The Life-Cycle of the HIV-1 Gag–RNA Complex

Mailler, Élodie; Bernacchi, Serena; Marquet, Roland; Paillart, Jean-Christophe; Vivet-Boudou, Valérie; Smyth, Redmond P.

doi:10.3390/v8090248

Cited by 79 publications

(90 citation statements)

References 190 publications

(301 reference statements)

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“…Surprisingly, the influence of the 5′-Cap moiety on dimerization was similar to that of adding a single phosphodiester-linked 5′ guanosine. Our findings support proposals that genome dimerization is a major determinant of RNA function (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(25)(26)(27)(28)(29)(30)(31)(32)(33) and suggest a paradigm in which the structure, function, and fate of the viral transcripts are modulated by heterogeneous TSS selection.…”

Section: Significancesupporting

confidence: 87%

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Transcriptional start site heterogeneity modulates the structure and function of the HIV-1 genome

Kharytonchyk

Monti

Smaldino

et al. 2016

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

119

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The promoter in HIV type 1 (HIV-1) proviral DNA contains three sequential guanosines at the U3-R boundary that have been proposed to function as sites for transcription initiation. Here we show that all three sites are used in cells infected with HIV-1 and that viral RNAs containing a single 5′ capped guanosine ( Cap 1G) are specifically selected for packaging in virions, consistent with a recent report [Masuda et al. (2015) Sci Rep 5:17680]. In addition, we now show that transcripts that begin with two or three capped guanosines ( Cap 2G or Cap 3G) are enriched on polysomes, indicating that RNAs synthesized from different transcription start sites have different functions in viral replication. Because genomes are selected for packaging as dimers, we examined the in vitro monomer-dimer equilibrium properties of Cap 1G, Cap 2G, and Cap 3G 5′-leader RNAs in the NL4-3 strain of HIV-1. Strikingly, under physiological-like ionic conditions in which the Cap 1G 5′-leader RNA adopts a dimeric structure, the Cap 2G and Cap 3G 5′-leader RNAs exist predominantly as monomers. Mutagenesis studies designed to probe for base-pairing interactions suggest that the additional guanosines of the 2G and 3G RNAs remodel the base of the PolyA hairpin, resulting in enhanced sequestration of dimerpromoting residues and stabilization of the monomer. Our studies suggest a mechanism through which the structure, function, and fate of the viral genome can be modulated by the transcriptionally controlled presence or absence of a single 5′ guanosine.HIV-1 | 5′-leader | transcription | RNA | structure

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Section: Significancesupporting

confidence: 87%

“…Because prior studies have established that genome dimerization likely controls packaging and other RNA-dependent functions (8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(25)(26)(27)(28)(29)(30)(31)(32)(33), we examined the influence of TSS use on the in vitro dimerization properties of the HIV-1 NL4-3 5′-L (Fig. 4A).…”

Section: Resultsmentioning

confidence: 99%

Transcriptional start site heterogeneity modulates the structure and function of the HIV-1 genome

Kharytonchyk

Monti

Smaldino

et al. 2016

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

119

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“…Several riboswitch models have been proposed for HIV-1 [50–53], and in the case of FIV, the existence of dual structures has been validated by SHAPE and RNA dimerization assays [54]. Flexibility in retroviral gRNA is created via long-range interactions that have now been observed in several retroviruses, from HIV-1, FIV, MLV, to MPMV and MMTV [6,25,26,33,55–59]. The DIS in one conformation is occluded by base pairing and not available for dimerization, thus earmarking this RNA for translation, while the DIS in the other conformation is available as an apical loop on a stem loop, thus allowing dimerization and packaging of the dimeric RNA [54].…”

Section: Discussionmentioning

confidence: 99%

The bifurcated stem loop 4 (SL4) is crucial for efficient packaging of mouse mammary tumor virus (MMTV) genomic RNA

et al. 2018

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Packaging the mouse mammary tumor virus (MMTV) genomic RNA (gRNA) requires the entire 5ʹ untranslated region (UTR) in conjunction with the first 120 nucleotides of the gag gene. This region includes several palindromic (pal) sequence(s) and stable stem loops (SLs). Among these, stem loop 4 (SL4) adopts a bifurcated structure consisting of three stems, two apical loops, and an internal loop. Pal II, located in one of the apical loops, mediates gRNA dimerization, a process intricately linked to packaging. We thus hypothesized that the bifurcated SL4 structure could constitute the major gRNA packaging determinant. To test this hypothesis, the two apical loops and the flanking sequences forming the bifurcated SL4 were individually mutated. These mutations all had deleterious effects on gRNA packaging and propagation. Next, single and compensatory mutants were designed to destabilize then recreate the bifurcated SL4 structure. A structure-function analysis using bioinformatics predictions and RNA chemical probing revealed that mutations that led to the loss of the SL4 bifurcated structure abrogated RNA packaging and propagation, while compensatory mutations that recreated the native SL4 structure restored RNA packaging and propagation to wild type levels. Altogether, our results demonstrate that SL4 constitutes the principal packaging determinant of MMTV gRNA. Our findings further suggest that SL4 acts as a structural switch that can not only differentiate between RNA for translation versus packaging/dimerization, but its location also allows differentiation between spliced and unspliced RNAs during gRNA encapsidation.

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“…Genetic studies indicate that the DIS is responsible for RNA:RNA partner selection [15,16], but other leader elements, including those overlapping the gag start codon (AUG, Figure 1), also play roles in dimerization [13,17,18,19]. Genome selection during virus assembly is mediated by interactions between the nucleocapsid (NC) domains of a small number of viral Gag polyproteins (~12 or fewer) [20] and packaging signals located within the 5′-leader of the viral RNA [8,10,11,14,21,22,23,24,25,26,27,28,29]. Additional Gag proteins associate with the genome after it is anchored to the PM, leading to further virus assembly and budding [30].…”

Section: Dimerization-dependent Control Of Human Immunodeficiency mentioning

confidence: 99%

“…In vitro, human immunodeficiency virus type 1 (HIV-1) 5′-leader RNA (residues 1–356, which includes the intact 5′-untranslated region and the first 21 nucleotides of the gag gene; Figure 1) exists as an equilibrium mixture of monomeric and dimeric species [8,10,11,14,21,22,23,24,25,26,27,28,29]. The secondary structure of the 5′-leader has been probed by combinations of nucleotide reactivity mapping, phylogenetic analyses, biochemical and molecular biological studies, and free energy calculations, resulting in more than 25 predicted secondary structures and multiple dimerization models [8].…”

Section: Probing the Secondary Structure Of The Intact 5′-leader Bmentioning

confidence: 99%

NMR Studies of the Structure and Function of the HIV-1 5′-Leader

Keane

Summers

2016

Viruses

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The 5′-leader of the human immunodeficiency virus type 1 (HIV-1) genome plays several critical roles during viral replication, including differentially establishing mRNA versus genomic RNA (gRNA) fates. As observed for proteins, the function of the RNA is tightly regulated by its structure, and a common paradigm has been that genome function is temporally modulated by structural changes in the 5′-leader. Over the past 30 years, combinations of nucleotide reactivity mapping experiments with biochemistry, mutagenesis, and phylogenetic studies have provided clues regarding the secondary structures of stretches of residues within the leader that adopt functionally discrete domains. More recently, nuclear magnetic resonance (NMR) spectroscopy approaches have been developed that enable direct detection of intra- and inter-molecular interactions within the intact leader, providing detailed insights into the structural determinants and mechanisms that regulate HIV-1 genome packaging and function.

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The Life-Cycle of the HIV-1 Gag–RNA Complex

Cited by 79 publications

References 190 publications

Transcriptional start site heterogeneity modulates the structure and function of the HIV-1 genome

Transcriptional start site heterogeneity modulates the structure and function of the HIV-1 genome

The bifurcated stem loop 4 (SL4) is crucial for efficient packaging of mouse mammary tumor virus (MMTV) genomic RNA

NMR Studies of the Structure and Function of the HIV-1 5′-Leader

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