Structural Characterization of the Rous Sarcoma Virus RNA Stability Element (original) (raw)
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Retrovirology, 2010
Background: Nonsense-mediated mRNA decay (NMD) is an mRNA quality control mechanism that selectively recognizes and targets for degradation mRNAs containing premature termination codons. Retroviral full-length RNA is presented to the host translation machinery with characteristics rarely observed among host cell mRNAs: a long 3′ UTR, retained introns, and multiple open reading frames. As a result, the viral RNA is predicted to be recognized by the host NMD machinery and degraded. In the case of the Rous sarcoma virus (RSV), we identified a stability element (RSE), which resides immediately downstream of the gag termination codon and facilitates NMD evasion. Results: We defined key RNA features of the RSE through directed mutagenesis of the virus. These data suggest that the minimal RSE is 155 nucleotides (nts) and functions independently of the nucleotide sequence of the stop codon or the first nucleotide following the stop codon. Further data suggested that the 3′UTRs of the RSV pol and src may also function as stability elements. Conclusions: We propose that these stability elements in RSV may be acting as NMD insulators to mask the preceding stop codon from the NMD machinery.
The structure and function of the rous sarcoma virus RNA stability element
Journal of Cellular Biochemistry, 2011
For simple retroviruses, such as the Rous sarcoma virus (RSV), post-transcriptional control elements regulate viral RNA splicing, export, stability, and packaging into virions. These RNA sequences interact with cellular host proteins to regulate and facilitate productive viral infections. One such element, known as the RSV stability element (RSE), is required for maintaining stability of the full-length unspliced RNA. This viral RNA serves as the mRNA for the Gag and Pol proteins and also as the genome packaged in progeny virions. When the RSE is deleted from the viral RNA, the unspliced RNA becomes unstable and is degraded in a Upf1-dependent manner. Current evidence suggests that the RSE inhibits recognition of the viral gag termination codon by the nonsense-mediated mRNA decay (NMD) pathway. We believe that the RSE acts as an insulator to NMD, thereby preventing at least one of the required functional steps that target an mRNA for degradation. Here, we discuss the history of the RSE and the current model of how the RSE is interacting with cellular NMD factors.
A 3' UTR sequence stabilizes termination codons in the unspliced RNA of Rous sarcoma virus
RNA, 2006
Eukaryotic cells target mRNAs to the nonsense-mediated mRNA decay (NMD) pathway when translation terminates within the coding region. In mammalian cells, this is presumably due to a downstream signal deposited during pre-mRNA splicing. In contrast, unspliced retroviral RNA undergoes NMD in chicken cells when premature termination codons (PTCs) are present in the gag gene. Surprisingly, deletion of a 401-nt 3¢ UTR sequence immediately downstream of the normal gag termination codon caused this termination event to be recognized as premature. We termed this 3¢ UTR region the Rous sarcoma virus (RSV) stability element (RSE). The RSE also stabilized the viral RNA when placed immediately downstream of a PTC in the gag gene. Deletion analysis of the RSE indicated a smaller functional element. We conclude that this 3¢ UTR sequence stabilizes termination codons in the RSV RNA, and termination codons not associated with such an RSE sequence undergo NMD.
Rous Sarcoma Virus RNA Stability Element Inhibits Deadenylation of mRNAs with Long 3′UTRs
Viruses
All retroviruses use their full-length primary transcript as the major mRNA for Group-specific antigen (Gag) capsid proteins. This results in a long 3′ untranslated region (UTR) downstream of the termination codon. In the case of Rous sarcoma virus (RSV), there is a 7 kb 3′UTR downstream of the gag terminator, containing the pol, env, and src genes. mRNAs containing long 3′UTRs, like those with premature termination codons, are frequently recognized by the cellular nonsense-mediated mRNA decay (NMD) machinery and targeted for degradation. To prevent this, RSV has evolved an RNA stability element (RSE) in the RNA immediately downstream of the gag termination codon. This 400-nt RNA sequence stabilizes premature termination codons (PTCs) in gag. It also stabilizes globin mRNAs with long 3′UTRs, when placed downstream of the termination codon. It is not clear how the RSE stabilizes the mRNA and prevents decay. We show here that the presence of RSE inhibits deadenylation severely. In add...
Retroviral strategy to stabilize viral RNA
Current Opinion in Microbiology, 2014
Unspliced Rous sarcoma virus (RSV) retroviral mRNA undergoes nonsense-mediated RNA decay (NMD) if it has premature termination codons in the gag gene. However, its normal gag termination codon is not subject to NMD despite being 7 kb from the 3 0 poly(A) sequence. An RNA stability element (RSE) has been identified immediately downstream of gag in the RSV genome. It appears to determine the proper context for translation termination and protects the RNA from NMD. The viral stability element may prevent Up-frameshift 1 (Upf1) protein from interacting with the terminating ribosome and release factors to initiate NMD.
Nonsense codons within the Rous sarcoma virus gag gene decrease the stability of unspliced viral RNA
Molecular and Cellular Biology, 1991
The intracellular accumulation of the unspliced RNA of Rous sarcoma virus was decreased when translation was prematurely terminated by the introduction of nonsense codons within its 5' proximal gene, the gag gene. In contrast, the levels of spliced viral RNAs were not affected in our transient expression assays in chicken cells. Experiments using the transcription inhibitor dactinomycin showed that mutant unspliced RNAs were degraded more rapidly than wild-type RNA. Furthermore, mutant RNAs could be partially stabilized by coexpression of wild-type gag proteins in trans; however, intact gag proteins were not required to maintain the stability of RNAs which did not contain premature termination codons. Thus, termination codons seemed to destabilize the RNA not because of their effect on gag protein function but instead because they disrupted the process of translating the gag region of the RNA. Analysis of double-mutant constructs containing both deletions and termaination codons within the gag gene also suggested that the stability of the unspliced RNA was affected by a cis-acting interaction between the RNA and ribosomes.
Structure-function relationship of Rous sarcoma virus leader RNA
Nucleic Acids Research, 1982
Cells infected by RSV synthesize viral 35S RNA as well as subgenomic 28S and 22S RN'As coding for the Env and Src genes respectively. In addition, at least the 5' 101 nucleotides of the leader are also conserved and we have shown previously that this sequence contains a strong ribosome binding site (J.-L. Darlix et al., J. Virol. 29, 597). We now report the RNA sequence of Rous Sarcoma virus (RSV) leader RNA and propose a folding of this 5' untranslated region which brings the Cap, the initiation codon for Gag and the strong ribosome binding site close to each other. We also show that ribosomes protect a sequence just upstream from initiator AUG of Gag in vitro, and believed to interact with part of the strong ribosome binding site according to the folding proposed for the leader RNA. 0 I RL Pres Umited, Oxford, England.
Nucleic Acids Research, 1980
The genarnes of mnmerous avian retroviruses contain at their 3' termini a conserved danain denoted "c". The precise boundaries and function of "c" have been enigmas. In an effort to resolve these issues, we determined the sequence of ovrer 900 rncleotides at the 3' end of the genom of the Schmidt-Ruppin subgroup A strain of avian sarcoa virus (ASV). We obtained the sequence from a suitable fragment of ASV IX that had been cloned into the single-stranded E phage M13np2. Coputer-assisted analysis of the sequence revealed the following structural features: i) the length of mc"-473 nucleotides; ii) the 3' terminal domain of src, ending in an amber codon at the 5' boundary of "c"; iii) terminator Cdons that preclude continuous translation from "c"; iv) suitably located sequences that my serve as signals for the initiaticn of viral RIA synthesis and for the processing and/or polyadenylaticn of viral M; v) a repeated sequence that flanks src and that could facilitate deletion of this gene; vi) repeated se quences within wc"; and vii) unexplained hanologies between sequences in "c" and sequences in several other reicacids, including the 5' terminal dmairt.W the ASV genomae, tR and its inversion, the cooplenwt of tR4 and its inversion, and the 18S MA of eukaryotic rib-ones. We conclude that "c" probably does not encode a protein, but its sequence may nevertheless serve several essential functions in viral replication. m1e haploid gerxne of avian saroma virus (ASV) is a singlestranded RI conpoxed of ca. 9500 nucleotides (1). This RW fills tw roles during the viral life cycle. First, it serves as niM for the synthesis of several viral polypeptides (1); accordingly, the RM is capped (2) and polyaderylated (3) at the 5' and 3' termini, respectively. Second, the viral genane is the tenplate for synthesis of viral CMz by reverse transcriptase (1,4); the product of this synthesis is integrated into the gerxne of the host cell and transcribed into progeny viral RA (1,5). Tw structural features of the viral genome have been inplicated in the synthesis of viral DNA: a molecule of tRaTrP located near the 5' end of the viral RIA serves as primer