The nuclear localization signal of CPSF6 governs post-nuclear import steps of HIV-1 infection

PLoS Pathogens, Journal Year: 2025, Volume and Issue: 21(1), P. e1012354 - e1012354

Published: Jan. 17, 2025

The early stages of HIV-1 infection include the trafficking viral core into nucleus infected cells. However, much remains to be understood about how accomplishes nuclear import and consequences pathways utilized on events. host factor cleavage polyadenylation specificity 6 (CPSF6) assists localization post-entry integration targeting. Here, we used a CPSF6 truncation mutant lacking functional signal (NLS), CPSF6-358, appended heterologous NLSs rescue localization. We show that some, but not all, drive CPSF6-358 nucleus. Interestingly, found some localized CPSF6-NLS chimeras supported inefficient infection. still enters in these cell lines fails traffic speckle-associated domains (SPADs). Additionally, efficiently integrate lines. Collectively, our results demonstrate NLS facilitates steps subsequent additionally identify ability canonical sequences influence cargo following import.

Language: Английский

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Structural insights into HIV-2 CA lattice formation and FG-pocket binding revealed by single-particle cryo-EM

Cell Reports, Journal Year: 2025, Volume and Issue: 44(2), P. 115245 - 115245

Published: Jan. 25, 2025

Language: Английский

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CPSF6 Promotes HIV-1 Preintegration Complex Function

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Jan. 29, 2025

ABSTRACT Cleavage and polyadenylation specificity factor 6 (CPSF6) is part of the cellular cleavage I mammalian (CFIm) complex that regulates mRNA processing polyadenylation. CPSF6 also functions as a HIV-1 capsid (CA) binding host promotes viral DNA integration targeting into gene dense regions genome. However, effects on activity preintegration (PIC) - machinery carries out to establish infection unknown. To study CPSF6’s role in PIC function, we extracted PICs from cells depleted or expressing mutant cannot bind CA. These exhibited significantly lower when compared control PICs. Addition recombinant restored cells, suggesting direct function. solidify effect inoculated CPSF6-depleted CPSF6-mutant with particles measured A significant reduction these was detected this defect not consequence reduced reverse transcription nuclear entry. Additionally, viruses deficient CA-CPSF6 showed no cells. Finally, sequencing analysis revealed redirected Collectively, results suggest CPSF6-CA interaction function both vitro infected IMPORTANCE dependent virus factors. molecular details virus-host interactions are fully understood. For instance, provides interfaces for several one such capsid-binding factor, whose regulate Initial work identified truncated cytosolic form restricted HIV by blocking it now established full-length primarily Here report complexes (PICs). We observed disruption target directed away gene-dense regions. findings demonstrate critical targeting.

Language: Английский

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Recent Advances in DNA‐Templated Protein Patterning

Small Methods, Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 2, 2025

Abstract In recent decades, the advancement of DNA nanotechnology enables precise nanoscale organization diverse functional materials with templates. Particularly, a variety DNA‐templated protein patterns are constructed as powerful tools for programming biomimetic complexes. this review, progress in patterning, including cutting‐edge methods arranging proteins templates, and across varying dimensions briefly summarized. Representative applications biological analysis biomedicine discussed. DNA‐protein programmable dynamics, which hold promise precision diagnosis therapeutics highlighted. Finally, current challenges opportunities fabrication application pattering

Language: Английский

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Spatiotemporal binding of cyclophilin A and CPSF6 to capsid regulates HIV-1 nuclear entry and integration

mBio, Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 27, 2025

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) capsid, which is the target of antiviral lenacapavir, protects viral genome and binds multiple host proteins to influence intracellular trafficking, nuclear import, integration. Previously, we showed that capsid binding cleavage polyadenylation specificity factor 6 (CPSF6) in cytoplasm competitively inhibited by cyclophilin A (CypA) regulates infection. Here, determined a mutant with increased CypA affinity had significantly reduced entry mislocalized However, disruption restored entry, integration, infection CPSF6-dependent manner. Furthermore, relocalization expression from cell nucleus failed restore HIV-1 Our results clarify sequential CPSF6 required for optimal integration targeting, providing insights development antiretroviral therapies, such as lenacapavir. IMPORTANCE (HIV) encodes protein forms conical shell, called surrounds its genome. The has been shown protect innate immune sensors cell, help transport toward into nucleus, keep components reverse transcription together conversion RNA DNA, DNA specific regions In this study, show HIV hijacks two bind sequentially order choreograph precise timing these replication steps. Disruption or their location leads defective Mutations exist infected individuals may reduce efficacy drugs capsid.

Language: Английский

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Correlative In Situ Cryo-ET Reveals Cellular and Viral Remodeling Associated with Selective HIV-1 Core Nuclear Import

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: March 4, 2025

Abstract Lentiviruses like HIV-1 infect non-dividing cells by traversing the nuclear pore, but studying this process has been challenging due to its scarcity and dynamic nature in infected cells. Here, we developed a robust cell-permeabilization system that recapitulates import established an integrated cryo-correlative workflow combining cryo-CLEM, cryo-FIB, cryo-ET for targeted imaging of process. These advancements enabled successful capture 1,899 cores at various stages import. Statistical structural analyses native wild-type mutant revealed depends on both capsid elasticity pore adaptability, as well factors such CPSF6. Brittle fail enter complex (NPC), while CPSF6-binding-deficient stall inside NPC, resulting impaired Intriguingly, pores function selective filters favoring smaller, tube-shaped cores. Our study opens new avenues dissecting biochemistry biology downstream events including core uncoating potentially integration, with unprecedented detail.

Language: Английский

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Nuclear warfare: pathogen manipulation of the nuclear pore complex and nuclear functions

mBio, Journal Year: 2025, Volume and Issue: unknown

Published: March 20, 2025

ABSTRACT Viruses and bacteria exploit the nuclear pore complex (NPC) host functions to bypass cellular barriers manipulate essential processes. frequently engage directly with NPC components, such as nucleoporins, enable genome import evade immune defenses. In contrast, bacterial pathogens rely on secreted effector proteins disrupt transport reprogram transcription. These strategies reflect a remarkable evolutionary convergence, both types of targeting promote infection. This minireview explores overlapping unique mechanisms by which hijack nucleus, shedding light their roles in disease potential avenues for therapeutic intervention.

Language: Английский

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CPSF6 promotes HIV-1 preintegration complex function

Journal of Virology, Journal Year: 2025, Volume and Issue: unknown

Published: April 9, 2025

ABSTRACT Cleavage and polyadenylation specificity factor 6 (CPSF6) is part of the cellular cleavage I mammalian (CFIm) complex that regulates mRNA processing polyadenylation. CPSF6 also functions as an HIV-1 capsid (CA) binding host to promote viral DNA integration targeting into gene-dense regions genome. However, effects on activity preintegration (PIC)—the sub-viral machinery carries out integration—are unknown. To study CPSF6’s role in PIC function, we extracted PICs from cells are either depleted or express a mutant form cannot bind CA. These exhibited significantly lower when compared control PICs. The addition purified recombinant restored CPSF6-mutant cells, suggesting direct function. solidify inoculated CPSF6-depleted with particles measured A significant reduction these was detected, this not consequence reverse transcription nuclear entry. Additionally, viruses deficient CA–CPSF6 showed no defect cells. Finally, sequencing analysis revealed cell genomes redirected away chromatin Collectively, results suggest CPSF6–CA interaction promotes function both vitro infected IMPORTANCE infection dependent virus factors. molecular details HIV–host interactions fully understood. For instance, provides interfaces for several one such capsid-binding factor, whose regulate Initial work identified truncated cytosolic restrict HIV by blocking it now established full-length primarily Here, provide evidence stimulates complexes (PICs). We describe disruption target reduces redirects low transcriptional activity. findings identify critical targeting.

Language: Английский

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Impact of HIV-1 capsid polymorphisms on viral infectivity and susceptibility to lenacapavir

mBio, Journal Year: 2025, Volume and Issue: unknown

Published: April 17, 2025

ABSTRACT Lenacapavir (LEN) is a first-in-class capsid (CA) inhibitor for the treatment and prevention of HIV-1 infection. While LEN has shown potent antiviral activity across all major subtypes, impact existing CA sequence diversity on remains to be determined. Here, we identified natural polymorphisms within LEN-binding site assessed each their viral infectivity susceptibility LEN. Using co-crystal structure in complex with hexamer, 29 binding residues five angstroms analyzed naturally occurring multiclade collection >10,000 unique gag sequences. Eleven these residues, including (M66, Q67, K70, N74, A105) previously associated resistance when mutated, were invariant The remaining 18 showed one or more substitutions ≥0.5% prevalence total 54 polymorphisms. When introduced as site-directed mutants (SDMs) an NL4.3-based reporter virus evaluated drug MT-4 cells, 74% (40/54) impaired (0.01%–77% wild type), 96% (46/48) exhibiting minimal change (less than threefold) L56V N57H conferred high-level (72- 4,890-fold, respectively), both variants diminished replication capacity primary T-cells relative wild-type virus. Collectively, results indicate that rare should minimally efficacy treatment-naïve individuals. IMPORTANCE protein mediates multiple essential functions throughout cycle, making it attractive target therapeutic intervention. (LEN), inhibitor, being long-acting option ongoing clinical studies HIV prevention. Twice-yearly lenacapavir approved countries adults multi-drug-resistant combination other antiretrovirals, its investigational use pre-exposure prophylaxis 99.9%–100% preventing infection among broad geographically diverse range study participants. In this report, investigated how may sensitivity Our data demonstrate high conservation large variants, majority having detrimental effect

Language: Английский

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HIV-1 Capsid Rapidly Induces Long-Lived CPSF6 Puncta in Non-Dividing Cells, but Similar Puncta Already Exist in Uninfected T-Cells

Viruses, Journal Year: 2024, Volume and Issue: 16(5), P. 670 - 670

Published: April 25, 2024

The HIV-1 capsid (CA) protein forms the outer shell of viral core that is released into cytoplasm upon infection. CA binds various cellular proteins, including CPSF6, direct integration speckle-associated domains in host chromatin. Upon infection, CPSF6 puncta nucleus. Here, we characterised these further HeLa cells, T-cells and macrophages confirmed reverse transcription are not required for formation. Indeed, found formed very rapidly after correlating with time entered In aphidicolin-treated cells macrophages, were detected length experiment, suggesting only lost cell division. still co-localised at latest points, considerably peak integration. Intriguingly, number induced did correlate MOI or total nuclear speckles present each cell, CA/CPSF6 directed to a few speckles. Furthermore, already uninfected T-cells, promotes natural behaviour CPSF6.

Language: Английский

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