Going beyond Integration: The Emerging Role of HIV-1 Integrase in Virion Morphogenesis

Elliott, Jennifer L; Kutluay, Sebla B.

doi:10.3390/v12091005

Cited by 23 publications

(25 citation statements)

References 224 publications

(348 reference statements)

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“…Thus, the maturation status of extracellular defective viruses becomes increasingly important for estimating their potential immunogenicity and our novel FRET labeling system would be suitable for investigating this matter. In addition, the viral integrase has been also shown to be necessary for correct HIV-1 maturation ( Fontana et al, 2015 ; Kessl et al, 2016 ; Elliott and Kutluay, 2020 ) and its mechanism of action could also be explored using HIV-1 Gag-iFRET.…”

Section: Discussionmentioning

confidence: 99%

FRET-Based Detection and Quantification of HIV-1 Virion Maturation

et al. 2021

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HIV-1 infectivity is achieved through virion maturation. Virus particles undergo structural changes via cleavage of the Gag polyprotein mediated by the viral protease, causing the transition from an uninfectious to an infectious status. The majority of proviruses in people living with HIV-1 treated with combination antiretroviral therapy are defective with large internal deletions. Defective proviral DNA frequently preserves intact sequences capable of expressing viral structural proteins to form virus-like particles whose maturation status is an important factor for chronic antigen-mediated immune stimulation and inflammation. Thus, novel methods to study the maturation capability of defective virus particles are needed to characterize their immunogenicity. To build a quantitative tool to study virion maturation in vitro, we developed a novel single virion visualization technique based on fluorescence resonance energy transfer (FRET). We inserted an optimized intramolecular CFP-YPF FRET donor-acceptor pair bridged with an HIV-1 protease cleavage sequence between the Gag MA-CA domains. This system allowed us to microscopically distinguish mature and immature virions via their FRET signal when the FRET donor and acceptor proteins were separated by the viral protease during maturation. We found that approximately 80% of the FRET labeled virus particles were mature with equivalent infectivity to wild type. The proportion of immature virions was increased by treatment of virus producer cells with a protease inhibitor in a dose-dependent manner, which corresponded to a relative decrease in infectivity. Potential areas of application for this tool are assessing maturation efficiency in different cell type settings of intact or deficient proviral DNA integrated cells. We believe that this FRET-based single-virion imaging platform will facilitate estimating the impact on the immune system of both extracellular intact and defective viruses by quantifying the Gag maturation status.

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Section: Discussionmentioning

confidence: 99%

FRET-Based Detection and Quantification of HIV-1 Virion Maturation

et al. 2021

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“…The canonical role of HIV-1 IN is to catalyze insertion of the reverse transcribed viral DNA into host chromatin [ 4 ]. However, IN mutations have been identified that not only impair viral integration, but also influence other steps of the viral lifecycle, including maturation (reviewed in [ 5 , 6 ]). Mutations that specifically impair viral DNA integration are known as class I, whereas class II mutations also affect other steps of the viral lifecycle [ 5 ].…”

Section: Introductionmentioning

confidence: 99%

HIV-1 integrase binding to genomic RNA 5′-UTR induces local structural changes in vitro and in virio

Liu

Koneru

et al. 2021

Retrovirology

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Background During HIV-1 maturation, Gag and Gag-Pol polyproteins are proteolytically cleaved and the capsid protein polymerizes to form the honeycomb capsid lattice. HIV-1 integrase (IN) binds the viral genomic RNA (gRNA) and impairment of IN-gRNA binding leads to mis-localization of the nucleocapsid protein (NC)-condensed viral ribonucleoprotein complex outside the capsid core. IN and NC were previously demonstrated to bind to the gRNA in an orthogonal manner in virio; however, the effect of IN binding alone or simultaneous binding of both proteins on gRNA structure is not yet well understood. Results Using crosslinking-coupled selective 2′-hydroxyl acylation analyzed by primer extension (XL-SHAPE), we characterized the interaction of IN and NC with the HIV-1 gRNA 5′-untranslated region (5′-UTR). NC preferentially bound to the packaging signal (Psi) and a UG-rich region in U5, irrespective of the presence of IN. IN alone also bound to Psi but pre-incubation with NC largely abolished this interaction. In contrast, IN specifically bound to and affected the nucleotide (nt) dynamics of the apical loop of the transactivation response element (TAR) and the polyA hairpin even in the presence of NC. SHAPE probing of the 5′-UTR RNA in virions produced from allosteric IN inhibitor (ALLINI)-treated cells revealed that while the global secondary structure of the 5′-UTR remained unaltered, the inhibitor treatment induced local reactivity differences, including changes in the apical loop of TAR that are consistent with the in vitro results. Conclusions Overall, the binding interactions of NC and IN with the 5′-UTR are largely orthogonal in vitro. This study, together with previous probing experiments, suggests that IN and NC binding in vitro and in virio lead to only local structural changes in the regions of the 5′-UTR probed here. Accordingly, disruption of IN-gRNA binding by ALLINI treatment results in local rather than global secondary structure changes of the 5′-UTR in eccentric virus particles. Graphical Abstract

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“…As mentioned before, recent works revealed that in HIV-1, IN is also an RBP with an essential role in virion morphogenesis, related to its ability to bind gRNA 8 25 26 . In fact, IN interacts with specific sites within gRNA ensuring the correct localization of viral RNP inside the capsid (reviewed in 27 ). Aberrant virions are obtained when IN-gRNA interaction is abolished, highlighting the importance of the proper formation of IN-containing RNPs for HIV-1 infection ( 8,25,28 ).…”

Section: Introductionmentioning

confidence: 99%

The HIV-1 Integrase C-Terminal domain induces TAR RNA structural changes promoting Tat binding

Rocchi

Louvat

Miele

et al. 2021

Preprint

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Recent evidence indicated that HIV-1 Integrase (IN) binds genomic viral RNA (gRNA) playing a critical role in viral particle morphogenesis and gRNA stability in host cells. Combining biophysical and biochemical approaches we show that the C-terminal flexible 18-residues tail of IN acts as a sensor of the peculiar apical structure of trans-activation response element RNA (TAR), directly interacting with its hexaloop. We highlighted how the whole IN C-terminal domain, once bound to TAR, can change its structure assisting the binding of Tat, the HIV trans-activator protein, which finally displaces IN from TAR. Our results are consistent with the emerging role of IN in early stage of proviral transcription and suggest new steps of HIV-1 life cycle that can be considered as therapeutic targets.

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Going beyond Integration: The Emerging Role of HIV-1 Integrase in Virion Morphogenesis

Cited by 23 publications

References 224 publications

FRET-Based Detection and Quantification of HIV-1 Virion Maturation

FRET-Based Detection and Quantification of HIV-1 Virion Maturation

HIV-1 integrase binding to genomic RNA 5′-UTR induces local structural changes in vitro and in virio

The HIV-1 Integrase C-Terminal domain induces TAR RNA structural changes promoting Tat binding

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