HIV-1 uncoating by release of viral cDNA from capsid-like structures in the nucleus of infected cells

eLife, Journal Year: 2021, Volume and Issue: 10

Published: April 27, 2021

HIV-1 replication commences inside the cone-shaped viral capsid, but timing, localization, and mechanism of uncoating are under debate. We adapted a strategy to visualize individual reverse-transcribed cDNA molecules their association with cellular proteins using fluorescence correlative-light-and-electron-microscopy (CLEM). specifically detected nuclei, not in cytoplasm. Nuclear initially co-localized fluorescent integrase fusion (IN-FP) CA (capsid) protein, cDNA-punctae separated from IN-FP/CA over time. This phenotype was conserved primary target cells, nuclear complexes exhibiting strong CA-signals all cell types. CLEM revealed capsid-like structures apparently broken capsid-remnants at position IN-FP signals elongated chromatin-like punctae lacking IN-FP. Our data argue for by physical disruption rather than cooperative disassembly CA-lattice, followed separation pre-integration complex.

Language: Английский

---

Clustering and reverse transcription of HIV‐1 genomes in nuclear niches of macrophages

The EMBO Journal, Journal Year: 2020, Volume and Issue: 40(1)

Published: Dec. 3, 2020

In order to replicate, human immunodeficiency virus (HIV-1) reverse-transcribes its RNA genome into DNA, which subsequently integrates host cell chromosomes. These two key events of the viral life cycle are commonly viewed as separate not only in time, but also cellular space, since reverse transcription (RT) is thought be completed cytoplasm before nuclear import and integration. However, spatiotemporal organization early replication macrophages, natural non-dividing target cells that constitute reservoirs HIV-1 an obstacle curing AIDS, remains unclear. Here, we demonstrate infected macrophages display large foci DNA (vDNA) RNA, multiple genomes cluster together. clusters form absence chromosomal integration, sequester paraspeckle protein CPSF6, localize speckles. Surprisingly, these consist mostly genomic, incoming both where pharmacologically suppressed untreated cells. We following temporary inhibition, can resume nucleus lead vDNA accumulation clusters. further show result transcription-competent DNA. findings change our understanding may have implications for addressing persistence.

Language: Английский

---

Nuclear Capsid Uncoating and Reverse Transcription of HIV-1

Annual Review of Virology, Journal Year: 2022, Volume and Issue: 9(1), P. 261 - 284

Published: June 15, 2022

After cell entry, human immunodeficiency virus type 1 (HIV-1) replication involves reverse transcription of the RNA genome, nuclear import subviral complex without envelope breakdown, and integration viral complementary DNA into host genome. Here, we discuss recent evidence indicating that completion genome uncoating occur in nucleus rather than cytoplasm, as previously thought, suggest a testable model for uncoating. Multiple studies indicated cone-shaped capsid, which encases proteins, not only serves reaction container shield from innate immune sensors but also may constitute elusive HIV-1 factor. Rupture capsid be triggered by transcription, yet-unknown factors, or physical damage, it appears to close temporal spatial association with process.

Language: Английский

---

Nuclear speckles – a driving force in gene expression

Journal of Cell Science, Journal Year: 2022, Volume and Issue: 135(13)

Published: July 1, 2022

Nuclear speckles are dynamic membraneless bodies located in the cell nucleus. They harbor RNAs and proteins, many of which splicing factors, that together display complex biophysical properties dictating nuclear speckle formation maintenance. Although these were discovered decades ago, only recently has in-depth genomic analysis begun to unravel their essential functions modulation gene activity. Major advancements mapping techniques combined with microscopy approaches have enabled insights into roles may play enhancing expression, how positioning specific landmarks can regulate expression RNA processing. Some studies drawn a link between disease. Certain maladies either involve directly or dictate localization reorganization factors. This is most striking during viral infection, as viruses alter entire architecture highjack host machinery. As discussed this Review, represent fascinating target study not reveal links positioning, genome subcompartments activity, but also potential for therapeutics.

Language: Английский

---

Deconstructing virus condensation

PLoS Pathogens, Journal Year: 2021, Volume and Issue: 17(10), P. e1009926 - e1009926

Published: Oct. 14, 2021

Viruses have evolved precise mechanisms for using the cellular physiological pathways their perpetuation. These virus-driven biochemical events must be separated in space and time from those of host cell. In recent years, granular structures, known over a century rabies virus, were shown to viral gene function named terms such as viroplasms, replication sites, inclusion bodies, or factories (VFs). More recently, these VFs liquid-like, sharing properties with membrane-less organelles driven by liquid-liquid phase separation (LLPS) process widely referred biomolecular condensation. Some best described examples structures come negative stranded RNA viruses, where micrometer size are formed toward end infectious cycle. We here discuss some basic principles LLPS connection several propose view, which integrates biochemistry underlying liquid-like organelles. this protein components gradually accumulate up critical point during infection is triggered. This yields an increase transcription that leads turn increased translation consequent growth initially condensates. According chemical behind separation, concentration increases condensate. A positive feedback cycle would thus generate crucial components, particular nucleoproteins polymerases, reach highest levels required genome replication. Progress understanding condensation exploration novel therapeutics. Furthermore, it provides insights into fundamentals regulation given virus transcription, requiring governed same principles.

Language: Английский

---

Phase separation in viral infections

Trends in Microbiology, Journal Year: 2022, Volume and Issue: 30(12), P. 1217 - 1231

Published: July 25, 2022

Language: Английский

---

Capsid–host interactions for HIV-1 ingress

Microbiology and Molecular Biology Reviews, Journal Year: 2023, Volume and Issue: 87(4)

Published: Sept. 26, 2023

The HIV-1 capsid, composed of approximately 1,200 copies the capsid protein, encases genomic RNA alongside viral nucleocapsid, reverse transcriptase, and integrase proteins. After cell entry, interacts with a myriad host factors to traverse cytoplasm, pass through nuclear pore complex (NPC), then traffic chromosomal sites for DNA integration. Integration may very well require dissolution but where when this uncoating event occurs remains hotly debated. Based on size constraints, long-prevailing view was that preceded transport, recent research has indicated remain largely intact during import, perhaps some structural remodeling required NPC traversal. Completion transcription in nucleus further aid uncoating. One canonical type factor, typified by CPSF6, leverages Phe-Gly (FG) motif bind capsid. Recent shown these peptides reside amid prion-like domains (PrLDs), which are stretches protein sequence devoid charged residues. Intermolecular PrLD interactions along exterior shell impart avid factor binding productive infection. Herein we overview capsid-host implicated ingress discuss important questions moving forward. Highlighting clinical relevance, long-acting ultrapotent inhibitor lenacapavir, engages same pocket as FG factors, recently approved treat people living HIV.

Language: Английский

---

Macrophages: Key Cellular Players in HIV Infection and Pathogenesis

Viruses, Journal Year: 2024, Volume and Issue: 16(2), P. 288 - 288

Published: Feb. 13, 2024

Although cells of the myeloid lineages, including tissue macrophages and conventional dendritic cells, were rapidly recognized, in addition to CD4+ T lymphocytes, as target HIV-1, their specific roles pathophysiology infection initially largely neglected. However, numerous studies performed over past decade, both vitro cell culture systems vivo monkey humanized mouse animal models, led growing evidence that play important direct indirect HIV-1 pathogenesis. It has been recently proposed are likely involved all stages pathogenesis, virus transmission dissemination, but above all, viral persistence through establishment, together with latently infected reservoirs many host tissues, major obstacle eradication people living HIV. Infected indeed found, very often multinucleated giant expressing antigens, almost lymphoid non-lymphoid tissues HIV-1-infected patients, where they can probably persist for long period time. In addition, also participate, directly targets or indirectly key regulators innate immunity inflammation, chronic inflammation associated clinical disorders observed HIV, even patients receiving effective antiretroviral therapy. The main objective this review is therefore summarize recent findings, revisit older data, regarding critical functions infection, found well during different

Language: Английский

---

HIV-1 uncoating requires long double-stranded reverse transcription products

Science Advances, Journal Year: 2024, Volume and Issue: 10(17)

Published: April 24, 2024

HIV-1 cores, which contain the viral genome and replication machinery, must disassemble (uncoat) during replication. However, host factors that trigger uncoating remain unidentified. Recent studies show infectious cores enter nucleus uncoat near site of integration. Here, we efficient nuclear requires synthesis a double-stranded DNA (dsDNA) >3.5 kb efficiency correlates with size. Core disruption by capsid inhibitors releases DNA, some integrates. most is degraded, indicating intact core safeguards DNA. Atomic force microscopy content estimation reveal full-length genomic dsDNA induces substantial internal strain on to promote uncoating. We conclude protect from degradation long reverse transcription products required

Language: Английский

---

HIV-1 usurps mixed-charge domain-dependent CPSF6 phase separation for higher-order capsid binding, nuclear entry and viral DNA integration

Nucleic Acids Research, Journal Year: 2024, Volume and Issue: unknown

Published: Aug. 25, 2024

Abstract HIV-1 integration favors nuclear speckle (NS)-proximal chromatin and viral infection induces the formation of capsid-dependent CPSF6 condensates that colocalize with speckles (NSs). Although displays liquid-liquid phase separation (LLPS) activity in vitro, contributions its different intrinsically disordered regions, which includes a central prion-like domain (PrLD) capsid binding FG motif C-terminal mixed-charge (MCD), to LLPS remain unclear. Herein, we determined PrLD MCD both contribute vitro. Akin mutant CPSF6, cells expressing MCD-deleted uncharacteristically arrested at rim. While heterologous MCDs effectively substituted for function during infection, Arg-Ser domains from related SR proteins were largely ineffective. wildtype displayed similar affinities, imparted LLPS-dependent higher-order co-aggregation capsids vitro cellulo. NS depletion reduced puncta without significantly affecting into NS-proximal chromatin, appending onto protein partially restored virus penetration targeting knockout cells. We conclude MCD-dependent condensation underlies post-nuclear incursion DNA pathogenesis.

Language: Английский

---