Herpesvirus-mediated stabilization of ICP0 expression neutralizes restriction by TRIM23

2021 ◽  
Vol 118 (51) ◽  
pp. e2113060118
Author(s):  
Xing Liu ◽  
Dhiraj Acharya ◽  
Eric Krawczyk ◽  
Chase Kangas ◽  
Michaela U. Gack ◽  
...  
Keyword(s):  
Viral Replication ◽  
Viral Gene Expression ◽  
Proteasomal Degradation ◽  
Viral Gene ◽  
Posttranslational Regulation ◽  
Early Proteins ◽  
Immediate Early Proteins ◽  
Simplex Virus ◽  
Functional Analyses ◽  
Hsv 1

Herpes simplex virus (HSV) infection relies on immediate early proteins that initiate viral replication. Among them, ICP0 is known, for many years, to facilitate the onset of viral gene expression and reactivation from latency. However, how ICP0 itself is regulated remains elusive. Through genetic analyses, we identify that the viral γ134.5 protein, an HSV virulence factor, interacts with and prevents ICP0 from proteasomal degradation. Furthermore, we show that the host E3 ligase TRIM23, recently shown to restrict the replication of HSV-1 (and certain other viruses) by inducing autophagy, triggers the proteasomal degradation of ICP0 via K11- and K48-linked ubiquitination. Functional analyses reveal that the γ134.5 protein binds to and inactivates TRIM23 through blockade of K27-linked TRIM23 autoubiquitination. Deletion of γ134.5 or ICP0 in a recombinant HSV-1 impairs viral replication, whereas ablation of TRIM23 markedly rescues viral growth. Herein, we show that TRIM23, apart from its role in autophagy-mediated HSV-1 restriction, down-regulates ICP0, whereas viral γ134.5 functions to disable TRIM23. Together, these results demonstrate that posttranslational regulation of ICP0 by virus and host factors determines the outcome of HSV-1 infection.

scholarly journals The HSV-1 Transcription Factor ICP4 Confers Liquid-Like Properties to Viral Replication Compartments

2021 ◽  
Vol 22 (9) ◽  
pp. 4447
Author(s):  
Michael Seyffert ◽  
Fanny Georgi ◽  
Kurt Tobler ◽  
Laurent Bourqui ◽  
Michela Anfossi ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Liquid Phase ◽  
Viral Replication ◽  
Viral Gene Expression ◽  
Intrinsically Disordered Protein ◽  
Viral Gene ◽  
Dependent Manner ◽  
Intrinsically Disordered ◽  
Simplex Virus ◽  
Hsv 1

Herpes Simplex Virus Type-1 (HSV-1) forms progeny in the nucleus within distinct membrane-less inclusions, the viral replication compartments (VRCs), where viral gene expression, DNA replication, and packaging occur. The way in which the VRCs maintain spatial integrity remains unresolved. Here, we demonstrate that the essential viral transcription factor ICP4 is an intrinsically disordered protein (IDP) capable of driving protein condensation and liquid–liquid phase separation (LLPS) in transfected cells. Particularly, ICP4 forms nuclear liquid-like condensates in a dose- and time-dependent manner. Fluorescence recovery after photobleaching (FRAP) assays revealed rapid exchange rates of EYFP-ICP4 between phase-separated condensates and the surroundings, akin to other viral IDPs that drive LLPS. Likewise, HSV-1 VRCs revealed by EYFP-tagged ICP4 retained their liquid-like nature, suggesting that they are phase-separated condensates. Individual VRCs homotypically fused when reaching close proximity and grew over the course of infection. Together, the results of this study demonstrate that the HSV-1 transcription factor ICP4 has characteristics of a viral IDP, forms condensates in the cell nucleus by LLPS, and can be used as a proxy for HSV-1 VRCs with characteristics of liquid–liquid phase-separated condensates.

scholarly journals Herpes Simplex Virus ICP27 Activation of Stress Kinases JNK and p38

2005 ◽  
Vol 79 (13) ◽  
pp. 8348-8360 ◽  
Author(s):  
Danna Hargett ◽  
Tim McLean ◽  
Steven L. Bachenheimer
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Viral Gene Expression ◽  
Immediate Early ◽  
Wild Type Virus ◽  
Viral Gene ◽  
Functional Domain ◽  
Viral Protein Synthesis ◽  
Immediate Early Proteins ◽  
Simplex Virus

ABSTRACT We previously reported that herpes simplex virus type 1 (HSV-1) can activate the stress-activated protein kinases (SAPKs) p38 and JNK. In the present study, we undertook a comprehensive and comparative analysis of the requirements for viral protein synthesis in the activation of JNK and p38. Infection with the UL36 mutant tsB7 or with UV-irradiated virus indicated that both JNK and p38 activation required viral gene expression. Cycloheximide reversal or phosphonoacetic acid treatment of wild-type virus-infected cells as well as infection with the ICP4 mutant vi13 indicated that only the immediate-early class of viral proteins were required for SAPK activation. Infection with ICP4, ICP27, or ICP0 mutant viruses indicated that only ICP27 was necessary. Additionally, we determined that in the context of virus infection ICP27 was sufficient for SAPK activation and activation of the p38 targets Mnk1 and MK2 by infecting with mutants deleted for various combinations of immediate-early proteins. Specifically, the d100 (0−/4−) and d103 (4−/22−/47−) mutants activated p38 and JNK, while the d106 (4−/22−/27−/47−) and d107 (4−/27−) mutants did not. Finally, infections with a series of ICP27 mutants demonstrated that the functional domain of ICP27 required for activation was located in the region encompassing amino acids 20 to 65 near the N terminus of the protein and that the C-terminal transactivation activity of ICP27 was not necessary.

scholarly journals Cellular Protein WDR11 Interacts with Specific Herpes Simplex Virus Proteins at thetrans-Golgi Network To Promote Virus Replication

2015 ◽  
Vol 89 (19) ◽  
pp. 9841-9852 ◽  
Author(s):  
Kathryne E. Taylor ◽  
Karen L. Mossman
Keyword(s):  
Gene Expression ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Viral Replication ◽  
Cellular Protein ◽  
Viral Gene ◽  
Late Gene Expression ◽  
Simplex Virus ◽  
Golgi Network ◽  
Hsv 1

ABSTRACTIt has recently been proposed that the herpes simplex virus (HSV) protein ICP0 has cytoplasmic roles in blocking antiviral signaling and in promoting viral replication in addition to its well-known proteasome-dependent functions in the nucleus. However, the mechanisms through which it produces these effects remain unclear. While investigating this further, we identified a novel cytoplasmic interaction between ICP0 and the poorly characterized cellular protein WDR11. During an HSV infection, WDR11 undergoes a dramatic change in localization at late times in the viral replication cycle, moving from defined perinuclear structures to a dispersed cytoplasmic distribution. While this relocation was not observed during infection with viruses other than HSV-1 and correlated with efficient HSV-1 replication, the redistribution was found to occur independently of ICP0 expression, instead requiring viral late gene expression. We demonstrate for the first time that WDR11 is localized to thetrans-Golgi network (TGN), where it interacts specifically with some, but not all, HSV virion components, in addition to ICP0. Knockdown of WDR11 in cultured human cells resulted in a modest but consistent decrease in yields of both wild-type and ICP0-null viruses, in the supernatant and cell-associated fractions, without affecting viral gene expression. Although further study is required, we propose that WDR11 participates in viral assembly and/or secondary envelopment.IMPORTANCEWhile the TGN has been proposed to be the major site of HSV-1 secondary envelopment, this process is incompletely understood, and in particular, the role of cellular TGN components in this pathway is unknown. Additionally, little is known about the cellular functions of WDR11, although the disruption of this protein has been implicated in multiple human diseases. Therefore, our finding that WDR11 is a TGN-resident protein that interacts with specific viral proteins to enhance viral yields improves both our understanding of basic cellular biology as well as how this protein is co-opted by HSV.

scholarly journals Viral DNA Sensors IFI16 and Cyclic GMP-AMP Synthase Possess Distinct Functions in Regulating Viral Gene Expression, Immune Defenses, and Apoptotic Responses during Herpesvirus Infection

mBio ◽  
2016 ◽  
Vol 7 (6) ◽  
Author(s):  
Benjamin A. Diner ◽  
Krystal K. Lum ◽  
Jared E. Toettcher ◽  
Ileana M. Cristea
Keyword(s):  
Gene Expression ◽  
Live Cell Imaging ◽  
Nuclear Periphery ◽  
Viral Gene Expression ◽  
Live Cell ◽  
Viral Gene ◽  
Viral Dna ◽  
Herpes Simplex Virus 1 ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACTThe human interferon-inducible protein IFI16 is an important antiviral factor that binds nuclear viral DNA and promotes antiviral responses. Here, we define IFI16 dynamics in space and time and its distinct functions from the DNA sensor cyclic dinucleotide GMP-AMP synthase (cGAS). Live-cell imaging reveals a multiphasic IFI16 redistribution, first to viral entry sites at the nuclear periphery and then to nucleoplasmic puncta upon herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV) infections. Optogenetics and live-cell microscopy establish the IFI16 pyrin domain as required for nuclear periphery localization and oligomerization. Furthermore, using proteomics, we define the signature protein interactions of the IFI16 pyrin and HIN200 domains and demonstrate the necessity of pyrin for IFI16 interactions with antiviral proteins PML and cGAS. We probe signaling pathways engaged by IFI16, cGAS, and PML using clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9-mediated knockouts in primary fibroblasts. While IFI16 induces cytokines, only cGAS activates STING/TBK-1/IRF3 and apoptotic responses upon HSV-1 and HCMV infections. cGAS-dependent apoptosis upon DNA stimulation requires both the enzymatic production of cyclic dinucleotides and STING. We show that IFI16, not cGAS or PML, represses HSV-1 gene expression, reducing virus titers. This indicates that regulation of viral gene expression may function as a greater barrier to viral replication than the induction of antiviral cytokines. Altogether, our findings establish coordinated and distinct antiviral functions for IFI16 and cGAS against herpesviruses.IMPORTANCEHow mammalian cells detect and respond to DNA viruses that replicate in the nucleus is poorly understood. Here, we decipher the distinct functions of two viral DNA sensors, IFI16 and cGAS, during active immune signaling upon infection with two herpesviruses, herpes simplex virus 1 (HSV-1) and human cytomegalovirus (HCMV). We show that IFI16 rapidly oligomerizes at incoming herpesvirus genomes at the nuclear periphery to transcriptionally repress viral gene expression and limit viral replicative capacity. We further demonstrate that IFI16 does not initiate upstream activation of the canonical STING/TBK-1/IRF3 signaling pathway but is required for downstream antiviral cytokine expression. In contrast, we find that, upon DNA sensing during herpesvirus infection, cGAS triggers apoptosis in a STING-dependent manner. Our live-cell imaging, mass spectrometry-based proteomics, CRISPR-based cellular assays, and optogenetics underscore the value of integrative approaches to uncover complex cellular responses against pathogens.

scholarly journals Differential Expression of Immune Checkpoint Molecules on CD8+ T Cells Specific for Immunodominant and Subdominant Herpes Simplex Virus 1 Epitopes

2019 ◽  
Vol 94 (2) ◽  
Author(s):  
Kate L. Carroll ◽  
Lyndsay Avery ◽  
Benjamin R. Treat ◽  
Lawrence P. Kane ◽  
Paul R. Kinchington ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Viral Gene Expression ◽  
Viral Reactivation ◽  
Viral Gene ◽  
Herpes Simplex Virus 1 ◽  
Latently Infected ◽  
Checkpoint Molecule ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACT Herpes simplex virus 1 (HSV-1) causes a lifelong infection of neurons that innervate barrier sites like the skin and mucosal surfaces like the eye. After primary infection of the cornea, the virus enters latency within the trigeminal ganglion (TG), from which it can reactivate throughout the life of the host. Viral latency is maintained, in part, by virus-specific CD8+ T cells that nonlethally interact with infected neurons. When CD8+ T cell responses are inhibited, HSV-1 can reactivate, and these recurrent reactivation events can lead to blinding scarring of the cornea. In the C57BL/6 mouse, CD8+ T cells specific for the immunodominant epitope from glycoprotein B maintain functionality throughout latency, while CD8+ T cells specific for subdominant epitopes undergo functional impairment that is associated with the expression of the inhibitory checkpoint molecule programmed death 1 (PD-1). Here, we investigate the checkpoint molecule T cell immunoglobulin and mucin domain-containing 3 (Tim-3), which has traditionally been associated with CD8+ T cell exhaustion. Unexpectedly, we found that Tim-3 was preferentially expressed on highly functional ganglionic CD8+ T cells during acute and latent HSV-1 infection. This, paired with data that show that Tim-3 expression on CD8+ T cells in the latently infected TG is influenced by viral gene expression, suggests that Tim-3 is an indicator of recent T cell stimulation, rather than functional compromise, in this model. We conclude that Tim-3 expression is not sufficient to define functional compromise during latency; however, it may be useful in identifying activated cells within the TG during HSV-1 infection. IMPORTANCE Without an effective means of eliminating HSV-1 from latently infected neurons, efforts to control the virus have centered on preventing viral reactivation from latency. Virus-specific CD8+ T cells within the infected TG have been shown to play a crucial role in inhibiting viral reactivation, and with a portion of these cells exhibiting functional impairment, checkpoint molecule immunotherapies have presented a potential solution to enhancing the antiviral response of these cells. In pursuing this potential treatment strategy, we found that Tim-3 (often associated with CD8+ T cell functional exhaustion) is not upregulated on impaired cells but instead is upregulated on highly functional cells that have recently received antigenic stimulation. These findings support a role for Tim-3 as a marker of activation rather than exhaustion in this model, and we provide additional evidence for the hypothesis that there is persistent viral gene expression in the HSV-1 latently infected TG.

scholarly journals Bovine Herpes Virus 1 (BHV-1) and Herpes Simplex Virus Type 1 (HSV-1) Promote Survival of Latently Infected Sensory Neurons, in Part by Inhibiting Apoptosis

2013 ◽  
Vol 6 ◽  
pp. JCD.S10803 ◽  
Author(s):  
Clinton Jones
Keyword(s):  
Sensory Neurons ◽  
Viral Gene Expression ◽  
Productive Infection ◽  
Viral Gene ◽  
Viral Transcription ◽  
Latently Infected ◽  
Simplex Virus ◽  
Cellular Transcription ◽  
Hsv 1

α-Herpesvirinae subfamily members, including herpes simplex virus type 1 (HSV-1) and bovine herpes virus 1 (BHV-1), initiate infection in mucosal surfaces. BHV-1 and HSV-1 enter sensory neurons by cell-cell spread where a burst of viral gene expression occurs. When compared to non-neuronal cells, viral gene expression is quickly extinguished in sensory neurons resulting in neuronal survival and latency. The HSV-1 latency associated transcript (LAT), which is abundantly expressed in latently infected neurons, inhibits apoptosis, viral transcription, and productive infection, and directly or indirectly enhances reactivation from latency in small animal models. Three anti-apoptosis genes can be substituted for LAT, which will restore wild type levels of reactivation from latency to a LAT null mutant virus. Two small non-coding RNAs encoded by LAT possess anti-apoptosis functions in transfected cells. The BHV-1 latency related RNA (LR-RNA), like LAT, is abundantly expressed during latency. The LR-RNA encodes a protein (ORF2) and two microRNAs that are expressed in certain latently infected neurons. Wild-type expression of LR gene products is required for stress-induced reactivation from latency in cattle. ORF2 has anti-apoptosis functions and interacts with certain cellular transcription factors that stimulate viral transcription and productive infection. ORF2 is predicted to promote survival of infected neurons by inhibiting apoptosis and sequestering cellular transcription factors which stimulate productive infection. In addition, the LR encoded microRNAs inhibit viral transcription and apoptosis. In summary, the ability of BHV-1 and HSV-1 to interfere with apoptosis and productive infection in sensory neurons is crucial for the life-long latency-reactivation cycle in their respective hosts.

scholarly journals Regulation of viral gene expression by the herpes simplex virus 1 UL24 protein (HSV-1 UL24 inhibits accumulation of viral transcripts)

Virology ◽  
2016 ◽  
Vol 495 ◽  
pp. 148-160 ◽  
Author(s):  
Carolina Sanabria-Solano ◽  
Carmen Elena Gonzalez ◽  
Nicolas Richerioux ◽  
Luc Bertrand ◽  
Slimane Dridi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Herpes Simplex Virus 1 ◽  
Simplex Virus ◽  
Hsv 1

scholarly journals Widespread remodeling of the m6A RNA-modification landscape by a viral regulator of RNA processing and export

2021 ◽  
Vol 118 (30) ◽  
pp. e2104805118
Author(s):  
Kalanghad Puthankalam Srinivas ◽  
Daniel P. Depledge ◽  
Jonathan S. Abebe ◽  
Stephen A. Rice ◽  
Ian Mohr ◽  
...  
Keyword(s):  
Gene Expression ◽  
Messenger Rna ◽  
Rna Binding ◽  
Viral Gene Expression ◽  
Rna Modification ◽  
Viral Gene ◽  
Viral Mrnas ◽  
Simplex Virus ◽  
Processing Stability ◽  
Hsv 1

N6-methyladenosine (m6A) is the most abundant internal messenger RNA (mRNA) modification, contributing to the processing, stability, and function of methylated RNAs. Methylation occurs in the nucleus during pre-mRNA synthesis and requires a core methyltransferase complex consisting of METTL3, METTL14, and WTAP. During herpes simplex virus (HSV-1) infection, cellular gene expression is profoundly suppressed, allowing the virus to monopolize the host transcription and translation apparatus and antagonize antiviral responses. The extent to which HSV-1 uses or manipulates the m6A pathway is not known. Here, we show that, in primary fibroblasts, HSV-1 orchestrates a striking redistribution of the nuclear m6A machinery that progresses through the infection cycle. METTL3 and METTL14 are dispersed into the cytoplasm, whereas WTAP remains nuclear. Other regulatory subunits of the methyltransferase complex, along with the nuclear m6A-modified RNA binding protein YTHDC1 and nuclear demethylase ALKBH5, are similarly redistributed. These changes require ICP27, a viral regulator of host mRNA processing that mediates the nucleocytoplasmic export of viral late mRNAs. Viral gene expression is initially reduced by small interfering RNA (siRNA)-mediated inactivation of the m6A methyltransferase but becomes less impacted as the infection advances. Redistribution of the nuclear m6A machinery is accompanied by a wide-scale reduction in the installation of m6A and other RNA modifications on both host and viral mRNAs. These results reveal a far-reaching mechanism by which HSV-1 subverts host gene expression to favor viral replication.

scholarly journals Dynamic Response of IFI16 and Promyelocytic Leukemia Nuclear Body Components to Herpes Simplex Virus 1 Infection

2015 ◽  
Vol 90 (1) ◽  
pp. 167-179 ◽  
Author(s):  
Roger D. Everett
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Viral Genome ◽  
Viral Gene Expression ◽  
Nuclear Body ◽  
Viral Gene ◽  
Intrinsic Resistance ◽  
Herpes Simplex Virus 1 ◽  
Simplex Virus ◽  
Hsv 1

ABSTRACTIntrinsic immunity is an aspect of antiviral defense that operates through diverse mechanisms at the intracellular level through a wide range of constitutively expressed cellular proteins. In the case of herpesviruses, intrinsic resistance involves the repression of viral gene expression during the very early stages of infection, a process that is normally overcome by viral tegument and/or immediate-early proteins. Thus, the balance between cellular repressors and virus-counteracting proteins determines whether or not a cell becomes productively infected. One aspect of intrinsic resistance to herpes simplex virus 1 (HSV-1) is conferred by components of promyelocytic leukemia nuclear bodies (PML NBs), which respond to infection by accumulating at sites that are closely associated with the incoming parental HSV-1 genomes. Other cellular proteins, including IFI16, which has been implicated in sensing pathogen DNA and initiating signaling pathways that lead to an interferon response, also respond to viral genomes in this manner. Here, studies of the dynamics of the response of PML NB components and IFI16 to invading HSV-1 genomes demonstrated that this response is extremely rapid, occurring within the first hour after addition of the virus, and that human Daxx (hDaxx) and IFI16 respond more rapidly than PML. In the absence of HSV-1 regulatory protein ICP0, which counteracts the recruitment process, the newly formed, viral-genome-induced PML NB-like foci can fuse with existing PML NBs. These data are consistent with a model involving viral genome sequestration into such structures, thereby contributing to the low probability of initiation of lytic infection in the absence of ICP0.IMPORTANCEHerpesviruses have intimate interactions with their hosts, with infection leading either to the productive lytic cycle or to a quiescent infection in which viral gene expression is suppressed while the viral genome is maintained in the host cell nucleus. Whether a cell becomes lytically or quiescently infected can be determined through the competing activities of cellular repressors and viral activators, some of which counteract cell-mediated repression. Therefore, the events that occur within the earliest stages of infection can be of crucial importance. This paper describes the extremely rapid response to herpes simplex virus 1 infection of cellular protein IFI16, a sensor of pathogen DNA, and also of the PML nuclear body proteins PML and hDaxx, as revealed by live-cell microscopy. The data imply that these proteins can accumulate on or close to the viral genomes in a sequential manner which may lead to their sequestration and repression.

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