An Internal Ribosome Entry Segment Promotes Translation of the Simian Immunodeficiency Virus Genomic RNA

The full-length genomic RNA of lentiviruses can be translated to produce proteins and incorporated as genomic RNA in the viral particle. Interestingly, both functions are driven by the genomic 5′-UTR (5′-untranslated region), which harbours structural RNA motifs for the replication cycle of the virus. Recent work has shown that this RNA architecture also functions as an IRES (internal ribosome entry site) in HIV-1 and -2, and in SIV (simian immunodeficiency virus). In addition, the IRES extends to the gag coding region for all these viruses and this leads to the synthesis of shorter isoforms of the Gag polyprotein from downstream initiation codons. In the present study, we have investigated how different members of the lentivirus family (namely HIV-1 and -2, and SIV) can initiate protein synthesis by distinct mechanisms. For this, we have used the competitive reticulocyte lysate that we have recently described. Our results show that HIV-1 is able to drive the synthesis of the Gag polyprotein both by a classical cap-dependent mechanism and an IRES, whereas HIV-2 and SIV appear to use exclusively an IRES mechanism.

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Optimization of PCR for quantification of simian immunodeficiency virus genomic RNA in plasma of rhesus macaques (Macaca mulatta ) using armored RNA

Journal of Medical Primatology ◽

10.1111/jmp.12088 ◽

2013 ◽

Vol 43 (1) ◽

pp. 31-43 ◽

Cited By ~ 17

Author(s):

C.J. Monjure ◽

C.D. Tatum ◽

A.T. Panganiban ◽

M. Arainga ◽

V. Traina-Dorge ◽

...

Keyword(s):

Simian Immunodeficiency Virus ◽

Macaca Mulatta ◽

Rhesus Macaques ◽

Genomic Rna ◽

Immunodeficiency Virus ◽

Armored Rna

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The Simian Immunodeficiency Virus 5′ Untranslated Leader Sequence Plays a Role in Intracellular Viral Protein Accumulation and in RNA Packaging

Journal of Virology ◽

10.1128/jvi.77.11.6284-6292.2003 ◽

2003 ◽

Vol 77 (11) ◽

pp. 6284-6292 ◽

Cited By ~ 11

Author(s):

Jignesh Patel ◽

Shainn-Wei Wang ◽

Elena Izmailova ◽

Anna Aldovini

Keyword(s):

Simian Immunodeficiency Virus ◽

Initiation Codon ◽

Primer Binding Site ◽

Deletion Mutants ◽

Protein Accumulation ◽

Genomic Rna ◽

Rna Packaging ◽

Packaging Signal ◽

Immunodeficiency Virus ◽

Selective Incorporation

ABSTRACT We investigated the role of 5′ untranslated leader sequences of simian immunodeficiency virus (SIVmac239) in RNA encapsidation and protein expression. A series of progressively longer deletion mutants was constructed with a common endpoint six nucleotides upstream of the gag initiation codon and another endpoint at the 3′ end of the primer binding site (PBS). We found that efficient intracellular Gag-Pol protein accumulation required the region between the PBS and splice donor (SD) site. Marked reduction of genomic RNA packaging was observed with all the deletion mutants that involved sequences at both the 5′ and at the 3′ ends of the major SD site, and increased nonspecific RNA incorporation could be detected in these mutants. RNA encapsidation was affected only modestly by a deletion of 54 nucleotides at the 3′ end of the SD site when the mutant construct pΔ54 was transfected alone. In contrast, the amount of pΔ54 genomic RNA incorporated into particles was reduced more than 10-fold when this mutant was cotransfected with a construct specifying an RNA molecule with a wild-type packaging signal. Therefore, we conclude that the 175 nucleotides located 5′ of the gag initiation codon are critical for efficient and selective incorporation of genomic RNA into virions. This location of the SIV Ψ element provides the means for efficient discrimination between viral genomic and spliced RNAs.

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A Stretch of Unpaired Purines in the Leader Region of Simian Immunodeficiency Virus (SIV) Genomic RNA is Critical for its Packaging into Virions

Journal of Molecular Biology ◽

10.1016/j.jmb.2021.167293 ◽

2021 ◽

pp. 167293

Author(s):

Vineeta N. Pillai ◽

Lizna Mohamed Ali ◽

Suresha G. Prabhu ◽

Anjana Krishnan ◽

Akhil Chameettachal ◽

...

Keyword(s):

Simian Immunodeficiency Virus ◽

Leader Region ◽

Genomic Rna ◽

Immunodeficiency Virus

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Rous Sarcoma Virus Translation Revisited: Characterization of an Internal Ribosome Entry Segment in the 5′ Leader of the Genomic RNA

Journal of Virology ◽

10.1128/jvi.74.24.11581-11588.2000 ◽

2000 ◽

Vol 74 (24) ◽

pp. 11581-11588 ◽

Cited By ~ 21

Author(s):

Clarence Deffaud ◽

Jean-Luc Darlix

Keyword(s):

Rous Sarcoma Virus ◽

Leukemia Virus ◽

Secondary Structures ◽

Open Reading Frames ◽

Start Codon ◽

Genomic Rna ◽

Internal Ribosome Entry ◽

Rous Sarcoma ◽

Internal Ribosome Entry Segment ◽

Sarcoma Virus

ABSTRACT The 5′ leader of Rous sarcoma virus (RSV) genomic RNA and of retroviruses in general is long and contains stable secondary structures that are critical in the early and late steps of virus replication such as RNA dimerization and packaging and in the process of reverse transcription. The initiation of RSV Gag translation has been reported to be 5′ cap dependent and controlled by three short open reading frames located in the 380-nucleotide leader upstream of the Gag start codon. Translation of RSV Gag would thus differ from that prevailing in other retroviruses such as murine leukemia virus, reticuloendotheliosis virus type A, and simian immunodeficiency virus, in which an internal ribosome entry segment (IRES) in the 5′ end of the genomic RNA directs efficient Gag expression despite stable 5′ secondary structures. This prompted us to investigate whether RSV Gag translation might be controlled by an IRES-dependent mechanism. The results show that the 5′ leaders of RSV and v-Src RNA exhibit IRES properties, since these viral elements can promote efficient translation of monocistronic RNAs in conditions inhibiting 5′ cap-dependent translation. When inserted between two cistrons in a canonical bicistronic construct, both the RSV and v-Src leaders promote expression of the 3′ cistron. A genetic analysis of the RSV leader allowed the identification of two nonoverlapping 5′ and 3′ leader domains with IRES activity. In addition, the v-Src leader was found to contain unique 3′ sequences promoting an efficient reinitiation of translation. Taken together, these data lead us to propose a new model for RSV translation.

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Alteration of Zinc-Binding Residues of Simian Immunodeficiency Virus p8NC Results in Subtle Differences in Gag Processing and Virion Maturation Associated with Degradative Loss of Mutant NC

Journal of Virology ◽

10.1128/jvi.75.1.115-124.2001 ◽

2001 ◽

Vol 75 (1) ◽

pp. 115-124 ◽

Cited By ~ 14

Author(s):

Jason L. Yovandich ◽

Elena N. Chertova ◽

Brad P. Kane ◽

Tracy D. Gagliardi ◽

Julian W. Bess ◽

...

Keyword(s):

Simian Immunodeficiency Virus ◽

High Performance ◽

Point Mutations ◽

Particle Morphology ◽

Reversed Phase ◽

Significant Loss ◽

Full Length ◽

Genomic Rna ◽

Rna Packaging ◽

Immunodeficiency Virus

ABSTRACT In all retroviruses analyzed to date (except for the spumaretroviruses), the Zn2+-coordinating residues of nucleocapsid (NC) perform or assist in crucial reactions necessary to complete the retrovirus life cycle. Six replication-defective mutations have been engineered in the two NC Zn2+ fingers (ZFs) of simian immunodeficiency virus [SIV(Mne)] that change or delete specific Zn2+-interacting Cys residues and were studied by using electron microscopy, reversed-phase high-performance liquid chromatography, immunoblotting, and RNA quantification. We focused on phenotypes of produced particles, specifically morphology, Gag polyprotein processing, and genomic RNA packaging. Phenotypes were similar among viruses containing a point or deletion mutation involving the same ZF. Mutations in the proximal ZF (ZF1) resulted in near-normal Gag processing and full-length genomic RNA incorporation and were most similar to wild-type (WT) virions with electron-dense, conical cores. Mutation of the distal ZF, as well as point mutations in both ZFs, resulted in more unprocessed Gag proteins than a deletion or point mutation in ZF1, with an approximate 30% reduction in levels of full-length genomic RNA in virions. These mutant virions contained condensed cores; however, the cores typically appeared less electron dense and more rod shaped than WT virions. Surprisingly, deletion of both ZFs, including the basic linker region between the ZFs, resulted in the most efficient Gag processing. However, genomic RNA packaging was ∼10% of WT levels, and those particles produced were highly abnormal with respect to size and core morphology. Surprisingly, all NC mutations analyzed demonstrated a significant loss of processed NC in virus particles, suggesting that Zn2+-coordinated NC is protected from excessive proteolytic cleavage. Together, these results indicate that Zn2+ coordination is important for correct Gag precursor processing and NC protein stability. Additionally, SIV particle morphology appears to be the result of proper and complete Gag processing and relies less on full-length genomic RNA incorporation, as dictated by the Zn2+ coordination in the ZFs of the NC protein.

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The Leader of Human Immunodeficiency Virus Type 1 Genomic RNA Harbors an Internal Ribosome Entry Segment That Is Active during the G2/M Phase of the Cell Cycle

Journal of Virology ◽

10.1128/jvi.77.7.3939-3949.2003 ◽

2003 ◽

Vol 77 (7) ◽

pp. 3939-3949 ◽

Cited By ~ 126

Author(s):

Ann Brasey ◽

Marcelo Lopez-Lastra ◽

Theophile Ohlmann ◽

Nancy Beerens ◽

Ben Berkhout ◽

...

Keyword(s):

Cell Cycle ◽

Human Immunodeficiency Virus ◽

Protein Synthesis ◽

Human Immunodeficiency Virus Type ◽

Genomic Rna ◽

M Cell ◽

Internal Ribosome Entry ◽

Immunodeficiency Virus ◽

Hiv 1

ABSTRACT The 5′ leader of the human immunodeficiency virus type 1 (HIV-1) genomic RNA contains highly structured domains involved in key steps of the viral life cycle. These RNA domains inhibit cap-dependent protein synthesis. Here we report that the HIV-1 5′ leader harbors an internal ribosome entry site (IRES) capable of driving protein synthesis during the G2/M cell cycle phase in which cap-dependent initiation is inhibited. The HIV-1 IRES was delineated with bicistronic mRNAs in in vitro and ex vivo assays. The HIV-1 leader IRES spans nucleotides 104 to 336 and partially overlaps the major determinants of genomic RNA packaging. These data strongly suggest that, as for HIV-1 transcription, IRES-mediated translation initiation could play an important role in virus replication during virus-induced G2/M cell cycle arrest.

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