Targeted Restriction of Viral Gene Expression and Replication by the ZAP Antiviral System

Ficarelli, Mattia; Neil, Stuart; Swanson, Chad M.

doi:10.1146/annurev-virology-091919-104213

Cited by 50 publications

(72 citation statements)

References 115 publications

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“…Mutating Y108 partially decreased ZAP-L antiviral activity for SARS-CoV-2, which could indicate that ZAP interacts with other motifs in the viral RNA in addition to CpGs. While it is clear that ZAP binds CpG dinucleotides in the context of single-stranded RNA, how the surrounding sequence and structural context of the CpG affect ZAP binding and antiviral activity as well as the role for non-CpG binding sites in viral RNA is still unclear [ 6 , 14 , 18 – 20 , 30 , 33 , 35 , 39 , 40 ].…”

Section: Discussionmentioning

confidence: 99%

“…The zinc finger antiviral protein (ZAP, also known as PARP13 and encoded by ZC3HAV1) is a broadly active antiviral protein that is induced by both type I and II interferons and is under positive selection in primates [2][3][4][5]. It restricts RNA and DNA viruses as well as endogenous retroelements, with retroviruses and positive-strand RNA viruses being the most commonly used viral systems to study ZAP [6].…”

Section: Introductionmentioning

confidence: 99%

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S-farnesylation is essential for antiviral activity of the long ZAP isoform against RNA viruses with diverse replication strategies

et al. 2021

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The zinc finger antiviral protein (ZAP) is a broad inhibitor of virus replication. Its best-characterized function is to bind CpG dinucleotides present in viral RNAs and, through the recruitment of TRIM25, KHNYN and other cofactors, target them for degradation or prevent their translation. The long and short isoforms of ZAP (ZAP-L and ZAP-S) have different intracellular localization and it is unclear how this regulates their antiviral activity against viruses with different sites of replication. Using ZAP-sensitive and ZAP-insensitive human immunodeficiency virus type I (HIV-1), which transcribe the viral RNA in the nucleus and assemble virions at the plasma membrane, we show that the catalytically inactive poly-ADP-ribose polymerase (PARP) domain in ZAP-L is essential for CpG-specific viral restriction. Mutation of a crucial cysteine in the C-terminal CaaX box that mediates S-farnesylation and, to a lesser extent, the residues in place of the catalytic site triad within the PARP domain, disrupted the activity of ZAP-L. Addition of the CaaX box to ZAP-S partly restored antiviral activity, explaining why ZAP-S lacks antiviral activity for CpG-enriched HIV-1 despite conservation of the RNA-binding domain. Confocal microscopy confirmed the CaaX motif mediated localization of ZAP-L to vesicular structures and enhanced physical association with intracellular membranes. Importantly, the PARP domain and CaaX box together jointly modulate the interaction between ZAP-L and its cofactors TRIM25 and KHNYN, implying that its proper subcellular localisation is required to establish an antiviral complex. The essential contribution of the PARP domain and CaaX box to ZAP-L antiviral activity was further confirmed by inhibition of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replication, which replicates in double-membrane vesicles derived from the endoplasmic reticulum. Thus, compartmentalization of ZAP-L on intracellular membranes provides an essential effector function in ZAP-L-mediated antiviral activity against divergent viruses with different subcellular replication sites.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

S-farnesylation is essential for antiviral activity of the long ZAP isoform against RNA viruses with diverse replication strategies

et al. 2021

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“…17 ZAP is an antiviral protein that is broadly expressed in human cells and restricts a diverse range of viruses. 18 It was initially identified based on its ability to inhibit MLV gene expression and targets viral RNA for degradation and/or inhibits its translation. [19][20][21][22] There are several ZAP isoforms, all of which contain four N-terminal zinc finger motifs that bind RNA.…”

Section: Introductionmentioning

confidence: 99%

Minimal impact of ZAP on lentiviral vector production and transduction efficiency

Sertkaya

Hidalgo

Ficarelli

et al. 2021

Molecular Therapy - Methods & Clinical Development

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The antiviral protein ZAP binds CpG dinucleotides in viral RNA to inhibit replication. This has likely led to the CpG suppression observed in many RNA viruses, including retroviruses. Sequences added to retroviral vector genomes, such as internal promoters, transgenes, or regulatory elements, substantially increase CpG abundance. Because these CpGs could allow retroviral vector RNA to be targeted by ZAP, we analyzed whether it restricts vector production, transduction efficiency, and transgene expression. Surprisingly, even though CpG-high HIV-1 was efficiently inhibited by ZAP in HEK293T cells, depleting ZAP did not substantially increase lentiviral vector titer using several packaging and genome plasmids. ZAP overexpression also did not inhibit lentiviral vector titer. In addition, decreasing CpG abundance in a lentiviral vector genome did not increase its titer, and a gammaretroviral vector derived from murine leukemia virus was not substantially restricted by ZAP. Overall, we show that the increased CpG abundance in retroviral vectors relative to the wild-type retroviruses they are derived from does not intrinsically sensitize them to ZAP. Further understanding of how ZAP specifically targets transcripts to inhibit their expression may allow the development of CpG sequence contexts that efficiently recruit or evade this antiviral system.

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“…The zinc finger antiviral protein (ZAP, also known as ZC3HAV1 or PARP13) is an RNA binding protein that targets viral RNA containing CpG dinucleotides for degradation and inhibits their translation [1]. Unlike many viral RNA sensors, ZAP binds single stranded RNA instead of double stranded RNA, allowing it to potentially bind cellular mRNAs [2][3][4].…”

Section: Introductionmentioning

confidence: 99%

“…Unlike many viral RNA sensors, ZAP binds single stranded RNA instead of double stranded RNA, allowing it to potentially bind cellular mRNAs [2][3][4]. Furthermore, as very few cellular transcripts are devoid of CpGs, ZAP activity must be highly regulated to prevent it from targeting many cellular mRNAs and causing genome-wide changes in gene expression [1,5,6].…”

Section: Introductionmentioning

confidence: 99%

Regulation of KHNYN antiviral activity by the extended di-KH domain and nucleo-cytoplasmic trafficking

Lista

Galão

Ficarelli

et al. 2021

Preprint

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The zinc finger antiviral protein (ZAP) restricts a broad range of viruses by binding CpG dinucleotides in viral RNA to target it for degradation and inhibit its translation. KHNYN was recently identified as an antiviral protein required for ZAP to inhibit retroviral replication, though little is known about its functional determinants. KHNYN contains an N-terminal extended di-KH-like domain, a PIN endoribonuclease domain and a C-terminal CUBAN domain that binds NEDD8 and ubiquitin. We show that deletion of the extended di-KH domain reduces its antiviral activity. However, despite its similarity to RNA binding KH domains, the extended di-KH domain in KHNYN does not appear to bind RNA. Mutation of residues in the CUBAN domain that bind NEDD8 increase KHNYN abundance but do not alter its antiviral activity, suggesting that this interaction regulates KHNYN homeostatic turnover. In contrast, a CRM1-dependent nuclear export signal (NES) at the C-terminus of the CUBAN domain is required for antiviral activity. Deletion of this signal retains KHNYN in the nucleus and inhibits its interaction with ZAP. Interestingly, this NES appeared in the KHNYN lineage at a similar time as when ZAP evolved in tetrapods, indicating that these proteins may have co-evolved to restrict viral replication.

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Targeted Restriction of Viral Gene Expression and Replication by the ZAP Antiviral System

Cited by 50 publications

References 115 publications

S-farnesylation is essential for antiviral activity of the long ZAP isoform against RNA viruses with diverse replication strategies

S-farnesylation is essential for antiviral activity of the long ZAP isoform against RNA viruses with diverse replication strategies

Minimal impact of ZAP on lentiviral vector production and transduction efficiency

Regulation of KHNYN antiviral activity by the extended di-KH domain and nucleo-cytoplasmic trafficking

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