremia and viral rebound remains underexplored. Here, we conducted an extensive analysis on four ART-treated individuals who underwent an analytical treatment interruption (ATI), characterizing the proviral genomes and associated integration sites of large infected clones and phylogenetically linking these to plasma viremia. We show discrepancies between different assays in their ability to assess clonal expansion. Furthermore, we demonstrate that proviruses could phylogenetically be linked to plasma virus obtained before or during an ATI. This study highlights a role for HIV-infected cell clones in the maintenance of the replication-competent reservoir and suggests that infected cell clones can directly contribute to rebound viremia upon ATI. ispartof: CELL REPORTS vol:39 issue:4 ispartof: location:United States status: published

The ability of the human immunodeficiency virus type 1 (HIV-1) to establish latent infections serves as a major barrier for its cure. This process could occur when its host cells undergo apoptosis, but it is uncertain whether the components of the apoptotic pathways affect viral latency. Using the susceptible Jurkat cell line, we investigated the relationship of apoptosis-associated components with HIV-1 DNA levels using the sensitive real-time PCR assay. Here, we found that the expression of proapoptotic proteins, including Fas ligand (FasL), FADD, and p53, significantly decreased HIV-1 viral DNA in cells. In contrast, the expression of antiapoptotic molecules, such as FLIP, Bcl2, and XIAP, increased the levels of viral DNA. Furthermore, promoting cellular antiapoptotic state via the knockdown of Bax with siRNA and FADD with antisense mRNA or the treatment with the Caspase-3 inhibitor, Z-DEVD, also raised viral DNA. We also simultaneously measured viral RNA from supernatants of these cell cultures and found that HIV-1 latency is inversely proportional to viral replication. Furthermore, we demonstrated that HIV-1-infected cells that underwent the transient expression of FLIP- or XIAP-induced viral latency would then produce an increased level of viral RNA upon the reversal of these antiapoptotic effects via PMA treatment compared to LacZ control cells. Taken together, these data suggest that HIV-1 infection could be adapted to employ or even manipulate the cellular apoptotic pathway to its advantage: when the host cell remains in a pro-apoptotic state, HIV-1 favors active replication, while when the host cell prefers an anti-apoptotic state, the virus establishes viral latency and promotes latent reservoir seeding in a way which would enhance viral replication and cytopathogenesis when the cellular conditions shift to encourage the productive infection phase.

Gammaherpesviruses are ubiquitous pathogens that establish lifelong infections in the vast majority of adults worldwide. Importantly, these viruses are associated with numerous malignancies and are responsible for significant human cancer burden. These virus-associated cancers are due, in part, to the ability of gammaherpesviruses to successfully evade the innate immune response throughout the course of infection. In this review, we will summarize the current understanding of how gammaherpesviruses are detected by innate immune sensors, how these viruses evade recognition by host cells, and how this knowledge can inform novel therapeutic approaches for these viruses and their associated diseases.

IntroductionLatency reversal agents (LRAs), such as protein kinase C (PKC) agonists, constitute a promising strategy for exposing and eliminating the HIV-1 latent reservoir. PKC agonists activate NF-κB and, in turn, induce deleterious pro-inflammatory cytokine production. Adjuvant pharmacological agents, such as ruxolitinib, a JAK inhibitor, and rapamycin, an mTOR inhibitor, have previously been combined with LRAs to reduce deleterious pro-inflammatory cytokine secretion without inhibiting HIV-1 viral reactivation in vitro. Histone deacetylase inhibitors (HDACi) are known to dampen pro-inflammatory cytokine secretion in the context of other diseases and can synergize with other LRAs to bring dormant proviruses out of latency. In this study we investigated whether a broad panel of epigenetic modifiers, including HDACi, could effectively dampen PKC-induced pro-inflammatory cytokine secretion during latency reversal.MethodsWe screened an epigenetic modifier library to identify compounds that reduced intracellular IL-6 production induced by the PKC agonist Ingenol-3,20-dibenzoate. We further tested the most promising epigenetic inhibitor class, HDACi, for their ability to reduce a broad panel of pro-inflammatory cytokines and reactivate latent HIV-1 ex vivo.ResultsWe identified nine epigenetic modulators that reduced PKC-induced intracellular IL-6. In cells from aviremic individuals living with HIV-1, the HDAC1-3 inhibitor, suberohydroxamic acid (SBHA), reduced secretion of pro-inflammatory cytokines TNF-α, IL-5, IL-2r, and IL-17 but did not significantly reactivate latent HIV-1 when used in combination with Ingenol-3,20-dibenzoate.ConclusionThe addition of SBHA to Ingenol-3,20-dibenzoate reduces deleterious cytokine production during latency reversal but does not induce significant viral reactivation in aviremic donor PBMCs. The ability of SBHA to reduce PKC-induced pro-inflammatory cytokines when used in combination with Ingenol-3,20-dibenzoate suggests that SBHA can be used to reduced PKC induced pro-inflammatory cytokines but not to achieve latency reversal in the context of HIV-1.

While combination antiretroviral therapy (cART) has successfully converted HIV to a chronic but manageable infection in many parts of the world, HIV continues to persist within latent cellular reservoirs, which can become reactivated at any time to produce infectious virus. New therapies are therefore needed not only for HIV suppression but also for containing or eliminating HIV reservoirs. Compounds derived from plant, marine, and other natural products have been found to combat HIV infection and/or target HIV reservoirs, and these discoveries have substantially guided current HIV therapy-based studies. Here we summarize the role of natural product-derived compounds in current HIV suppression, remission, and cure strategies.

Transmission of HIV is known to occur by two mechanisms in vivo: the free virus pathway, where viral particles bud off an infected cell before attaching to an uninfected cell, and the cell-cell pathway, where infected cells form virological synapses through close contact with an uninfected cell. It has also been shown that HIV replication includes a positive feedback loop controlled by the viral protein Tat, which may act as a stochastic switch in determining whether an infected cell enters latency. In this paper, we introduce a simple mathematical model of HIV replication containing both the free virus and cell-cell pathways. Using this model, we demonstrate that the high multiplicity of infection in cell-cell transmission results in a suppression of latent infection, and that this modulation of latency through balancing the two transmission mechanisms can provide an evolutionary benefit to the virus. This benefit increases with decreasing overall viral fitness, which may provide a within-host evolutionary pressure toward more cell-cell transmission in late-stage HIV infection.

Although combination antiretroviral therapy (cART) is highly effective in suppressing human immunodeficiency virus type 1 (HIV-1) replication, it fails to eradicate the virus from HIV-1-infected individuals because HIV-1 integrates into the resting CD4+ T cells, establishing latently infected reservoirs. Histone deacetylation is a key element in regulating HIV-1 latent infection. Chidamide, a new anticancer drug, is a novel type of selective histone deacetylase inhibitor. Here we showed that chidamide effectively reactivated HIV-1 latent provirus in different latently infected cell lines in a dose- and time-dependent manner. Chidamide had relatively low cytotoxicity to peripheral blood mononuclear cells (PBMCs) and other latent cell lines. We have demonstrated that chidamide reactivated HIV-1 latent provirus through the NF-κB signaling pathway. The replication of the newly reactivated HIV-1 could then be effectively inhibited by the anti-HIV-1 drugs Zidovudine, Nevirapine, and Indinavir. Therefore, chidamide might be used in combination with cART for functional HIV-1 cure.

Epstein-Barr virus (EBV) is a gamma-herpesvirus that establishes lifelong infection in the majority of people worldwide. EBV uses epigenetic reprogramming to switch between multiple latency states in order to colonize the memory B-cell compartment and to then periodically undergo lytic reactivation upon plasma cell differentiation. This review focuses on recent advances in the understanding of epigenetic mechanisms that EBV uses to control its lifecycle and to subvert the growth and survival pathways that underly EBV-driven B-cell differentiation versus B-cell growth transformation, a hallmark of the first human tumor virus. These include the formation of viral super-enhancers that drive expression of key host dependency factors, evasion of tumor suppressor responses, prevention of plasmablast differentiation, and regulation of the B-cell lytic switch.

Virus infection contributes to nearly 15% of human cancers worldwide. Many of the oncogenic viruses tend to cause cancer in immunosuppressed individuals, but maintain asymptomatic, persistent infection for decades in the general population. In this review, we discuss the tactics employed by two small DNA tumor viruses, Human papillomavirus (HPV) and Merkel cell polyomavirus (MCPyV), to establish persistent infection. We will also highlight recent key findings as well as outstanding questions regarding the mechanisms by which HPV and MCPyV evade host immune control to promote their survival. Since persistent infection enables virus-induced tumorigenesis, identifying the mechanisms by which small DNA tumor viruses achieve latent infection may inform new approaches for preventing and treating their respective human cancers.

Herpes zoster is caused by the reactivation of latent varicella-zoster virus from dorsal root ganglia. Although infrequent, simultaneous damage to the anterior horn cells or anterior nerve roots at the same level may result in motor neuropathy. When motor involvement is localized in the abdominal wall, a pseudohernia may be the clinical presentation. We report a case of abdominal wall post-herpetic pseudohernia, with clinical, ultrasound and MRI correlation. MRI demonstrated increased T2/STIR signal intensity in the abdominal wall muscles, suggesting acute denervation. To our knowledge, this is the first case report of postherpetic pseudohernia with acute denervation demonstrated on MRI.

We developed a set of rabbit antisera to characterize infections by the macaque RV2 rhadinovirus homologs of KSHV. We analyzed tissues from rhesus and pig-tailed macaques naturally infected with rhesus rhadinovirus (RRV) or Macaca nemestrina rhadinovirus 2 (MneRV2). Our study demonstrates that RV2 rhadinoviruses have a tropism for epithelial cells, lymphocytes and gonadal germ cells in vivo. We observed latent infections in both undifferentiated and differentiated epithelial cells with expression of the latency marker, LANA. Expression of the early (ORF59) and late (glycoprotein B) lytic markers were detected in highly differentiated cells in epithelial ducts in oral, renal, dermal and gastric mucosal tissue as well as differentiated germ cells in male and female gonads. Our data provides evidence that epithelial and germ cell differentiation in vivo induces rhadinovirus reactivation and suggests that infected epithelial and germ cells play a role in transmission and dissemination of RV2 rhadinovirus infections in vivo.

Antiretroviral therapy can effectively block HIV-1 replication and prevent or reverse immunodeficiency in HIV-1-infected individuals. However, viral replication resumes within weeks of treatment interruption. The major barrier to a cure is a small pool of resting memory CD4+ T cells that harbor latent HIV-1 proviruses. This latent reservoir is now the focus of an intense international research effort. We describe how the reservoir is established, challenges involved in eliminating it, and pharmacologic and immunologic strategies for targeting this reservoir. The development of a successful cure strategy will most likely require understanding the mechanisms that maintain HIV-1 proviruses in a latent state and pathways that drive the proliferation of infected cells, which slows reservoir decay. In addition, a cure will require the development of effective immunologic approaches to eliminating infected cells. There is renewed optimism about the prospect of a cure, and the interventions discussed here could pave the way.

Primary effusion lymphoma (PEL), strongly linked with latent infection of Kaposi's sarcoma-associated herpesvirus (KSHV), constitutively expresses cellular interferon regulatory factor 4 (IRF4) while suppressing the expression of B cell lymphoma 6 (BCL6). Recently, it was shown that IRF4, a key transcriptional repressor of BCL6, might be a pivotal regulator of KSHV for balancing between latency and its reactivation in PEL cells. However, the action of the BCL6-IRF4 transcription factor axis during KSHV's life cycle is not clear. Herein we found that the KSHV lytic protein viral interferon regulatory factor 4 (vIRF4) dramatically enhanced the transcriptional activity of the BCL6 through the inhibition of its negative regulator IRF4. Using a chromatin immunoprecipitation (ChIP) assay, we further showed that vIRF4 bound to the specific promoter region of IRF4, contributing to a dramatic suppression of IRF4 gene expression. Correspondingly, we also found BCL6 expression to be positively and inversely correlated with vIRF4 and IRF4 expression, respectively, during KSHV reactivation. Finally, we observed that these processes require efficient KSHV lytic replication. Thus, our findings suggest a crucial role of the BCL6-IRF4 axis in triggering the transition between KSHV latency and lytic reactivation.

Ethnopharmacological relevance Current HIV therapies do not act on latent cellular HIV reservoirs; hence they are not curative. While experimental latency reversal agents (LRAs) can promote HIV expression in these cells, thereby exposing them to immune recognition, existing LRAs exhibit limited clinical efficacy and high toxicity. We previously described a traditional 3-step medicinal plant regimen used for HIV/AIDS management in Northern Botswana that inhibits HIV replication in vitro. Here we describe use of one component of the regimen that additionally contains novel phorbol esters possessing HIV latency-reversal properties. Aim of the study We sought to document experiences of traditional medicine users, assess the ability of traditional medicine components to reverse HIV latency in vitro, and identify pure compounds that conferred these activities. Materials and methods Experiences of two HIV-positive traditional medicine users (patients) were documented using qualitative interview techniques. Latency reversal activity was assessed using a cell-based model (J-Lat, clone 9.2). Crude plant extracts were fractionated by open column chromatography and reverse-phase HPLC. Compound structures were elucidated using NMR spectroscopy and mass spectrometry. Results Patients using the 3-step regimen reported improved health over several years despite no reported use of standard HIV therapies. Crude extracts from Croton megalobotrys Mull Arg. (“Mukungulu”), the third component of the 3-step regimen, induced HIV expression in J-lat cells to levels comparable to the known LRA prostratin. Co-incubation with known LRAs and pharmacological inhibitors indicated that the active agent(s) in C. megalobotrys were likely to be protein kinase C (PKC) activator(s). Consistent with these results, two novel phorbol esters (Namushen 1 and 2) were isolated as abundant components of C. megalobotrys and were sufficient to confer HIV latency reversal in vitro. Conclusion We have identified novel LRAs of the phorbol ester class from a medicinal plant used in HIV/AIDS management. These data, combined with self-reported health effects and previously-described in vitro anti-HIV activities of this traditional 3-step regimen, support the utility of longitudinal observational studies of patients undergoing this regimen to quantify its effects on plasma viral loads and HIV reservoir size in vivo.

A population at low census might go extinct, or instead transition into exponential growth to become firmly established. Whether this pivotal event occurs for a within-host pathogen can be the difference between health and illness. Here we define the principles governing whether HIV-1 spread among cells fails or becomes established, by coupling stochastic modeling with laboratory experiments. Following ex vivo activation of latently-infected CD4 T cells without de novo infection, stochastic cell division and death contributes to high variability in the magnitude of initial virus release. Transition to exponential HIV-1 spread often fails due to release of an insufficient amount of replication-competent virus. Establishment of exponential growth occurs when virus produced from multiple infected cells exceeds a critical population size. We quantitatively define the crucial transition to exponential viral spread. Thwarting this process would prevent HIV transmission or rebound from the latent reservoir.

In this issue of Molecular Cell, Hu et al. (2019) discover that coordinated regulation of AKT activity emanating from neurotrophic-factor stimulation and endogenous DNA damage maintains HSV latency. These studies provide novel insights into the role of AKT in integrating multiple signals to maintain neuronal homeostasis.

Unmethylated CpG motifs activate toll-like receptor 9 (TLR9), leading to sequence- and species-specific immune stimulation. Here, we engineered a CpG oligodeoxyribonucleotide (ODN) with multiple hydrophobic moieties, so-called lipid-modified uracil, which resulted in a facile micelle formation of the stimulant. The self-assembled CpG nanostructure (U4CpG) containing the ODN 2216 sequence was characterized by various spectroscopic and microscopic methods together with molecular dynamics simulations. Next, we evaluated the nano-immunostimulant for enhancement of anti-HIV immunity. U4CpG treatment induced activation of plasmacytoid dendritic cells (pDCs) and natural killer (NK) cells in healthy human peripheral blood, which produced type I interferons (IFNs) and IFN-γ in human peripheral blood mononuclear cells (PBMCs). Moreover, we validated the activation and promotion efficacy of U4CpG in patient-derived blood cells, and HIV-1 spread was significantly suppressed by a low dosage of the immunostimulant. Furthermore, U4CpG-treated PBMC cultured medium elicited transcription of latent HIV-1 in U1 cells indicating that U4CpG reversed HIV-1 latency. Thus, the functions of U4CpG in eradicating HIV-1 by enhancing immunity and reversing latency make the material a potential candidate for clinical studies dealing with viral infection.

Due to the emergence of multidrug-resistant bacteria, bacteriophages have become a viable alternative in controlling bacterial growth or biofilm formation. Biofilm is formed by extracellular polymeric substances (EPS) and is one of the factors responsible for increasing bacterial resistance . Bacteriophages have been studied as a bacterial control agent by use of phage enzymes or due to their bactericidal activities . A specific phage against Serratia marcescens was isolated in this work and was evaluated its biological and genomic aspects. The object of this study was UFV01, a bacteriophage belonging to the Podoviridae family, genus Teseptimavirus (group of lytic viruses), specific to the species S. marcescens , which may be related to several amino acid substitutions in the virus tail fibers. Despite this high specificity, the phage reduced the biofilm formation of several Escherichia coli strains without infecting them. UFV01 presents a relationship with phages of the genus Teseptimavirus, although it does not infect any of the E. coli strains evaluated, as these others do. All the characteristics make the phage an interesting alternative in biofilm control in hospital environments since small breaks in the biofilm matrix can lead to a complete collapse.