title: Novel expression vectors enabling induction of gene expression by small-interfering RNAs and microRNAs.
abstract: Small-interfering RNAs and microRNAs are small ∼21-22 nucleotide long RNAs capable of posttranscriptional suppression of gene expression. The synthetic siRNAs are especially designed to target pre-specified genes and are common molecular biology tools. The miRNAs are endogenous regulators of gene expression found in a wide variety of eukaryotes. miRNAs are currently utilized for diagnostics applications. Therapeutically, various miRNA-antagonizing tools are being explored and miRNAs are also utilized for cell-specific inhibition of the expression of gene therapy vectors harboring target sites for specific miRNAs. Here we show, for the first time, that siRNAs and miRNAs can be harnessed to induce gene expression. We designed special expression vectors in which target sites for artificial siRNAs or endogenous miRNAs are located between the transgene and an Upstream Inhibitory Region (UIR). We hypothesized that cleavage of the mRNA by siRNAs or miRNAs will separate the transgene from the UIR and the resulting uncapped mRNA will be capable of being translated. A UIR composed of seven open reading frames was found to be the most efficient inhibitor of the translation of the downstream transgene. We show that under such a configuration both artificial siRNAs and endogenous miRNAs were capable of inducing transgene expression. We show that using the diphtheria toxin A-chain gene, in combination with target sites for highly expressed miRNAs, specific induction of cell-death can be achieved, setting the stage for application to cancer therapy.
authors.affiliation: Nanodoc Biotechnology, 7 Sapir Str., Nes Ziona 7403630, Israel.
authors.forename: Liraz
authors.initials: L
authors.lastname: Harel
authors.affiliation: Nanodoc Biotechnology, 7 Sapir Str., Nes Ziona 7403630, Israel.
authors.forename: Nir
authors.initials: N
authors.lastname: Gefen
authors.affiliation: Nanodoc Biotechnology, 7 Sapir Str., Nes Ziona 7403630, Israel.
authors.forename: Ofira
authors.initials: O
authors.lastname: Carmi
authors.affiliation: Nanodoc Biotechnology, 7 Sapir Str., Nes Ziona 7403630, Israel.
authors.forename: Pini
authors.initials: P
authors.lastname: Orbach
authors.affiliation: Dr. Paz Einat Biotechnology Projects & Consulting, 45a Moshe Levi Str., Nes Ziona 74207, Israel.
authors.forename: Paz
authors.initials: P
authors.lastname: Einat
authors.affiliation: Nanodoc Biotechnology, 7 Sapir Str., Nes Ziona 7403630, Israel.
authors.forename: Guy
authors.initials: G
authors.lastname: Abitbol
mesh_terms: D004167:Diphtheria Toxin; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D016366:Open Reading Frames / Q000235:genetics; D010446:Peptide Fragments; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D019076:Transgenes / Q000235:genetics
chemical_list: D004167:Diphtheria Toxin; D035683:MicroRNAs; D010446:Peptide Fragments; D034741:RNA, Small Interfering; C029795:diphtheria toxin fragment A