Adenovirus Virus-Associated RNA Is Processed to Functional Interfering RNAs Involved in Virus Production

Aparicio, Oscar M.; Razquin, Nerea; Zaratiegui, Mikel; Narvaiza, Iñigo; Fortes, Puri

doi:10.1128/jvi.80.3.1376-1384.2006

Cited by 133 publications

(170 citation statements)

References 50 publications

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“…9 Luciferase was quantified from images obtained using the Living Image 2.20 software (Xenogen). Firefly luciferase activity was quantified in liver extracts by using the Luciferase System (Promega), as described previously, 35 in a Berthold Luminometer (Lumat LB 9507). Protein concentration was assayed by the Bradford method following the recommendations of the supplier.…”

Section: Analysis Of Luciferase Expressionmentioning

confidence: 99%

AAV vectors transduce hepatocytes in vivo as efficiently in cirrhotic as in healthy rat livers

et al. 2011

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In liver cirrhosis, abnormal liver architecture impairs efficient transduction of hepatocytes with large viral vectors such as adenoviruses. Here we evaluated the ability of adeno-associated virus (AAV) vectors, small viral vectors, to transduce normal and cirrhotic rat livers. Using AAV serotype-1 (AAV1) encoding luciferase (AAV1Luc) we analyzed luciferase expression with a CCD camera. AAV1Luc was injected through the hepatic artery (intra-arterial (IA)), the portal vein (intra-portal (IP)), directly into the liver (intra-hepatic (IH)) or infused into the biliary tree (intra-biliar). We found that AAV1Luc allows long-term and constant luciferase expression in rat livers. Interestingly, IP administration leads to higher expression levels in healthy than in cirrhotic livers, whereas the opposite occurs when using IA injection. IH administration leads to similar transgene expression in cirrhotic and healthy rats, whereas intra-biliar infusion is the least effective route. After 70% partial hepatectomy, luciferase expression decreased in the regenerating liver, suggesting lack of efficient integration of AAV1 DNA into the host genome. AAV1Luc transduced mainly the liver but also the testes and spleen. Within the liver, transgene expression was found mainly in hepatocytes. Using a liver-specific promoter, transgene expression was detected in hepatocytes but not in other organs. Our results indicate that AAVs are convenient vectors for the treatment of liver cirrhosis.

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Section: Analysis Of Luciferase Expressionmentioning

confidence: 99%

AAV vectors transduce hepatocytes in vivo as efficiently in cirrhotic as in healthy rat livers

et al. 2011

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“…VA1 RNAs are 160-nt long and can be recognized by Exp-5 via their minihelix RNA motif (63), which are dsRNA molecules similar in structure to pre-miRNAs (64). Exp-5 directly interacts with VA1 RNA in a Ran·GTP-dependent manner (63).…”

Section: Exportin-5 and Adenovirus Infectionmentioning

confidence: 99%

“…Later, it was found that small RNAs derived from VA1 RNAs, similar to miRNAs, can be found in adenovirus-infected cells (64). These small RNAs are efficiently bound by Ago2, and behave as functional siRNAs, in that they inhibit the expression of reporter genes with complementary sequences.…”

Section: Exportin-5 and Adenovirus Infectionmentioning

confidence: 99%

“…These small RNAs are efficiently bound by Ago2, and behave as functional siRNAs, in that they inhibit the expression of reporter genes with complementary sequences. Inhibition of small VA1 RNA function negatively affected adenovirus production, indicating that they are required for optimal virus replication (64). Adenoviruses thus appear to utilize the endogenous miRNA-guided RNA silencing pathway to their advantage in viral production.…”

Section: Exportin-5 and Adenovirus Infectionmentioning

confidence: 99%

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Protein Components of the microRNA Pathway and Human Diseases

Perron¹,

Provost²

2008

Methods in Molecular Biology

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SummaryMicroRNAs (miRNAs) are key regulators of messenger RNA (mRNA) translation known to be involved in a wide variety of cellular processes. In fact, their individual importance is reflected in the diseases that may arise upon the loss, mutation or dysfunction of specific miRNAs. It has been appreciated only recently that diseases may also develop when the protein components of the miRNA machinery itself are affected. The core enzymes of the major protein complexes involved in miRNA biogenesis and function, such as the ribonucleases III (RNases III) Drosha and Dicer as well as Argonaute 2 (Ago2), appear to be essential. However, the accessory proteins of the miRNA pathway, such as the DiGeorge syndrome critical region gene 8 (DGCR8) protein, Exportin-5 (Exp-5), TAR RNA binding protein (TRBP) and Fragile X mental retardation protein (FMRP), are each related, in various ways, to specific genetic diseases.

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“…43,44 In light of this, we carried out additional experiments to address whether EBER RNAs might influence RNAi. Results from our control experiments shown in Figures 3a and b indicated that the partial EBER sequences produced from pEBER1-T6 and pE-BER2-T6 had minimal influence on the expression of luciferase reporter.…”

Section: Induction Of Rnai By Eber-shrna Transcriptsmentioning

confidence: 99%

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

et al. 2007

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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.

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Adenovirus Virus-Associated RNA Is Processed to Functional Interfering RNAs Involved in Virus Production

Cited by 133 publications

References 50 publications

AAV vectors transduce hepatocytes in vivo as efficiently in cirrhotic as in healthy rat livers

AAV vectors transduce hepatocytes in vivo as efficiently in cirrhotic as in healthy rat livers

Protein Components of the microRNA Pathway and Human Diseases

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

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