The Zinc Finger Antiviral Protein ZAP Restricts Human Cytomegalovirus and Selectively Binds and Destabilizes Viral UL4/UL5 Transcripts
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE159853 | GEO | 2021/02/01
REPOSITORIES: GEO
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Project description:Interferon-stimulated gene products (ISGs) play a crucial role in early infection control. The ISG zinc finger CCCH-type antiviral protein 1 (ZAP/ZC3HAV1) antagonizes several RNA viruses by binding to CG-rich RNA sequences, whereas its effect on DNA viruses is less well understood. Here, we decipher the role of ZAP in the context of human cytomegalovirus (HCMV) infection, a β-herpesvirus that is associated with high morbidity in immunosuppressed individuals and newborns. We show that expression of the two major isoforms of ZAP, ZAP-S and ZAP-L, is induced during HCMV infection and that both negatively affect HCMV replication. Transcriptome and proteome analyses demonstrated that the expression of ZAP results in reduced viral mRNA and protein levels and decelerates the progression of HCMV infection. Metabolic RNA labeling combined with high-throughput sequencing (SLAM-seq) revealed that most of the gene expression changes late in infection result from the general attenuation of HCMV. Furthermore, at early stages of infection, ZAP restricts HCMV by destabilizing a distinct subset of viral mRNAs, particularly those from the previously uncharacterized UL4-UL6 HCMV gene locus. Through enhanced cross-linking immunoprecipitation and sequencing analysis (eCLIP-seq), we identified the transcripts expressed from this HCMV locus as the direct targets of ZAP. Moreover, our data show that ZAP preferentially recognizes not only CG, but also other cytosine-rich sequences, thereby expanding its target specificity. In summary, this report is the first to reveal direct targets of ZAP during HCMV infection, which strongly indicates that transcripts from the UL4-UL6 locus may play an important role for HCMV replication.IMPORTANCE Viral infections have a large impact on society, leading to major human and economic losses and even global instability. So far, many viral infections, including human cytomegalovirus (HCMV) infection, are treated with a small repertoire of drugs, often accompanied by the occurrence of resistant mutants. There is no licensed HCMV vaccine in sight to protect those most at risk, particularly immunocompromised individuals or pregnant women who might otherwise transmit the virus to the fetus. Thus, the identification of novel intervention strategies is urgently required. In this study, we show that ZAP decelerates the viral gene expression cascade, presumably by selectively handpicking a distinct set of viral transcripts for degradation. Our study illustrates the potent role of ZAP as an HCMV restriction factor and sheds light on a possible role for UL4 and/or UL5 early during infection, paving a new avenue for the exploration of potential targets for novel therapies.
| S-EPMC8263000 | biostudies-literature
Project description:Interferon-stimulated gene products (ISGs) play a crucial role in early infection control. The ISG zinc finger CCCH-type antiviral protein 1 (ZAP/ZC3HAV1) antagonises several RNA viruses by binding to CG-rich RNA sequences, whereas its effect on DNA viruses is largely unknown. Here, we decipher the role of ZAP in the context of human cytomegalovirus (HCMV) infection, a β-herpesvirus that is associated with high morbidity in immunosuppressed individuals and newborns. We show that expression of the two major isoforms of ZAP, the long (ZAP-L) and short (ZAP-S), is induced during HCMV infection and that both negatively affect HCMV replication. Transcriptome and proteome analyses demonstrated that the expression of ZAP decelerates the progression of HCMV infection. SLAM-sequencing revealed that ZAP restricts HCMV at early stages of infection by destabilising a distinct subset of viral transcripts with low CG content. In summary, this report provides evidence of an important antiviral role for ZAP in host defense against HCMV infection and highlights its differentiated function during DNA virus infection.
2021-04-22 | PXD023559 | Pride
Project description:The Zinc Finger Antiviral Protein ZAP Restricts Human Cytomegalovirus and Selectively Binds and Destabilizes Viral UL4/UL5 Transcripts
| PRJNA670698 | ENA
Project description:The zinc finger antiviral protein (ZAP) depletes non-self RNAs through recognition of their elevated CpG dinucleotide content. CpG dinucleotides are sparse in most endogenous mammalian mRNAs, but a subset might potentially be modulated by ZAP. While CpG frequency alone is insufficient to predict ZAP-regulation, we developed an algorithm using experimentally determined compositional features to predict which endogenous mRNAs may be ZAP-regulated. Using ZAP-knockout mice, we demonstrate that levels of many host mRNAs that are algorithmically predicted ZAP targets are indeed increased when ZAP is absent. ZAP is interferon-inducibel and we also identify genes that are downregulated by ZAP during an innate immune response. Many ZAP-regulated gene products are extracellular matrix or of nucleosome components, whose ZAP mediated control is conserved in human cells. Overall, we provide a new tool for the prediction of ZAP target genes and reveal host mRNAs that are ZAP-regulated.
2024-09-01 | GSE274582 | GEO
Project description:The zinc finger antiviral protein (ZAP) depletes non-self RNAs through recognition of their elevated CpG dinucleotide content. CpG dinucleotides are sparse in most endogenous mammalian mRNAs, but a subset might potentially be modulated by ZAP. While CpG frequency alone is insufficient to predict ZAP-regulation, we developed an algorithm using experimentally determined compositional features to predict which endogenous mRNAs may be ZAP-regulated. Using ZAP-knockout mice, we demonstrate that levels of many host mRNAs that are algorithmically predicted ZAP targets are indeed increased when ZAP is absent. ZAP is interferon-inducibel and we also identify genes that are downregulated by ZAP during an innate immune response. Many ZAP-regulated gene products are extracellular matrix or of nucleosome components, whose ZAP mediated control is conserved in human cells. Overall, we provide a new tool for the prediction of ZAP target genes and reveal host mRNAs that are ZAP-regulated.
2024-09-01 | GSE274578 | GEO
Project description:B-cell chronic lymphocytic leukemia (B-CLL) is a heterogenous disease with a highly variable clinical course and analysis of ZAP-70 and CD38 expression on B-CLL cells allowed for identification of patients with good (ZAP-70-CD38-), intermediate (discordant expression of ZAP-70 and CD38) and poor (ZAP-70+CD38+) prognosis. In an attempt to identify a molecular basis that may underly this diverse clinical behaviour DNA microarray technology was employed to compare eight ZAP-70+CD38+ with eight ZAP-70-CD38- B-CLL cases. We used microarrays to detail the global programme of gene expression distinguising B-CLL from patient with good (samples 1 to 8) and poor prognosis (sample 9 to 16) and identified distinct classes of up- and down-regulated genes. Keywords: Disease progression
2006-09-22 | GSE4392 | GEO
Project description:Intrinsic immunity describes the set of recently discovered but poorly understood cellular mechanisms that specifically target viral pathogens. Their discovery derives in large part from intensive studies of HIV and SIV that revealed restriction factors acting at various stages of the retroviral life cycle. Recent studies indicate that some factors restrict both retroviruses and retrotransposons but surprisingly in ways that may differ. We screened known interferon-stimulated antiviral proteins previously untested for their effects on cell culture retrotransposition. Several factors, including BST2, ISG20, MAVS, MX2, and ZAP, showed strong L1 inhibition. We focused on ZAP (PARP13/ZC3HAV1), a zinc-finger protein that targets viruses of several families, including Retroviridae, Tiloviridae, and Togaviridae, and show that ZAP expression also strongly restricts retrotransposition in cell culture through loss of L1 RNA and ribonucleoprotein particle integrity. Association of ZAP with the L1 ribonucleoprotein particle is supported by co-immunoprecipitation and co-localization with ORF1p in cytoplasmic stress granules. We also used mass spectrometry to determine the protein components of the ZAP interactome, and identified many proteins that directly interact and colocalize with ZAP, including MOV10, an RNA helicase previously shown to suppress retrotransposons. The detection of a chaperonin complex, RNA degradation proteins, helicases, post-translational modifiers, and components of chromatin modifying complexes suggest mechanisms of ZAP anti-retroelement activity that function in the cytoplasm and perhaps also in the nucleus. The association of the ZAP ribonucleoprotein particle with many interferon-stimulated gene products indicates it may be a key player in the interferon response.
| S-EPMC4441479 | biostudies-literature
Project description:The host factor and interferon (IFN)-stimulated gene (ISG) product, zinc-finger antiviral protein (ZAP), inhibits a number of diverse viruses by usurping and intersecting with multiple cellular pathways. To elucidate its antiviral mechanism, we perform a loss-of-function genome-wide RNAi screen to identify cellular cofactors required for ZAP antiviral activity against the prototype alphavirus, Sindbis virus (SINV). In order to exclude off-target effects, we carry out stringent confirmatory assays to verify the top hits. Important ZAP-liaising partners identified include proteins involved in membrane ion permeability, type I IFN signaling, and post-translational protein modification. The factor contributing most to the antiviral function of ZAP is TRIM25, an E3 ubiquitin and ISG15 ligase. We demonstrate here that TRIM25 interacts with ZAP through the SPRY domain, and TRIM25 mutants lacking the RING or coiled coil domain fail to stimulate ZAP's antiviral activity, suggesting that both TRIM25 ligase activity and its ability to form oligomers are critical for its cofactor function. TRIM25 increases the modification of both the short and long ZAP isoforms by K48- and K63-linked polyubiquitin, although ubiquitination of ZAP does not directly affect its antiviral activity. However, TRIM25 is critical for ZAP's ability to inhibit translation of the incoming SINV genome. Taken together, these data uncover TRIM25 as a bona fide ZAP cofactor that leads to increased ZAP modification enhancing its translational inhibition activity.
| S-EPMC5245905 | biostudies-literature
Project description:Synonymous recoding of viral genome can attenuate their replication, but can have pleiotropic effects, with multiple mechanisms contributing to attenuation. We set out to design recoded viral genomes whose attenuation was specific and conditional. The zinc finger antiviral protein (ZAP) recognizes CpG dinucleotides and targets CpG-rich RNAs for depletion, but RNA features such as CpG numbers, spacing and surrounding nucleotide composition that enable specific modulation by ZAP are undescribed. Using synonymously mutated HIV-1 genomes, we define several sequence features that govern ZAP sensitivity and stable attenuation. Using features defined using HIV-1, we then designed a mutant enterovirus A71 genome whose attenuation was also stable and strictly ZAP-dependent, both in cell culture and in mice. This conditionally attenuated enterovirus A71 elicited neutralizing antibodies that were protective against wild-type enterovirus 71 infection and disease. Elucidation of the determinants of ZAP sensitivity can thus enable the rational design of conditionally attenuated viral vaccines.
2022-07-26 | GSE208611 | GEO
Project description:B-cell chronic lymphocytic leukemia (B-CLL) is a heterogenous disease with a highly variable clinical course and analysis of ZAP-70 and CD38 expression on B-CLL cells allowed for identification of patients with good (ZAP-70-CD38-), intermediate (discordant expression of ZAP-70 and CD38) and poor (ZAP-70+CD38+) prognosis. In an attempt to identify a molecular basis that may underly this diverse clinical behaviour DNA microarray technology was employed to compare eight ZAP-70+CD38+ with eight ZAP-70-CD38- B-CLL cases. We used microarrays to detail the global programme of gene expression distinguising B-CLL from patient with good (samples 1 to 8) and poor prognosis (sample 9 to 16) and identified distinct classes of up- and down-regulated genes. Experiment Overall Design: To compare the transcriptosomes of good prognosis CLL cases (ZAP-70-CD38-) to poor prognosis cases (ZAP-70+CD38+), we purified CD19+ cells from peripheral blood samples by immunomagnetic isolation using MidiMacs, resulting in >95% purity of leukemic cells as detected by FACS analysis of CD19+CD5+ cells. The leukemic cells were freshly purified from untreated patients and RNA was directly isolated from fresh cells without further ex vivo treatment of the cells. Eight immunomagnetically purified peripheral blood derived ZAP-70+CD38+ CLL cases were compared with eight ZAP-70-CD38- B-CLL cases.
2009-04-23 | E-GEOD-4392 | biostudies-arrayexpress