Uracil DNA glycosylase initiates degradation of HIV-1 cDNA containing misincorporated dUTP and prevents viral integration

Weil, Amy F.; Ghosh, Dipak; Zhou, Yan; Seiple, Lauren; McMahon, Moira A.; Spivak, Adam M.; Siliciano, Robert F.; Stivers, James T.

doi:10.1073/pnas.1219702110

Cited by 65 publications

(129 citation statements)

References 64 publications

(71 reference statements)

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“…Although HIV-1 does not encode dUTPase, its Vpr protein effectively prevents UNG2-initiated uracil excision repair (31). This alternative mechanism may allow HIV-1 to alleviative negative consequences of UNG2-mediated uracil excision and downstream pathways for the integrity of HIV-1 cDNA (19).…”

Section: Discussionmentioning

confidence: 99%

“…Although the exact mechanism by which Vpr mediates the induction of replication stress is unclear, it was recently linked to the Vpr-mediated recruitment of SLX4-SLX1/MUS81-EME1 structure-specific endonucleases to DCAF1, resulting in activation of the MUS81-EME1 endonuclease and, surprisingly, DCAF1-and proteasome-dependent MUS81 degradation (26). Separately, HIV-1 and HIV-2 Vpr were reported to bind a base excision repair enzyme, uracil DNA glycosylase (UNG2), which can restrict HIV-1 infection in cells with high concentrations of dUTP (19,(27)(28)(29). HIV-1 Vpr was shown to target UNG2 to the ubiquitin proteasome pathway via the CRL4 DCAF1 E3 complex and thereby disrupt UNG2-initiated base excision repair in HIV-1-infected cells (30)(31)(32).…”

Section: Significancementioning

confidence: 99%

“…Although HIV-1 does not counteract SAMHD1 directly, it was proposed to bypass the SAMHD1-imposed restriction as a result of a more efficient reverse transcriptase that can synthesize viral cDNA in a low-dNTP environment (16). The failure of the aforementioned viral countermeasures flags HIV cDNA for processing by DNA repair enzymes, which, if not successfully completed in a low-dNTP environment set up by SAMHD1, could lead to the initiation of an innate response to viral nucleic acids, increased HIV-1 mutation rate, and/or inhibition of HIV-1 infection (17)(18)(19).…”

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confidence: 99%

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HIV-1 and HIV-2 exhibit divergent interactions with HLTF and UNG2 DNA repair proteins

Hrecka

Hao

Shun

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

103

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HIV replication in nondividing host cells occurs in the presence of high concentrations of noncanonical dUTP, apolipoprotein B mRNA-editing, enzyme-catalytic, polypeptide-like 3 (APOBEC3) cytidine deaminases, and SAMHD1 (a cell cycle-regulated dNTP triphosphohydrolase) dNTPase, which maintains low concentrations of canonical dNTPs in these cells. These conditions favor the introduction of marks of DNA damage into viral cDNA, and thereby prime it for processing by DNA repair enzymes. Accessory protein Vpr, found in all primate lentiviruses, and its HIV-2/simian immunodeficiency virus (SIV) SIVsm paralogue Vpx, hijack the CRL4DCAF1 E3 ubiquitin ligase to alleviate some of these conditions, but the extent of their interactions with DNA repair proteins has not been thoroughly characterized. Here, we identify HLTF, a postreplication DNA repair helicase, as a common target of HIV-1/SIVcpz Vpr proteins. We show that HIV-1 Vpr reprograms CRL4DCAF1 E3 to direct HLTF for proteasome-dependent degradation independent from previously reported Vpr interactions with base excision repair enzyme uracil DNA glycosylase (UNG2) and crossover junction endonuclease MUS81, which Vpr also directs for degradation via CRL4DCAF1 E3. Thus, separate functions of HIV-1 Vpr usurp CRL4DCAF1 E3 to remove key enzymes in three DNA repair pathways. In contrast, we find that HIV-2 Vpr is unable to efficiently program HLTF or UNG2 for degradation. Our findings reveal complex interactions between HIV-1 and the DNA repair machinery, suggesting that DNA repair plays important roles in the HIV-1 life cycle. The divergent interactions of HIV-1 and HIV-2 with DNA repair enzymes and SAMHD1 imply that these viruses use different strategies to guard their genomes and facilitate their replication in the host.

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Section: Discussionmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

mentioning

confidence: 99%

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HIV-1 and HIV-2 exhibit divergent interactions with HLTF and UNG2 DNA repair proteins

Hrecka

Hao

Shun

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

103

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“…Given our findings that dUTP is an excellent substrate, there must be undiscovered regulatory mechanisms that allow dUTP levels to persist or accumulate after the other dNTPs have been depleted. Because dUTP incorporation into HIV-1 cDNA has been implicated as a related antiviral defense mechanism (28,29), additional understanding of the mechanisms that regulate nucleotide pools during the transition from active to resting states of cells will likely be relevant to viral infectivity and persistence in these cell types.…”

Section: Discussionmentioning

confidence: 99%

GTP activator and dNTP substrates of HIV-1 restriction factor SAMHD1 generate a long-lived activated state

Hansen

Seamon

Cravens

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

187

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The HIV-1 restriction factor sterile α-motif/histidine-aspartate domain-containing protein 1 (SAMHD1) is a tetrameric protein that catalyzes the hydrolysis of all dNTPs to the deoxynucleoside and tripolyphosphate, which effectively depletes the dNTP substrates of HIV reverse transcriptase. Here, we establish that SAMHD1 is activated by GTP binding to guanine-specific activator sites (A1) as well as coactivation by substrate dNTP binding to a distinct set of nonspecific activator sites (A2). Combined activation by GTP and dNTPs results in a long-lived tetrameric form of SAMHD1 that persists for hours, even after activating nucleotides are withdrawn from the solution. These results reveal an ordered model for assembly of SAMHD1 tetramer from its inactive monomer and dimer forms, where GTP binding to the A1 sites generates dimer and dNTP binding to the A2 and catalytic sites generates active tetramer. Thus, cellular regulation of active SAMHD1 is not determined by GTP alone but instead, the levels of all dNTPs and the generation of a persistent tetramer that is not in equilibrium with free activators. The significance of the long-lived activated state is that SAMHD1 can remain active long after dNTP pools have been reduced to a level that would lead to inactivation. This property would be important in resting CD4 + T cells, where dNTP pools are reduced to nanomolar levels to restrict infection by HIV-1. dNTP induced oligomerization | enzyme catalysis | innate immunity T he steady-state composition and concentration of deoxynucleotide triphosphate pools in mammalian cells are highly regulated because of the mutagenic consequences of dNTP imbalances in dividing cells (1, 2) as well as the important antiviral effects of dNTP pool depletion in quiescent cells (3,4). In all cell types, the ultimate pool balance is determined by dNTP-dependent regulatory pathways that affect the activities of enzymes involved in both synthesis and degradation of dNTPs (5-7). The most important highly up-regulated synthetic enzyme during S phase of dividing cells is the R1/R2 isoform of ribonucleotide triphosphate reductase, which ensures that dNTP precursors are plentiful for DNA synthesis (8). However, in quiescent cells of the immune system (resting CD4 + T cells, macrophages, and dendritic cells), where dNTP pools are ∼10-fold lower than dividing cells, the ultimate pool levels are likely determined by a balance between the activities of the R1/p53R2 isoform of ribonucleotide triphosphate reductase and the degradative dNTP triphosphohydrolase sterile α-motif/histidineaspartate domain-containing protein 1 (SAMHD1) (9). The highly dynamic nature of dNTP pools demands finely tuned mechanisms for feedback regulation of these enzymes by dNTPs as well as coarse regulatory mechanisms (posttranslational modifications, transcriptional regulation, and proteasomal targeting) that serve to turn these activities on and off at appropriate stages of the cell cycle and in specific cell types (10, 11). dNTP triphosphohydrolase enzymes, such as SAMHD1,...

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“…Work from several laboratories showed that UNG2 has a positive impact on HIV-1 (39,82,83). In contrast, some laboratories found that UNG2 hinders HIV-1 (84,85), whereas others found UNG2 to have no measurable impact on the virus (7,86). Interestingly, one work linked the requirement for UNG2 to coreceptor usage (83).…”

Section: Resultsmentioning

confidence: 99%

Cullin4A and Cullin4B Are Interchangeable for HIV Vpr and Vpx Action through the CRL4 Ubiquitin Ligase Complex

Sharifi

Furuya

Jellinger

et al. 2014

J Virol

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IMPORTANCEThe work presented here shows for the first time that HIV Vpr and Vpx do not rely exclusively on CUL4A to cause ubiquitination through the CRL4 ubiquitin ligase complex. Furthermore, our finding that intracellular CUL4 and SAMHD1 distributions can vary with cell type provides the basis for reconciling previous disparate findings regarding the site of SAMHD1 depletion. Finally, our observations with primary immune cells provide insight into the cell biology of CUL4A and CUL4B that will help differentiate the functions of these similar proteins.

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Uracil DNA glycosylase initiates degradation of HIV-1 cDNA containing misincorporated dUTP and prevents viral integration

Cited by 65 publications

References 64 publications

HIV-1 and HIV-2 exhibit divergent interactions with HLTF and UNG2 DNA repair proteins

HIV-1 and HIV-2 exhibit divergent interactions with HLTF and UNG2 DNA repair proteins

GTP activator and dNTP substrates of HIV-1 restriction factor SAMHD1 generate a long-lived activated state

Cullin4A and Cullin4B Are Interchangeable for HIV Vpr and Vpx Action through the CRL4 Ubiquitin Ligase Complex

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