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Influence of untranslated regions on retroviral mRNA transfer and expression.

Abstract : BACKGROUND: Deliberate cellular reprogramming is becoming a realistic objective in the clinic. While the origin of the target cells is critical, delivery of bioactive molecules to trigger a shift in cell-fate remains the major hurdle. To date, several strategies based either on non-integrative vectors, protein transfer or mRNA delivery have been investigated. In a recent study, a unique modification in the retroviral genome was shown to enable RNA transfer and its expression. RESULTS: Here, we used the retroviral mRNA delivery approach to study the impact of modifying gene-flanking sequences on RNA transfer. We designed modified mRNAs for retroviral packaging and used the quantitative luciferase assay to compare mRNA expression following viral transduction of cells. Cloning the untranslated regions of the vimentin or non-muscular myosin heavy chain within transcripts improved expression and stability of the reporter gene while slightly modifying reporter-RNA retroviral delivery. We also observed that while the modified retroviral platform was the most effective for retroviral mRNA packaging, the highest expression in target cells was achieved by the addition of a non-viral UTR to mRNAs containing the packaging signal. CONCLUSIONS: Through molecular engineering we have assayed a series of constructs to improve retroviral mRNA transfer. We showed that an authentic RNA retroviral genomic platform was most efficiently transferred but that adding UTR sequences from highly expressed genes could improve expression upon transfection while having only a slight effect on expression from transferred RNA. Together, these data should contribute to the optimisation of retroviral mRNA-delivery systems that test combinations of UTRs and packaging platforms.
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Submitted on : Wednesday, May 1, 2013 - 5:03:14 PM
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Anne Prel, Luc Sensébé, Jean-Christophe Pagès. Influence of untranslated regions on retroviral mRNA transfer and expression.. BMC Biotechnology, BioMed Central, 2013, 13 (1), pp.35. ⟨10.1186/1472-6750-13-35⟩. ⟨inserm-00819499⟩



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