scholarly journals An epitranscriptomic switch at the 5′-UTR controls genome selection during HIV-1 genomic RNA packaging

2019 ◽  
Author(s):  
Camila Pereira-Montecinos ◽  
Daniela Toro-Ascuy ◽  
Cecilia Rojas-Fuentes ◽  
Sebastián Riquelme-Barrios ◽  
Bárbara Rojas-Araya ◽  
...  
Keyword(s):  
Full Length ◽  
Genomic Rna ◽  
Rna Packaging ◽  
Gag Protein ◽  
Rna Molecules ◽  
Viral Genomes ◽  
Viral Particles ◽  
Demethylase Activity ◽  
Retroviral Replication ◽  
Hiv 1

ABSTRACTDuring retroviral replication, the full-length RNA serves both as mRNA and genomic RNA (gRNA). While the simple retrovirus MLV segregates its full-length RNA into two functional populations, the HIV-1 full-length RNA was proposed to exist as a single population used indistinctly for protein synthesis or packaging. However, the mechanisms by which the HIV-1 Gag protein selects the two RNA molecules that will be packaged into nascent virions remain poorly understood. Here, we demonstrate that HIV-1 full-length RNA packaging is regulated through an epitranscriptomic switch requiring demethylation of two conserved adenosine residues present within the 5′-UTR. As such, while m6A deposition by METTL3/METTL14 onto the full-length RNA was associated with increased Gag synthesis and reduced packaging, FTO-mediated demethylation was required for the incorporation of the full-length RNA into viral particles. Interestingly, HIV-1 Gag associates with the RNA demethylase FTO in the nucleus and drives full-length RNA demethylation. Finally, the specific inhibition of the FTO RNA demethylase activity suppressed HIV-1 full-length RNA packaging. Together, our data propose a novel epitranscriptomic mechanism allowing the selection of the full-length RNA molecules that will be used as viral genomes.

scholarly journals Human Immunodeficiency Virus Types 1 and 2 Differ in the Predominant Mechanism Used for Selection of Genomic RNA for Encapsidation

1999 ◽  
Vol 73 (4) ◽  
pp. 3023-3031 ◽  
Author(s):  
Jane F. Kaye ◽  
Andrew M. L. Lever
Keyword(s):  
Human Immunodeficiency Virus ◽  
Genomic Rna ◽  
Rna Packaging ◽  
Packaging Signal ◽  
Initiation Mechanism ◽  
Gag Protein ◽  
Gag Proteins ◽  
Viral Genomes ◽  
Immunodeficiency Virus

ABSTRACT Retroviral RNA encapsidation is a highly selective process mediated through recognition by the viral Gag proteins of cis-acting RNA packaging signals in genomic RNA. This RNA species is also translated, producing the viral gag gene products. The relationship between these processes is poorly understood. Unlike that of human immunodeficiency virus type 1 (HIV-1), the dominant packaging signal of HIV-2 is upstream of the major splice donor and present in both unspliced and spliced viral RNAs, necessitating additional mechanisms for preferential packaging of unspliced genomic RNA. Encapsidation studies of a series of HIV-2-based vectors showed efficient packaging of viral genomes only if the unspliced, encapsidated RNA expressed full-length Gag protein, including functional nucleocapsid. We propose a novel encapsidation initiation mechanism, providing selectivity, in which unspliced HIV-2 RNA is captured in cis by the Gag protein. This has implications for the use of HIV-2 and other lentiviruses as vectors.

scholarly journals A Heterologous, High-Affinity RNA Ligand for Human Immunodeficiency Virus Gag Protein Has RNA Packaging Activity

2000 ◽  
Vol 74 (1) ◽  
pp. 541-546 ◽  
Author(s):  
Jared L. Clever ◽  
Randy A. Taplitz ◽  
Michael A. Lochrie ◽  
Barry Polisky ◽  
Tristram G. Parslow
Keyword(s):  
Human Immunodeficiency Virus ◽  
Specific Binding ◽  
Single Point Mutation ◽  
Genomic Rna ◽  
Rna Packaging ◽  
Gag Protein ◽  
High Affinity ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Retroviral RNA encapsidation depends on the specific binding of Gag proteins to packaging (ψ) signals in genomic RNA. We investigated whether an in vitro-selected, high-affinity RNA ligand for the nucleocapsid (NC) portion of the Gag protein from human immunodeficiency virus type 1 (HIV-1) could mediate packaging into HIV-1 virions. We find that this ligand can functionally substitute for one of the Gag-binding elements (termed SL3) in the HIV-1 ψ locus to support packaging and viral infectivity in cis. By contrast, this ligand, which fails to dimerize spontaneously in vitro, is unable to replace a different ψ element (termed SL1) which is required for both Gag binding and dimerization of the HIV-1 genome. A single point mutation within the ligand that eliminates high-affinity in vitro Gag binding also abolishes its packaging activity at the SL3 position. These results demonstrate that specific binding of Gag or NC protein is a critical determinant of genomic RNA packaging.

scholarly journals RNA and Protein Requirements for Incorporation of the Pol Protein into Foamy Virus Particles

2005 ◽  
Vol 79 (11) ◽  
pp. 7005-7013 ◽  
Author(s):  
Katrin Peters ◽  
Tatiana Wiktorowicz ◽  
Martin Heinkelein ◽  
Axel Rethwilm
Keyword(s):  
Foamy Virus ◽  
Genomic Rna ◽  
Rna Packaging ◽  
Gag Protein ◽  
Protein Precursor ◽  
Protein Requirements ◽  
Virus Particles ◽  
Foamy Viruses ◽  
Protein Incorporation ◽  
The Individual

ABSTRACT Foamy viruses (FVs) generate their Pol protein precursor molecule independently of the Gag protein from a spliced mRNA. This mode of expression raises the question of the mechanism of Pol protein incorporation into the viral particle (capsid). We previously showed that the packaging of (pre)genomic RNA is essential for Pol encapsidation (M. Heinkelein, C. Leurs, M. Rammling, K. Peters, H. Hanenberg, and A. Rethwilm, J. Virol. 76:10069-10073, 2002). Here, we demonstrate that distinct sequences in the RNA, which we termed Pol encapsidation sequences (PES), are required to incorporate Pol protein into the FV capsid. Two PES were found, which are contained in the previously identified cis-acting sequences necessary to transfer an FV vector. One PES is located in the U5 region of the 5′ long terminal repeat and one at the 3′ end of the pol gene region. Neither element has any significant effect on RNA packaging. However, deletion of either PES resulted in a significant reduction in Pol encapsidation. On the protein level, we show that only the Pol precursor, but not the individual reverse transcriptase (RT) and integrase (IN) subunits, is incorporated into FV particles. However, enzymatic activities of the protease (PR), RT, or IN are not required. Our results strengthen the view that in FVs, (pre)genomic RNA functions as a bridging molecule between Gag and Pol precursor proteins.

scholarly journals Life of psi: How full-length HIV-1 RNAs become packaged genomes in the viral particles

Virology ◽  
2014 ◽  
Vol 454-455 ◽  
pp. 362-370 ◽  
Author(s):  
Malika Kuzembayeva ◽  
Kari Dilley ◽  
Luca Sardo ◽  
Wei-Shau Hu
Keyword(s):  
Full Length ◽  
Viral Particles ◽  
Hiv 1

scholarly journals Selective and Nonselective Packaging of Cellular RNAs in Retrovirus Particles

2007 ◽  
Vol 81 (12) ◽  
pp. 6623-6631 ◽  
Author(s):  
Samuel J. Rulli ◽  
Catherine S. Hibbert ◽  
Jane Mirro ◽  
Thoru Pederson ◽  
Shyam Biswal ◽  
...  
Keyword(s):  
Leukemia Virus ◽  
Signal Recognition Particle ◽  
Viral Rna ◽  
Genomic Rna ◽  
Gag Protein ◽  
Mrna Species ◽  
Reverse Transcription Pcr ◽  
Cellular Mrnas ◽  
A Minor ◽  
Hiv 1

ABSTRACT Assembly of retrovirus particles normally entails the selective encapsidation of viral genomic RNA. However, in the absence of packageable viral RNA, assembly is still efficient, and the released virus-like particles (termed “Ψ−” particles) still contain roughly normal amounts of RNA. We have proposed that cellular mRNAs replace the genome in Ψ− particles. We have now analyzed the mRNA content of Ψ− and Ψ+ murine leukemia virus (MLV) particles using both microarray analysis and real-time reverse transcription-PCR. The majority of mRNA species present in the virus-producing cells were also detected in Ψ− particles. Remarkably, nearly all of them were packaged nonselectively; that is, their representation in the particles was simply proportional to their representation in the cells. However, a small number of low-abundance mRNAs were greatly enriched in the particles. In fact, one mRNA species was enriched to the same degree as Ψ+ genomic RNA. Similar results were obtained with particles formed from the human immunodeficiency virus type 1 (HIV-1) Gag protein, and the same mRNAs were enriched in MLV and HIV-1 particles. The levels of individual cellular mRNAs were ∼5- to 10-fold higher in Ψ− than in Ψ+ MLV particles, in agreement with the idea that they are replacing viral RNA in the former. In contrast, signal recognition particle RNA was present at the same level in Ψ− and Ψ+ particles; a minor fraction of this RNA was weakly associated with genomic RNA in Ψ+ MLV particles.

scholarly journals Visualizing the translation and packaging of HIV-1 full-length RNA

2020 ◽  
Vol 117 (11) ◽  
pp. 6145-6155 ◽  
Author(s):  
Jianbo Chen ◽  
Yang Liu ◽  
Bin Wu ◽  
Olga A. Nikolaitchik ◽  
Preeti R. Mohan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Molecule ◽  
Rna Binding ◽  
Virus Assembly ◽  
Full Length ◽  
Rna Molecules ◽  
Rna Translation ◽  
Translation Template ◽  
A Genome ◽  
Hiv 1

HIV-1 full-length RNA (HIV-1 RNA) plays a central role in viral replication, serving as a template for Gag/Gag-Pol translation and as a genome for the progeny virion. To gain a better understanding of the regulatory mechanisms of HIV-1 replication, we adapted a recently described system to visualize and track translation from individual HIV-1 RNA molecules in living cells. We found that, on average, half of the cytoplasmic HIV-1 RNAs are being actively translated at a given time. Furthermore, translating and nontranslating RNAs are well mixed in the cytoplasm; thus, Gag biogenesis occurs throughout the cytoplasm without being constrained to particular subcellular locations. Gag is an RNA binding protein that selects and packages HIV-1 RNA during virus assembly. A long-standing question in HIV-1 gene expression is whether Gag modulates HIV-1 RNA translation. We observed that despite its RNA-binding ability, Gag expression does not alter the proportion of translating HIV-1 RNA. Using single-molecule tracking, we found that both translating and nontranslating RNAs exhibit dynamic cytoplasmic movement and can reach the plasma membrane, the major HIV-1 assembly site. However, Gag selectively packages nontranslating RNA into the assembly complex. These studies illustrate that although HIV-1 RNA serves two functions, as a translation template and as a viral genome, individual RNA molecules carry out only one function at a time. These studies shed light on previously unknown aspects of HIV-1 gene expression and regulation.

scholarly journals New Structure Sheds Light on Selective HIV-1 Genomic RNA Packaging

Viruses ◽  
2015 ◽  
Vol 7 (8) ◽  
pp. 4826-4835 ◽  
Author(s):  
Erik Olson ◽  
William Cantara ◽  
Karin Musier-Forsyth
Keyword(s):  
Genomic Rna ◽  
Rna Packaging ◽  
Hiv 1

scholarly journals Distinct binding interactions of HIV-1 Gag to Psi and non-Psi RNAs: Implications for viral genomic RNA packaging

RNA ◽  
2013 ◽  
Vol 19 (8) ◽  
pp. 1078-1088 ◽  
Author(s):  
J. A. Webb ◽  
C. P. Jones ◽  
L. J. Parent ◽  
I. Rouzina ◽  
K. Musier-Forsyth
Keyword(s):  
Genomic Rna ◽  
Rna Packaging ◽  
Binding Interactions ◽  
Viral Genomic Rna ◽  
Hiv 1

scholarly journals Coordination of Genomic RNA Packaging with Viral Assembly in HIV-1

Viruses ◽  
2016 ◽  
Vol 8 (7) ◽  
pp. 192 ◽  
Author(s):  
Chris Hellmund ◽  
Andrew Lever
Keyword(s):  
Viral Assembly ◽  
Genomic Rna ◽  
Rna Packaging ◽  
Hiv 1
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