Obálka

Utility of Epstein-Barr virus-encoded small RNA promoters for driving the expression of fusion transcripts harboring short hairpin RNAs

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Vydáno v:Gene Therapy vol. 15, no. 3 (Feb 2008), p. 191-202
Hlavní autor: Choy, E Y-W
Další autoři: Kok, K-H, Tsao, S W, Jin, D-Y
Vydáno:
Nature Publishing Group
Témata:
Cell Line, Tumor
Feasibility Studies
Gene Expression
Gene Silencing
Genetic Engineering
Genetic Therapy > methods
Genetic Vectors > pharmacology
Herpesvirus 4, Human > genetics
Humans
Nasopharyngeal Neoplasms > metabolism
Nasopharyngeal Neoplasms > therapy
Promoter Regions, Genetic
RNA Interference
RNA Polymerase III > genetics
RNA, Messenger > analysis
RNA, Small Interfering
Recombinant Fusion Proteins > genetics
Viruses
Gene therapy
Cellular biology
Ribonucleic acid > RNA
Epstein-Barr virus
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Abstrakt:  To induce RNA interference (RNAi), either small interfering RNAs (siRNAs) are directly introduced into the cell or short hairpin RNAs (shRNAs) are expressed from a DNA vector. At present, shRNAs are commonly synthesized by RNA polymerase III (Pol III) promoters of the H1 and U6 RNAs. In this study, we designed and characterized a new set of plasmid vectors driven by promoters of the Epstein-Barr virus (EBV)-encoded small RNAs (EBERs). The EBERs are the most abundant transcript in infected cells and they are transcribed by Pol III. We showed that the EBER promoters were able to drive the expression of shRNA fusion transcripts. siRNAs processed from these fusion transcripts specifically and effectively inhibited the expression of homologous reporter or endogenous genes in various types of cells. The partial EBER sequences in the fusion transcripts did not activate double-stranded RNA-dependent protein kinase or suppress RNAi. In nasopharyngeal carcinoma cells, the EBER2 promoter was stronger than the H1 and U6 promoters in shRNA synthesis, leading to more effective knockdown of the target genes. Taken together, our findings suggest that the EBER promoters fundamentally different from those of H1 and U6 can be used to drive the intracellular expression of shRNAs for effective silencing of target genes in mammalian cells and particularly, in EBV-infected cells.
ISSN:0969-7128
1476-5462
DOI:10.1038/sj.gt.3303055
Zdroj:Medical Database