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1
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Wu S, Li X, Wang G. tRNA-like structures and their functions. FEBS J 2021; 289:5089-5099. [PMID: 34117728 DOI: 10.1111/febs.16070] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2021] [Revised: 05/12/2021] [Accepted: 06/10/2021] [Indexed: 11/27/2022]
Abstract
tRNA-like structures (TLSs) were first identified in the RNA genomes of turnip yellow mosaic virus. Since then, TLSs have been found in many other species including mammals, and the RNAs harboring these structures range from viral genomic RNAs to mRNAs and noncoding RNAs. Some progress has also been made on understanding their functions that include regulation of RNA replication, translation enhancement, RNA-protein interaction, and more. In this review, we summarize the current knowledge about the regulations and functions of these TLSs. Possible future directions of the field are also briefly discussed.
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Affiliation(s)
- Sipeng Wu
- State Key Laboratory for Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Fujian, China
| | - Xiang Li
- State Key Laboratory for Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Fujian, China
| | - Geng Wang
- State Key Laboratory for Cellular Stress Biology, Innovation Center for Cell Signaling Network, School of Life Sciences, Xiamen University, Fujian, China
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Ariza-Mateos A, Gómez J. Viral tRNA Mimicry from a Biocommunicative Perspective. Front Microbiol 2017; 8:2395. [PMID: 29259593 PMCID: PMC5723415 DOI: 10.3389/fmicb.2017.02395] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2017] [Accepted: 11/20/2017] [Indexed: 12/20/2022] Open
Abstract
RNA viruses have very small genomes which limits the functions they can encode. One of the strategies employed by these viruses is to mimic key factors of the host cell so they can take advantage of the interactions and activities these factors typically participate in. The viral RNA genome itself was first observed to mimic cellular tRNA over 40 years ago. Since then researchers have confirmed that distinct families of RNA viruses are accessible to a battery of cellular factors involved in tRNA-related activities. Recently, potential tRNA-like structures have been detected within the sequences of a 100 mRNAs taken from human cells, one of these being the host defense interferon-alpha mRNA; these are then additional to the examples found in bacterial and yeast mRNAs. The mimetic relationship between tRNA, cellular mRNA, and viral RNA is the central focus of two considerations described below. These are subsequently used as a preface for a final hypothesis drawing on concepts relating to mimicry from the social sciences and humanities, such as power relations and creativity. Firstly, the presence of tRNA-like structures in mRNAs indicates that the viral tRNA-like signal could be mimicking tRNA-like elements that are contextualized by the specific carrier mRNAs, rather than, or in addition to, the tRNA itself, which would significantly increase the number of potential semiotic relations mediated by the viral signals. Secondly, and in particular, mimicking a host defense mRNA could be considered a potential new viral strategy for survival. Finally, we propose that mRNA's mimicry of tRNA could be indicative of an ancestral intracellular conflict in which species of mRNAs invaded the cell, but from within. As the meaning of the mimetic signal depends on the context, in this case, the conflict that arises when the viral signal enters the cell can change the meaning of the mRNAs' internal tRNA-like signals, from their current significance to that they had in the distant past.
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Affiliation(s)
- Ascensión Ariza-Mateos
- Laboratory of RNA Archaeology, Instituto de Parasitología y Biomedicina “López Neyra” (Consejo Superior de Investigaciones Científicas), Granada, Spain
- Centro de Biología Molecular “Severo Ochoa” (CSIC-UAM), Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, Madrid, Spain
| | - Jordi Gómez
- Laboratory of RNA Archaeology, Instituto de Parasitología y Biomedicina “López Neyra” (Consejo Superior de Investigaciones Científicas), Granada, Spain
- Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Madrid, Spain
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Exploiting tRNAs to Boost Virulence. Life (Basel) 2016; 6:life6010004. [PMID: 26797637 PMCID: PMC4810235 DOI: 10.3390/life6010004] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2015] [Revised: 01/08/2016] [Accepted: 01/14/2016] [Indexed: 01/22/2023] Open
Abstract
Transfer RNAs (tRNAs) are powerful small RNA entities that are used to translate nucleotide language of genes into the amino acid language of proteins. Their near-uniform length and tertiary structure as well as their high nucleotide similarity and post-transcriptional modifications have made it difficult to characterize individual species quantitatively. However, due to the central role of the tRNA pool in protein biosynthesis as well as newly emerging roles played by tRNAs, their quantitative assessment yields important information, particularly relevant for virus research. Viruses which depend on the host protein expression machinery have evolved various strategies to optimize tRNA usage—either by adapting to the host codon usage or encoding their own tRNAs. Additionally, several viruses bear tRNA-like elements (TLE) in the 5′- and 3′-UTR of their mRNAs. There are different hypotheses concerning the manner in which such structures boost viral protein expression. Furthermore, retroviruses use special tRNAs for packaging and initiating reverse transcription of their genetic material. Since there is a strong specificity of different viruses towards certain tRNAs, different strategies for recruitment are employed. Interestingly, modifications on tRNAs strongly impact their functionality in viruses. Here, we review those intersection points between virus and tRNA research and describe methods for assessing the tRNA pool in terms of concentration, aminoacylation and modification.
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Díaz-Toledano R, Gómez J. Messenger RNAs bearing tRNA-like features exemplified by interferon alfa 5 mRNA. Cell Mol Life Sci 2015; 72:3747-68. [PMID: 25900662 PMCID: PMC4565877 DOI: 10.1007/s00018-015-1908-0] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2015] [Revised: 03/31/2015] [Accepted: 04/10/2015] [Indexed: 12/24/2022]
Abstract
The purpose of this work was to ascertain whether liver mRNA species share common structural features with hepatitis C virus (HCV) mRNA that allow them to support the RNase-P (pre-tRNA/processing enzyme) cleavage reaction in vitro. The presence of RNase-P competitive elements in the liver mRNA population was determined by means of biochemical techniques, and a set of sensitive mRNA species were identified through microarray screening. Cleavage specificity and substrate length requirement of around 200 nts, were determined for three mRNA species. One of these cleavage sites was found in interferon-alpha 5 (IFNA5) mRNA between specific base positions and with the characteristic RNase-P chemistry of cleavage. It was mapped within a cloverleaf-like structure revealed by a comparative structural analysis based on several direct enzymes and chemical probing methods of three RNA fragments of increasing size, and subsequently contrasted against site-directed mutants. The core region was coincident with the reported signal for the cytoplasmic accumulation region (CAR) in IFNAs. Striking similarities with the tRNA-like element of the antagonist HCV mRNA were found. In general, this study provides a new way of looking at a variety of viral tRNA-like motifs as this type of structural mimicry might be related to specific host mRNA species rather than, or in addition to, tRNA itself.
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Affiliation(s)
- Rosa Díaz-Toledano
- Laboratorio de Arqueología del RNA, Departamento de Bioquímica y Biología Molecular, Instituto de Parasitología y Biomedicina López Neyra (IPBLN-CSIC), Armilla, Granada, Spain.,Centro de Investigación Biológica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Madrid, Spain.,Centro de Biología Molecular Severo Ochoa (UAM-CSIC) Cantoblanco, Madrid, Spain
| | - Jordi Gómez
- Laboratorio de Arqueología del RNA, Departamento de Bioquímica y Biología Molecular, Instituto de Parasitología y Biomedicina López Neyra (IPBLN-CSIC), Armilla, Granada, Spain. .,Centro de Investigación Biológica en Red de Enfermedades Hepáticas y Digestivas (CIBERehd), Madrid, Spain.
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Lee CH, Kim JH, Lee SW. Prospects for nucleic acid-based therapeutics against hepatitis C virus. World J Gastroenterol 2013; 19:8949-8962. [PMID: 24379620 PMCID: PMC3870548 DOI: 10.3748/wjg.v19.i47.8949] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/05/2013] [Revised: 11/10/2013] [Accepted: 11/30/2013] [Indexed: 02/06/2023] Open
Abstract
In this review, we discuss recent advances in nucleic acid-based therapeutic technologies that target hepatitis C virus (HCV) infection. Because the HCV genome is present exclusively in RNA form during replication, various nucleic acid-based therapeutic approaches targeting the HCV genome, such as ribozymes, aptamers, siRNAs, and antisense oligonucleotides, have been suggested as potential tools against HCV. Nucleic acids are potentially immunogenic and typically require a delivery tool to be utilized as therapeutics. These limitations have hampered the clinical development of nucleic acid-based therapeutics. However, despite these limitations, nucleic acid-based therapeutics has clinical value due to their great specificity, easy and large-scale synthesis with chemical methods, and pharmaceutical flexibility. Moreover, nucleic acid therapeutics are expected to broaden the range of targetable molecules essential for the HCV replication cycle, and therefore they may prove to be more effective than existing therapeutics, such as interferon-α and ribavirin combination therapy. This review focuses on the current status and future prospects of ribozymes, aptamers, siRNAs, and antisense oligonucleotides as therapeutic reagents against HCV.
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Abstract
The ability to interfere with gene expression is of crucial importance to unravel the function of genes and is also a promising therapeutic strategy. Here we discuss methodologies for inhibition of target RNAs based on the cleavage activity of the essential enzyme, Ribonuclease P (RNase P). RNase P-mediated cleavage of target RNAs can be directed by external guide sequences (EGSs) or by the use of the catalytic M1 RNA from E. coli linked to a guide sequence (M1GSs). These are not only basic tools for functional genetic studies in prokaryotic and eukaryotic cells but also promising antibacterial, anticancer and antiviral agents.
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Affiliation(s)
- Eirik Wasmuth Lundblad
- Reference Centre for Detection of Antimicrobial Resistance, Department of Microbiology and Infection Control, University Hospital of North Norway, 9038 Tromsø, Norway.
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Abstract
The current standard of care for the treatment of hepatitis C virus infection, pegylated interferon-alpha and ribavirin, is costly, associated with significant side effects, and effective in only 50% of patients. There is therefore a need for the development of novel antiviral therapies. One such approach involves the application of gene silencing technologies, including antisense oligonucleotides, ribozymes, RNA interference, and aptamers. However, despite great scientific advances over the past decade, and promising in vitro data, several significant challenges continue to limit the translation of this technology to the clinical setting. This review provides a concise update of the current literature.
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Affiliation(s)
- Alexander J V Thompson
- Division of Gastroenterology/Hepatology, Duke Clinical Research Institute, Duke University, Durham, NC 27715, USA
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Nadal A, Coll A, Cook N, Pla M. A molecular beacon-based real time NASBA assay for detection of Listeria monocytogenes in food products: role of target mRNA secondary structure on NASBA design. J Microbiol Methods 2007; 68:623-32. [PMID: 17258831 DOI: 10.1016/j.mimet.2006.11.011] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2006] [Revised: 11/06/2006] [Accepted: 11/20/2006] [Indexed: 11/20/2022]
Abstract
A molecular beacon-based real-time NASBA (QNASBA) assay for detection and identification of Listeria monocytogenes has been developed. A correlation between targeting highly accessible mRNA sequences and QNASBA efficiency and sensitivity was demonstrated. The assay targets a sequence from the mRNA transcript of the hly gene which is specific for this bacterium; and includes an internal amplification control to disclose failure of the reaction. It was fully selective and consistently detected down to 100 target molecules and 40 L. monocytogenes exponentially growing cells per reaction. In addition, it was capable of accurate quantification of target RNA molecules independently of the presence of DNA in the sample. In combination with a short RNase treatment prior to nucleic acids extraction our QNASBA specifically detected viable L. monocytogenes cells. It was successfully applied to rapid detection of this pathogen in meat and salmon products, and is therefore a useful tool for the study of L. monocytogenes in food samples. We finally discuss considerations of target secondary structure with regard to development of NASBA assays.
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Affiliation(s)
- Anna Nadal
- Institute of Food and Agricultural Technology, University of Girona, E-17071 Girona, Spain
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Mueller D, Stahl U, Meyer V. Application of hammerhead ribozymes in filamentous fungi. J Microbiol Methods 2006; 65:585-95. [PMID: 16298445 DOI: 10.1016/j.mimet.2005.10.003] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2005] [Revised: 10/11/2005] [Accepted: 10/11/2005] [Indexed: 11/25/2022]
Abstract
Metabolic engineering in filamentous fungi is a emerging field of research as many fungi produce high value primary and secondary metabolites. Ribozyme technology can be used as a tool for metabolic engineering to influence metabolic pathways and to knock down the expression of specific genes of interest. Hammerhead ribozymes can target virtually any mRNA sequence of choice and prevent gene expression on the post-transcriptional level. They are thus a versatile tool for timed and spatial elimination of unwanted gene products. As current research has only investigated the application of ribozymes in bacteria, yeast and mammalian cells, we decided to carry out a study on whether this technology can also function with filamentous fungi. We employed a sensitive, quantitative reporter-based model system as a proof of concept, using the Escherichia coli beta-glucuronidase transcript (uidA) as the target mRNA and Aspergillus giganteus as the host. This system was used to validate the in vivo activities of seven different hammerhead ribozymes, which were selected by in silico analysis of the uidA mRNA. All ribozymes tested were able to reduce the reporter activity up to a maximum of 100%, demonstrating that ribozyme technology is indeed a useful tool in fungal metabolic engineering.
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Affiliation(s)
- Dirk Mueller
- Technische Universität Berlin, Institut für Biotechnologie, Fachgebiet Mikrobiologie und Genetik, Germany
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Trepanier JB, Tanner JE, Alfieri C. Oligonucleotide-Based Therapeutic Options against Hepatitis C Virus Infection. Antivir Ther 2006. [DOI: 10.1177/135965350601100315] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
The hepatitis C virus (HCV) is the cause of a silent pandemic that, due to the chronic nature of the disease and the absence of curative therapy, continues to claim an ever-increasing number of lives. Current antiviral regimens have proven largely unsatisfactory for patients with HCV drug-resistant genotypes. It is therefore important to explore alternative therapeutic stratagems whose mode of action allows them to bypass viral resistance. Antisense oligonucleotides, ribozymes, small interfering RNAs, aptamers and deoxyribozymes constitute classes of oligonucleotide-based compounds designed to target highly conserved or functionally crucial regions contained within the HCV genome. The therapeutic expectation for such compounds is the elimination of HCV from infected individuals. Progress in oligonucleotide-based HCV antivirals towards clinical application depends on development of nucleotide designs that bolster efficacy while minimizing toxicity, improvement in liver-targeting delivery systems, and refinement of small-animal models for preclinical testing.
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Affiliation(s)
- Janie B Trepanier
- Sainte-Justine Hospital Research Centre, and the Department of Microbiology and Immunology, Université de Montréal, Montréal, Québec, Canada
| | | | - Caroline Alfieri
- Sainte-Justine Hospital Research Centre, and the Department of Microbiology and Immunology, Université de Montréal, Montréal, Québec, Canada
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Nadal A, Eritja R, Esteve T, Pla M. "Parallel" and "antiparallel tail-clamps" increase the efficiency of triplex formation with structured DNA and RNA targets. Chembiochem 2005; 6:1034-42. [PMID: 15880676 DOI: 10.1002/cbic.200400358] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Sequence-specific triple-helix structures can be formed by parallel and antiparallel DNA clamps interacting with single-stranded DNA or RNA targets. Single-stranded nucleic acid molecules are known to adopt secondary structures that might interfere with intermolecular interactions. We demonstrate the correlation between a secondary structure involving the target--a stable stem predicted by in silico folding and experimentally confirmed by thermal stability and competition analyses--and an inhibitory effect on triplex formation. We overcame structural impediments by designing a new type of clamp: "tail-clamps". A combination of gel-shift, kinetic analysis, UV thermal melting and thermodynamic techniques was used to demonstrate that tail-clamps efficiently form triple helices with a structured target sequence. The performance of parallel and antiparallel tail-clamps was compared: antiparallel tail-clamps had higher binding efficiencies than parallel tail-clamps both with structured DNA and RNA targets. In addition, the reported triplex-stabilizing property of 8-aminopurine residues was confirmed for tail-clamps. Finally, we discuss the possible use of this improved triplex technology as a new tool for applications in molecular biology.
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Affiliation(s)
- Anna Nadal
- Departament de Genètica Molecular, Institut de Biologia Molecular de Barcelona, Consejo Superior de Investigaciones Científicas, Jordi Girona 18-26, 08034 Barcelona, Spain
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12
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Dinkova TD, Zepeda H, Martínez-Salas E, Martínez LM, Nieto-Sotelo J, de Jiménez ES. Cap-independent translation of maize Hsp101. THE PLANT JOURNAL : FOR CELL AND MOLECULAR BIOLOGY 2005; 41:722-31. [PMID: 15703059 DOI: 10.1111/j.1365-313x.2005.02333.x] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/10/2023]
Abstract
Maize embryonic axes contain stored mRNAs, some of which are able to undergo cap-independent translation initiation during germination. The Hsp101 mRNA, encoding a heat shock protein, is essential for thermo-tolerance induction and is present among the stored transcripts. This research aimed to investigate whether the Hsp101 transcript is IRES-driven regulated upon heat stress. Hsp101 transcribed either in vitro or in vivo was efficiently translated via a cap-independent mechanism. This was observed either in an animal in vitro translation system containing proteolytically cleaved eukaryotic initiation factor eIF4G or in a plant system lacking both eIF4E and eIFiso4E initiation factors. Deletion of the 5' untranslated region (UTR) from the Hsp101 mRNA abolished its cap-independent translation indicating that this nucleotide sequence is required to confer cap-independent initiation. Bicistronic constructs containing the Hsp101 mRNA 5'UTR in sense and anti-sense directions between two reporter genes were translated in both cap-independent systems. A similar bicistronic construct containing a viral internal ribosome entry site (IRES) element between the reporter genes was used as control. Internal translation of the second reporter gene was observed when the Hsp101 5'UTR was in the sense but not in the anti-sense orientation in the bicistronic construct. Taken together, these data suggest that the 5'UTR of maize Hsp101, a plant cellular mRNA, functions as an IRES-like element accounting for its cap-independent translation during heat stress.
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Affiliation(s)
- Tzvetanka D Dinkova
- Departamento de Bioquímica, Facultad de Química, Universidad Nacional Autónoma de México, 04510, México DF
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N/A. N/A. Shijie Huaren Xiaohua Zazhi 2004; 12:2269-2275. [DOI: 10.11569/wcjd.v12.i10.2269] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
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Martinand-Mari C, Lebleu B, Robbins I. Oligonucleotide-based strategies to inhibit human hepatitis C virus. Oligonucleotides 2004; 13:539-48. [PMID: 15025918 DOI: 10.1089/154545703322860834] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022]
Abstract
Hepatitis C virus (HCV) infection represents a worldwide problem, and current antiviral regimens are not satisfactory. The need to develop novel, specific, anti-HCV antiviral drugs is clear. Antisense oligonucleotides (AS-ON), ribozymes, and more recently, small interfering RNAs (siRNAs) have been widely used to control gene expression, and several clinical trials are in progress. The potential to use AS-ON as tools to control HCV infection, either by promoting an RNase H mediated cleavage of viral genomic RNA or by interfering with the assembly of a translation initiation complex on the internal ribosome entry site (IRES) is reviewed. Extensive knowledge of IRES structure and conservation among HCV genotypes have rendered the HCV IRES (and, in particular, its IIId loop) particularly attractive for antisense approaches. Encouraging data have been obtained with IRES-targeted RNase H-competent and incompetent ON analogs. We demonstrate here that very short steric blocking ONs can inhibit the formation of translation preinitiation complexes on the IRES and block IRES-mediated translation in a cell-free translation assay and in a transfected hepatoma cell line.
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Affiliation(s)
- Camille Martinand-Mari
- UMR 5124 CNRS, Laboratoire des Défenses Antivirales et Antitumorales, Université Montpellier 2, 34293 Montpellier Cedex 5, France
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Martell M, Briones C, de Vicente A, Piron M, Esteban JI, Esteban R, Guardia J, Gómez J. Structural analysis of hepatitis C RNA genome using DNA microarrays. Nucleic Acids Res 2004; 32:e90. [PMID: 15247323 PMCID: PMC443556 DOI: 10.1093/nar/gnh088] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Abstract
Many studies have tried to identify specific nucleotide sequences in the quasispecies of hepatitis C virus (HCV) that determine resistance or sensitivity to interferon (IFN) therapy, unfortunately without conclusive results. Although viral proteins represent the most evident phenotype of the virus, genomic RNA sequences determine secondary and tertiary structures which are also part of the viral phenotype and can be involved in important biological roles. In this work, a method of RNA structure analysis has been developed based on the hybridization of labelled HCV transcripts to microarrays of complementary DNA oligonucleotides. Hybridizations were carried out at non-denaturing conditions, using appropriate temperature and buffer composition to allow binding to the immobilized probes of the RNA transcript without disturbing its secondary/tertiary structural motifs. Oligonucleotides printed onto the microarray covered the entire 5' non-coding region (5'NCR), the first three-quarters of the core region, the E2-NS2 junction and the first 400 nt of the NS3 region. We document the use of this methodology to analyse the structural degree of a large region of HCV genomic RNA in two genotypes associated with different responses to IFN treatment. The results reported here show different structural degree along the genome regions analysed, and differential hybridization patterns for distinct genotypes in NS2 and NS3 HCV regions.
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Affiliation(s)
- María Martell
- Laboratorio Medicina Interna-Hepatología, Hospital Vall d'Hebron, Barcelona, Spain.
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