APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction

Salamango, Daniel J.; Becker, Jordan T.; McCann, John D.; Cheng, Adam Z; Demir, Özlem; Amaro, Rommie E.; Brown, William L.; Shaban, Nadine M.; Harris, Reuben S.

doi:10.1128/mcb.00356-18

Cited by 17 publications

(15 citation statements)

References 100 publications

(89 reference statements)

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“…We therefore performed immunofluorescent (IF) microscopy studies of U2OS cells overexpressing A3-mCherry constructs with either empty vector or BORF2-FLAG. As reported, A3B is nuclear, A3A has a cell-wide localization, A3H is cytoplasmic and nucleolar, and the other A3s are cytoplasmic [46][47][48][49][50]. Also as expected, BORF2 caused a robust and complete relocalization of nuclear A3B to perinuclear aggregates (Fig 2).…”

Section: Ebv Borf2 and Kshv Orf61 Bind And Relocalize Both A3b And A3asupporting

confidence: 85%

A Conserved Mechanism of APOBEC3 Relocalization by Herpesviral Ribonucleotide Reductase Large Subunits

Cheng

Moraes

Attarian

et al. 2019

J Virol

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An integral part of the antiviral innate immune response is the APOBEC3 family of single-stranded DNA cytosine deaminases, which inhibits virus replication through deamination-dependent and -independent activities. Viruses have evolved mechanisms to counteract these enzymes, such as HIV-1 Vif-mediated formation of a ubiquitin ligase to degrade virus-restrictive APOBEC3 enzymes. A new example is Epstein-Barr virus (EBV) ribonucleotide reductase (RNR)-mediated inhibition of cellular APOBEC3B (A3B). The large subunit of the viral RNR, BORF2, causes A3B relocalization from the nucleus to cytoplasmic bodies and thereby protects viral DNA during lytic replication. Here, we use coimmunoprecipitation and immunofluorescence microscopy approaches to ask whether this mechanism is shared with the closely related gammaherpesvirus Kaposi’s sarcoma-associated herpesvirus (KSHV) and the more distantly related alphaherpesvirus herpes simplex virus 1 (HSV-1). The large RNR subunit of KSHV, open reading frame 61 (ORF61), coprecipitated multiple APOBEC3s, including A3B and APOBEC3A (A3A). KSHV ORF61 also caused relocalization of these two enzymes to perinuclear bodies (A3B) and to oblong cytoplasmic structures (A3A). The large RNR subunit of HSV-1, ICP6, also coprecipitated A3B and A3A and was sufficient to promote the relocalization of these enzymes from nuclear to cytoplasmic compartments. HSV-1 infection caused similar relocalization phenotypes that required ICP6. However, unlike the infectivity defects previously reported for BORF2-null EBV, ICP6 mutant HSV-1 showed normal growth rates and plaque phenotypes. Combined, these results indicate that both gamma- and alphaherpesviruses use a conserved RNR-dependent mechanism to relocalize A3B and A3A and furthermore suggest that HSV-1 possesses at least one additional mechanism to neutralize these antiviral enzymes. IMPORTANCE The APOBEC3 family of DNA cytosine deaminases constitutes a vital innate immune defense against a range of different viruses. A novel counterrestriction mechanism has recently been uncovered for the gammaherpesvirus EBV, in which a subunit of the viral protein known to produce DNA building blocks (ribonucleotide reductase) causes A3B to relocalize from the nucleus to the cytosol. Here, we extend these observations with A3B to include a closely related gammaherpesvirus, KSHV, and a more distantly related alphaherpesvirus, HSV-1. These different viral ribonucleotide reductases also caused relocalization of A3A, which is 92% identical to A3B. These studies are important because they suggest a conserved mechanism of APOBEC3 evasion by large double-stranded DNA herpesviruses. Strategies to block this host-pathogen interaction may be effective for treating infections caused by these herpesviruses.

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Section: Ebv Borf2 and Kshv Orf61 Bind And Relocalize Both A3b And A3asupporting

confidence: 85%

A Conserved Mechanism of APOBEC3 Relocalization by Herpesviral Ribonucleotide Reductase Large Subunits

Cheng

Moraes

Attarian

et al. 2019

J Virol

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“…Disrupting RNA binding prevents dimerization, and monomeric A3H may be susceptible to increased processing by ubiquitination. Indeed, mutagenesis of RNA binding residues results in decreased expression, nuclear localization, and a loss of packaging and antiviral activity [25][26][27]40,41], suggesting a shared mechanism of instability and loss of function to the R105G and N15del mutations. Recent biochemical work has more directly implicated nucleic acid binding as critical for A3H antiviral activity and is required for stabilization, virion incorporation, deaminase-independent inhibition of reverse transcriptase, and ssDNA substrate recognition [42].…”

Section: Discussionmentioning

confidence: 99%

Polymorphisms in Human APOBEC3H Differentially Regulate Ubiquitination and Antiviral Activity

Chesarino

Emerman

2020

Viruses

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The APOBEC3 family of cytidine deaminases are an important part of the host innate immune defense against endogenous retroelements and retroviruses like Human Immunodeficiency Virus (HIV). APOBEC3H (A3H) is the most polymorphic of the human APOBEC3 genes, with four major haplotypes circulating in the population. Haplotype II is the only antivirally-active variant of A3H, while the majority of the population possess independently destabilizing polymorphisms present in haplotype I (R105G) and haplotypes III and IV (N15del). In this paper, we show that instability introduced by either polymorphism is positively correlated with degradative ubiquitination, while haplotype II is protected from this modification. Inhibiting ubiquitination by mutating all of the A3H lysines increased the expression of haplotypes III and IV, but these stabilized forms of haplotype III and IV had a strict nuclear localization, and did not incorporate into virions, nor exhibit antiviral activity. Fusion chimeras with haplotype II allowed for stabilization, cytoplasmic retention, and packaging of the N15del-containing haplotype III, but the haplotype III component of these chimeras was unable to restrict HIV-1 on its own. Thus, the evolutionary loss of A3H activity in many humans involves functional deficiencies independent of protein stability.

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“…Furthermore, RNA binding has been implicated for proper subcellular localization of A3F, A3G, and A3H [ 33 , 34 , 55 – 61 ]. Treatment with RNase A can disrupt interactions between A3s and cellular proteins, hinting at RNA playing an important role in regulating A3 activities [ 33 – 35 , 62 ].…”

Section: Discussionmentioning

confidence: 99%

“…This correlates with weaker RNA binding for A3G in comparison to A3C-A3H (Fig 4), which has less disparity between the charge of the two domains. Furthermore, RNA binding has been implicated for proper subcellular localization of A3F, A3G, and A3H [33,34,[55][56][57][58][59][60][61]. Treatment with RNase A can disrupt interactions between A3s and cellular proteins, hinting at RNA playing an important role in regulating A3 activities [33][34][35]62].…”

Section: Plos Pathogensmentioning

confidence: 99%

Highly-potent, synthetic APOBEC3s restrict HIV-1 through deamination-independent mechanisms

et al. 2021

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The APOBEC3 (A3) genes encode cytidine deaminase proteins with potent antiviral and anti-retroelement activity. This locus is characterized by duplication, recombination, and deletion events that gave rise to the seven A3s found in primates. These include three single deaminase domain A3s (A3A, A3C, and A3H) and four double deaminase domain A3s (A3B, A3D, A3F, and A3G). The most potent of the A3 proteins against HIV-1 is A3G. However, it is not clear if double deaminase domain A3s have a generalized functional advantage to restrict HIV-1. In order to test whether superior restriction factors could be created by genetically linking single A3 domains into synthetic double domains, we linked A3C and A3H single domains in novel combinations. We found that A3C/A3H double domains acquired enhanced antiviral activity that is at least as potent, if not better than, A3G. Although these synthetic double domain A3s package into budding virions more efficiently than their respective single domains, this does not fully explain their gain of antiviral potency. The antiviral activity is conferred both by cytidine-deaminase dependent and independent mechanisms, with the latter correlating to an increase in RNA binding affinity. T cell lines expressing this A3C-A3H super restriction factor are able to control replicating HIV-1ΔVif infection to similar levels as A3G. Together, these data show that novel combinations of A3 domains are capable of gaining potent antiviral activity to levels similar to the most potent genome-encoded A3s, via a primarily non-catalytic mechanism.

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APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction

Cited by 17 publications

References 100 publications

A Conserved Mechanism of APOBEC3 Relocalization by Herpesviral Ribonucleotide Reductase Large Subunits

A Conserved Mechanism of APOBEC3 Relocalization by Herpesviral Ribonucleotide Reductase Large Subunits

Polymorphisms in Human APOBEC3H Differentially Regulate Ubiquitination and Antiviral Activity

Highly-potent, synthetic APOBEC3s restrict HIV-1 through deamination-independent mechanisms

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