Effect of single-base substitutions in the central domain of virus-associated RNA I on its function

Rahman, A; Malhotra, Pawan; Dhar, Ravi; Kewalramani, T; Thimmapaya, Bayar

doi:10.1128/jvi.69.7.4299-4307.1995

Cited by 19 publications

(9 citation statements)

References 31 publications

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“…This sequence is short and heterogeneous, and insertion of two bases between nt 92 and 93 in Ad2 VA RNA I (VAI-CB [24]) is not deleterious to VA RNA function in vitro. In contrast, mutant pm91 was nonfunctional, presumably because it disrupted the base of the apical stem (56).…”

Section: Discussionmentioning

confidence: 95%

“…As described above, the terminal stem region contains transcription signals (A box; initiation and termination sites). Mutagenesis of Ad2 VA RNA I suggests that the top part of the stem, adjacent to the central domain, is important for inhibiting PKR activation in vitro, while sequences near to the termini can be altered without loss of function (18,24,56). Possibly the duplex adjacent to the central domain acts as a clamp to secure the bottom part of the central domain as in the three-dimensional structure recently proposed (40); the terminal part of the duplex may play a stabilizing role in vivo by protecting the molecule from attack by exonucleases.…”

Section: Discussionmentioning

confidence: 99%

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Structure, function, and evolution of adenovirus-associated RNA: a phylogenetic approach

Mathews

1996

J Virol

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To explore the structure and function of a small regulatory RNA, we examined the virus-associated (VA) RNA species of all 47 known human adenovirus serotypes and of one simian virus, SA7. The VA RNA gene regions of 43 human adenoviruses were amplified and sequenced, and the structures of 10 representative VA RNAs were probed by nuclease sensitivity analysis. Most human viruses have two VA RNA species, VA RNA I and VA RNA II , but nine viruses (19%) have a single VA RNA gene. Sequence alignments classified the RNAs into eight families, corresponding broadly to the known virus groups, and three superfamilies. One superfamily contains the single VA RNAs of groups A and F and the VA RNA I species of group C; the second contains the VA RNA I species of groups B1, D, and E and the unclassified viruses (adenovirus types 42 to 47), as well as the single VA RNAs of group B2; and the third contains all VA RNA II species. Fourteen regions of homology occur throughout the molecule. The longest of these correspond to transcription signals; most of the others participate in RNA secondary structure. The previously identified tetranucleotide pair, GGGU:ACCC, is nearly invariant, diverging slightly (to GGGU:ACCU) only in the two group F viruses and forming a stem in the central domain that is critical for VA RNA structure and function. Secondary structure models which accommodate the nuclease sensitivity data and sequence variations within each family were generated. The major structural features-the terminal stem, apical stem-loop, and central domain-are conserved in all VA RNAs, but differences exist in the apical stem and central domains, especially of the VA RNA II species. Sequence analysis suggests that an ancestral VA RNA gene underwent duplication during the evolution of viruses containing two VA RNA genes. Although the VA RNA II gene seems to have been lost or inactivated by secondary deletion events in some viruses, the high degree of homology among the VA RNA II species implies that this RNA may play an undiscovered role in virus survival. We speculate that the VA RNA genes originated from cellular sequences containing multiple tRNA genes.

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Section: Discussionmentioning

confidence: 95%

Section: Discussionmentioning

confidence: 99%

Structure, function, and evolution of adenovirus-associated RNA: a phylogenetic approach

Mathews

1996

J Virol

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“…was used, as the effect of L3 on ADAR inhibition has not been established); or VA1-PKR5 (PKRÀRNAI+ADAR+inhibitor) in which VA1 incorporated a 1 bp change (pm91) in the central domain. 27 (d) VA1 did not increase transgene expression after genomic integration. Postintegration expression of (1) kanR gWIZ (CMV) compared with (2) AF gWIZ derivative containing the CMV-HTLV-I R promoter (instead of CMV) and the VA1 gene (CMV/AF-HTLV-I-VA1).…”

Section: Nuclear Membrane Transit: Sv40 Enhancer Transient Expressionmentioning

confidence: 86%

“…This is a 2 bp change in the central domain that has been shown to be inactive for PKR inhibition, 26 yet is transcribed at high level in cell culture and maintains the VA RNAIlike RNA secondary structure necessary for RNA interference inhibition. VA1-PKR5 incorporated a 1 bp change (pm91) in the central domain that is strongly reduced for PKR inhibition, 27 but retained adenosine deaminase acting on RNA-inhibiting activity 23 and the VA RNAI-like RNA secondary structure required for RNA interference inhibition. The results ( Figure 4c) suggest that PKR inhibition, and not adenosine deaminase acting on RNA or RNA interference inhibition, was critical for VA1-mediated transgene expression enhancement, as this effect was lost with both PKR3 and PKR5 point mutants.…”

Section: Protein Kinase Regulated By Rna (Pkr) Inhibitor: Va1 Transiementioning

confidence: 99%

Improved antibiotic-free plasmid vector design by incorporation of transient expression enhancers

Luke

Vincent

et al. 2010

Gene Ther

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Methods to improve plasmid-mediated transgene expression are needed for gene medicine and gene vaccination applications. To maintain a low risk of insertional mutagenesis-mediated gene activation, expression-augmenting sequences would ideally function to improve transgene expression from transiently transfected intact plasmid, but not from spurious genomically integrated vectors. We report herein the development of potent minimal, antibiotic-free, high-manufacturing-yield mammalian expression vectors incorporating rationally designed additive combinations of expression enhancers. The SV40 72 bp enhancer incorporated upstream of the cytomegalovirus (CMV) enhancer selectively improved extrachromosomal transgene expression. The human T-lymphotropic virus type I (HTLV-I) R region, incorporated downstream of the CMV promoter, dramatically increased mRNA translation efficiency, but not overall mRNA levels, after transient transfection. A similar mRNA translation efficiency increase was observed with plasmid vectors incorporating and expressing the protein kinase R-inhibiting adenoviral viral associated (VA)1 RNA. Strikingly, HTLV-I R and VA1 did not increase transgene expression or mRNA translation efficiency from plasmid DNA after genomic integration. The vector platform, when combined with electroporation delivery, further increased transgene expression and improved HIV-1 gp120 DNA vaccine-induced neutralizing antibody titers in rabbits. These antibioticfree vectors incorporating transient expression enhancers are safer, more potent alternatives to improve transgene expression for DNA therapy or vaccination.

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“…This is also the case for a pair of highly conserved based-paired tetranucleotides GGGU/ACCC located in the central domain [44,45] This suggests that for VA functionality, the nucleotide sequence is less important than the specific structure. However, since some single point mutations in the central domain reduce VA activity, specific sequences may also have a role in VA functionality [52]. Recently, it has been described that VAI apical stem can adopt two different conformations with different functional activities that co-exist in the infected cell [53].…”

Section: Va Rna Structurementioning

confidence: 99%

Adenovirus and miRNAs

Carnero

Sutherland

Fortes

2011

Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanism

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Adenovirus infection has a tremendous impact on the cellular silencing machinery. Adenoviruses express high amounts of non-coding virus associated (VA) RNAs able to saturate key factors of the RNA interference (RNAi) processing pathway, such as Exportin 5 and Dicer. Furthermore, a proportion of VA RNAs is cleaved by Dicer into viral microRNAs (mivaRNAs) that can saturate Argonaute, an essential protein for miRNA function. Thus, processing and function of cellular miRNAs is blocked in adenoviral-infected cells. However, viral miRNAs actively target the expression of cellular genes involved in relevant functions such as cell proliferation, DNA repair or RNA regulation. Interestingly, the cellular silencing machinery is active at early times post-infection and can be used to control the adenovirus cell cycle. This is relevant for therapeutic purposes against adenoviral infections or when recombinant adenoviruses are used as vectors for gene therapy. Manipulation of the viral genome allows the use of adenoviral vectors to express therapeutic miRNAs or to be silenced by the RNAi machinery leading to safer vectors with a specific tropism. This article is part of a "Special Issue entitled:MicroRNAs in viral gene regulation".

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Effect of single-base substitutions in the central domain of virus-associated RNA I on its function

Cited by 19 publications

References 31 publications

Structure, function, and evolution of adenovirus-associated RNA: a phylogenetic approach

Structure, function, and evolution of adenovirus-associated RNA: a phylogenetic approach

Improved antibiotic-free plasmid vector design by incorporation of transient expression enhancers

Adenovirus and miRNAs

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