Dataset Information

CGAS-mediated stabilization of IFI16 promotes innate signaling during herpes simplex virus infection.

ABSTRACT: Interferon ?-inducible protein 16 (IFI16) and cGMP-AMP synthase (cGAS) have both been proposed to detect herpesviral DNA directly in herpes simplex virus (HSV)-infected cells and initiate interferon regulatory factor-3 signaling, but it has been unclear how two DNA sensors could both be required for this response. We therefore investigated their relative roles in human foreskin fibroblasts (HFFs) infected with HSV or transfected with plasmid DNA. siRNA depletion studies showed that both are required for the production of IFN in infected HFFs. We found that cGAS shows low production of cGMP-AMP in infected cells, but instead cGAS is partially nuclear in normal human fibroblasts and keratinocytes, interacts with IFI16 in fibroblasts, and promotes the stability of IFI16. IFI16 is associated with viral DNA and targets to viral genome complexes, consistent with it interacting directly with viral DNA. Our results demonstrate that IFI16 and cGAS cooperate in a novel way to sense nuclear herpesviral DNA and initiate innate signaling.

SUBMITTER: Orzalli MH

PROVIDER: S-EPMC4394261 | biostudies-literature | 2015 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

cGAS-mediated stabilization of IFI16 promotes innate signaling during herpes simplex virus infection.

Orzalli Megan H MH Broekema Nicole M NM Diner Benjamin A BA Hancks Dustin C DC Elde Nels C NC Cristea Ileana M IM Knipe David M DM

Proceedings of the National Academy of Sciences of the United States of America 20150323 14

Interferon γ-inducible protein 16 (IFI16) and cGMP-AMP synthase (cGAS) have both been proposed to detect herpesviral DNA directly in herpes simplex virus (HSV)-infected cells and initiate interferon regulatory factor-3 signaling, but it has been unclear how two DNA sensors could both be required for this response. We therefore investigated their relative roles in human foreskin fibroblasts (HFFs) infected with HSV or transfected with plasmid DNA. siRNA depletion studies showed that both are requ ...[more]

PMID: 25831530

Similar Datasets

Project description:UnlabelledIntrinsic 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.

Project description:The innate immune system pattern recognition receptors (PRR) are the first line of host defenses recognizing the various pathogen- or danger-associated molecular patterns and eliciting defenses by regulating the production of pro-inflammatory cytokines such as IL-1β, IL-18 or interferon β (IFN-β). NOD-like receptors (NLRs) and AIM2-like receptors (ALRs) are cytoplasmic inflammasome sensors of foreign molecules, including DNA. IFI16, a sequence-independent nuclear innate sensor ALR, recognizes episomal dsDNA genomes of herpes viruses such as KSHV, EBV, and HSV-1 in the infected cell nuclei, forms an inflammasome complex with ASC and procaspase1, and relocates into the cytoplasm leading into Caspase-1 and IL-1β generation. IFI16 also induces IFN-β during HSV-1 infection via the cytoplasmic STING-TBK1-IRF3 pathway. Thus far, whether IFI16 recognizes foreign DNA directly or utilizes other host protein(s) is unknown. Here, we demonstrate that BRCA1, a DNA damage repair sensor and transcription regulator, is in complex with IFI16 in the host cell nucleus, and their association increases in the presence of nuclear viral genomes during de novo KSHV, EBV and HSV-1 infection, and in latent KSHV or EBV infection, but not by DNA damage responses (DDR) induced by bleomycin and vaccinia virus cytoplasmic dsDNA. BRCA1 is a constituent of the triggered IFI16-inflammasome and is translocated into the cytoplasm after genome recognition along with the IFI16-inflammasome. The absence of BRCA1 abrogated IFI16-viral genome association, inflammasome assembly, IFI16 cytoplasmic localization, and Caspase-1 and IL-1β production. The absence of BRCA1 also abolished the cytoplasmic IFI16-STING interaction, downstream IRF3 phosphorylation, nuclear translocation of pIRF3 and IFN-β production during de novo KSHV and HSV-1 infection. These findings highlight that BRCA1 plays a hitherto unidentified innate immunomodulatory role by facilitating nuclear foreign DNA sensing by IFI16, subsequent assembly and cytoplasmic distribution of IFI16-inflammasomes leading into IL-1β formation and the induction of IFN-β via cytoplasmic signaling through IFI16-STING, TBK1 and IRF3.

Project description:UnlabelledHerpes simplex virus 2 (HSV-2) is one of the most common sexually transmitted infections globally, with a very high prevalence in many countries. During HSV-2 infection, viral particles become coated with complement proteins and antibodies, both present in genital fluids, which could influence the activation of immune responses. In genital mucosa, the primary target cells for HSV-2 infection are epithelial cells, but resident immune cells, such as dendritic cells (DCs), are also infected. DCs are the activators of the ensuing immune responses directed against HSV-2, and the aim of this study was to examine the effects opsonization of HSV-2, either with complement alone or with complement and antibodies, had on the infection of immature DCs and their ability to mount inflammatory and antiviral responses. Complement opsonization of HSV-2 enhanced both the direct infection of immature DCs and their production of new infectious viral particles. The enhanced infection required activation of the complement cascade and functional complement receptor 3. Furthermore, HSV-2 infection of DCs required endocytosis of viral particles and their delivery into an acid endosomal compartment. The presence of complement in combination with HSV-1- or HSV-2-specific antibodies more or less abolished HSV-2 infection of DCs. Our results clearly demonstrate the importance of studying HSV-2 infection under conditions that ensue in vivo, i.e., conditions under which the virions are covered in complement fragments and complement fragments and antibodies, as these shape the infection and the subsequent immune response and need to be further elucidated.ImportanceDuring HSV-2 infection, viral particles should become coated with complement proteins and antibodies, both present in genital fluids, which could influence the activation of the immune responses. The dendritic cells are activators of the immune responses directed against HSV-2, and the aim of this study was to examine the effects of complement alone or complement and antibodies on HSV-2 infection of dendritic cells and their ability to mount inflammatory and antiviral responses. Our results demonstrate that the presence of antibodies and complement in the genital environment can influence HSV-2 infection under in vitro conditions that reflect the in vivo situation. We believe that our findings are highly relevant for the understanding of HSV-2 pathogenesis.

Project description:The initial events after DNA virus infection involve a race between epigenetic silencing of the incoming viral DNA by host cell factors and expression of viral genes. Several host gene products, including the nuclear domain 10 (ND10) components PML (promyelocytic leukemia) and Daxx (death domain-associated protein 6), as well as IFI16 (interferon-inducible protein 16), have been shown to restrict herpes simplex virus 1 (HSV-1) replication. Whether IFI16 and ND10 components work together or separately to restrict HSV-1 replication is not known. To determine the combinatorial effects of IFI16 and ND10 proteins on viral infection, we depleted Daxx or PML in primary human foreskin fibroblasts (HFFs) in the presence or absence of IFI16. Daxx or IFI16 depletion resulted in higher ICP0 mutant viral yields, and the effects were additive. Surprisingly, small interfering RNA (siRNA) depletion of PML in the HFF cells led to decreased ICP0-null virus replication, while short hairpin RNA (shRNA) depletion led to increased ICP0-null virus replication, arguing that different PML isoforms or PML-related proteins may have restrictive or proviral functions. In normal human cells, viral DNA replication increases expression of all classes of HSV-1 genes. We observed that IFI16 repressed transcription from both parental and progeny DNA genomes. Taken together, our results show that the mechanisms of action of IFI16 and ND10 proteins are independent, at least in part, and that IFI16 exerts restrictive effects on both input and replicated viral genomes. These results raise the potential for distinct mechanisms of action of IFI16 on parental and progeny viral DNA molecules.IMPORTANCE Many human DNA viruses transcribe their genomes and replicate in the nucleus of a host cell, where they exploit the host cell nuclear machinery for their own replication. Host factors attempt to restrict viral replication by blocking such events, and viruses have evolved mechanisms to neutralize the host restriction factors. In this study, we provide information about the mechanisms of action of three host cell factors that restrict replication of herpes simplex virus (HSV). We found that these factors function independently and that one acts to restrict viral transcription from parental and progeny viral DNA genomes. These results provide new information about how cells counter DNA virus replication in the nucleus and provide possible approaches to enhance the ability of human cells to resist HSV infection.

Dataset Information

CGAS-mediated stabilization of IFI16 promotes innate signaling during herpes simplex virus infection.

Publications

cGAS-mediated stabilization of IFI16 promotes innate signaling during herpes simplex virus infection.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets