Inhibition of HIV-1 infection by TNPO3 depletion is determined by capsid and detectable after viral cDNA enters the nucleus

Iaco, Alberto De; Luban, Jeremy

doi:10.1186/1742-4690-8-98

Cited by 94 publications

(151 citation statements)

References 60 publications

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“…The decrease in WT A3F-YFP particles upon Nup153 knockdown was similar to the decrease in 2-LTR circles (1.7-vs. 2.3-fold decrease, respectively). There was no decrease in the number of nuclear A3F-YFP particles upon TNPO3 depletion in cells infected with WT or D110E virus, consistent with reports indicating that TNPO3 depletion acts on HIV-1 at a step after entry of the PIC into the nucleus (12,15,17,18). The decrease in 2-LTR circles (2.7-fold decrease) upon TNPO3 depletion, but not nuclear import of A3F-YFP particles, indicates that TNPO3 may affect the formation and/or stability of 2-LTR circles, which is consistent with a previous report (15).…”

Section: Significancesupporting

confidence: 76%

Nuclear import of APOBEC3F-labeled HIV-1 preintegration complexes

Burdick

Pathak

2013

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

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Human cytidine deaminases APOBEC3F (A3F) and APOBEC3G (A3G) are host factors that incorporate into virions and restrict virus replication. We labeled HIV-1 particles with yellow fluorescent protein (YFP)-tagged APOBEC3 proteins and examined their association with preintegration complexes (PICs) in infected cells. Labeling of PICs with A3F-YFP, and to a lesser extent A3G-YFP, could be used to visualize PICs in the nuclei, which was dependent on nuclear pore protein Nup153 but not TNPO3. We show that reverse transcription is not required for nuclear import of PICs, indicating that a viral core uncoating event associated with reverse transcription, and the central DNA flap that forms during reverse transcription, are not required for nuclear import. We also quantify association of cytoplasmic PICs with nuclear envelope (NE) and report that capsid mutations that increase or decrease core stability dramatically reduce NE association and nuclear import of PICs. In addition, we find that nuclear PICs remain close to the NE and are not distributed throughout the nuclei. These results provide tools for tracking retroviral PICs in infected cells and reveal insights into HIV-1 replication.retrovirus | microscopy | early events

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Section: Significancesupporting

confidence: 76%

Nuclear import of APOBEC3F-labeled HIV-1 preintegration complexes

Burdick

Pathak

2013

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

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“…However, previous reports differ in implicating either HIV-1 IN or CA as direct binding partners of TNPO3 (12,19,22,23). Because these studies examined the interactions employing different experimental approaches, we compared TNPO3 interactions with HIV-1 IN and CA in parallel experiments using purified recombinant proteins and pulldown assays.…”

Section: Resultsmentioning

confidence: 99%

“…Recent studies have suggested that the TNPO3 within the nucleus could be important for promoting HIV-1 integration (22)(23)(24). A mechanism was proposed whereby some PICassociated HIV-1 CA enters the nucleus and TNPO3 strips the CA from PICs to promote effective integration (24).…”

Section: Tnpo3 Does Not Interact Directly With Ca Tubes In Vitro-mentioning

confidence: 99%

“…Recent publications have extended the debate about a TNPO3-dependent mechanism in later stages of viral replication by suggesting that TNPO3 affects HIV-1 integration only after PICs enter the nucleus (22)(23)(24). One study (24) reported that some CA travels with PICs into the nucleus, after which the TNPO3-RanGTP complex strips the residual CA from the PICs to facilitate the integration and subsequently exports CA to the cytoplasm.…”

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confidence: 99%

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Interaction of the HIV-1 Intasome with Transportin 3 Protein (TNPO3 or TRN-SR2)

Larue

Gupta

Wuensch

et al. 2012

Journal of Biological Chemistry

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Background: TNPO3 is a key cellular factor involved in early steps of HIV-1 replication. Results: TNPO3 is highly structured, interacts with the HIV-1 intasome by engaging the C-terminal domain of integrase, and does not directly bind capsid tubes. Conclusion: TNPO3 interacts with HIV-1 intasomes and not capsid cores. Significance: Our findings aid future genetic analysis to elucidate the role of TNPO3 in HIV-1 replication.

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“…protein (30)(31)(32)(33). A surprising discovery that a CPSF6 deletion mutant lacking the RS domain can potently inhibit HIV-1 replication through direct binding to capsid provides for a link between Tnpo3 and a viral component (34,35).…”

mentioning

confidence: 99%

Structural basis for nuclear import of splicing factors by human Transportin 3

Maertens

Cook

Wang

et al. 2014

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

128

167

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Transportin 3 (Tnpo3, Transportin-SR2) is implicated in nuclear import of splicing factors and HIV-1 replication. Herein, we show that the majority of cellular Tnpo3 binding partners contain arginineserine (RS) repeat domains and present crystal structures of human Tnpo3 in its free as well as GTPase Ran-and alternative splicing factor/splicing factor 2 (ASF/SF2)-bound forms. The flexible β-karyopherin fold of Tnpo3 embraces the RNA recognition motif and RS domains of the cargo. A constellation of charged residues on and around the arginine-rich helix of Tnpo3 HEAT repeat 15 engage the phosphorylated RS domain and are critical for the recognition and nuclear import of ASF/SF2. Mutations in the same region of Tnpo3 impair its interaction with the cleavage and polyadenylation specificity factor 6 (CPSF6) and its ability to support HIV-1 replication. Steric incompatibility of the RS domain and RanGTP engagement by Tnpo3 provides the mechanism for cargo release in the nucleus. Our results elucidate the structural bases for nuclear import of splicing factors and the Tnpo3-CPSF6 nexus in HIV-1 biology.T he transport of macromolecules between cytoplasm and nucleus is orchestrated by a family of nuclear import and export receptors (1). Referred to as importins and exportins, these proteins bind their specific cargoes and translocate them across the nuclear pore complex. The process is regulated by the small GTPase Ran that partitions between cytoplasm and nucleus in the predominantly GDP-and GTP-bound form, respectively. Importins associate with their cargoes in the cytoplasm, and the competitive binding of RanGTP induces them to release their cargoes in the nucleus (2). Most nuclear import/export receptors belong to the β-karyopherin family of proteins, with 22 members encoded in the human genome (3). The majority of β-karyopherins bind their cargoes directly, recognizing a linear nuclear localization or export signal and/or a specific tertiary/quaternary structural feature (4, 5).A fundamentally important type of a nuclear localization signal (NLS), comprising sequences rich in Arg-Ser and/or ArgAsp/Glu/Gly dipeptides (referred to as RS or RS-like domains), belongs to the family of Ser/Arg-rich (SR) proteins. These nuclear proteins also contain RNA recognition motif (RRM) domains and play essential roles in pre-mRNA splicing and 3′ processing and participate in transcription regulation, mRNA transport, translation, and nonsense-mediated mRNA decay (6). The splicing factors alternative splicing factor/splicing factor 2 (ASF/SF2) and SC35 along with the cleavage and polyadenylation specificity factor 6 (CPSF6, also known as CF-Im-68) are among the best-characterized metazoan SR proteins (7-10). The SR protein family can be further extended by inclusion of a structurally and functionally diverse group of nuclear proteins that possess RS repeats but lack RRM domains (11). The RS domains are processively phosphorylated on their Ser residues by a set of dedicated kinases (12-16). Phosphorylation of SR proteins is thought to...

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Inhibition of HIV-1 infection by TNPO3 depletion is determined by capsid and detectable after viral cDNA enters the nucleus

Cited by 94 publications

References 60 publications

Nuclear import of APOBEC3F-labeled HIV-1 preintegration complexes

Nuclear import of APOBEC3F-labeled HIV-1 preintegration complexes

Interaction of the HIV-1 Intasome with Transportin 3 Protein (TNPO3 or TRN-SR2)

Structural basis for nuclear import of splicing factors by human Transportin 3

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