title: Expanding the utility of heptamer-type sgRNA for TRUE gene silencing.
abstract: tRNase Z(L)-utilizing efficacious gene silencing (TRUE gene silencing) is a novel technology for suppressing gene expression. TRUE gene silencing is based on a unique enzymatic property of mammalian tRNase Z(L), which is that it can cleave any target RNA at any desired site by recognizing a pre-tRNA-like or micro-pre-tRNA-like complex formed between the target RNA and artificial small guide RNA (sgRNA). sgRNA is divided into four groups, 5'-half-tRNA, RNA heptamer, hook RNA, and ∼14-nt linear RNA. One of the final destinations of TRUE gene silencing is to generate cancer therapeutic sgRNAs, and from a pharmacological point of view, heptamer-type sgRNA appears to be the most appropriate for this purpose. In this paper, we present two strategies to expand the utility of heptamer-type sgRNA: one is about locked nucleic acid (LNA) modifications of heptamers and the other is about usage of double heptamers. We showed that RNA heptamers with LNA modifications can work as sgRNA in vitro and in vivo. We also demonstrated that two consecutively aligned heptamers can guide target RNA cleavage by human tRNase Z(L) as efficiently as a corresponding 14-nt sgRNA in vitro and that a double heptamer can work much better than a 14-nt sgRNA in vivo.
authors.affiliation: Department of Applied Life Sciences, Niigata University of Pharmacy and Applied Life Sciences, Niigata 956-8603, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Sano
authors.forename: Masayuki
authors.initials: M
authors.lastname: Takahashi
authors.forename: Tadasuke
authors.initials: T
authors.lastname: Nozaki
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Takahashi
authors.forename: Masato
authors.initials: M
authors.lastname: Tamura
authors.forename: Masayuki
authors.initials: M
authors.lastname: Nashimoto
mesh_terms: D004722:Endoribonucleases / Q000737:chemistry / Q000378:metabolism*; D055785:Gene Knockdown Techniques / Q000379:methods*; D020868:Gene Silencing; D057809:HEK293 Cells; D006801:Humans; D019169:Jurkat Cells; D009841:Oligonucleotides / Q000737:chemistry*; D059367:RNA Cleavage; D034741:RNA, Small Interfering / Q000737:chemistry* / Q000235:genetics; D058727:RNA, Small Untranslated
chemical_list: D009841:Oligonucleotides; D034741:RNA, Small Interfering; C477371:locked nucleic acid; D004722:Endoribonucleases; C531011:tRNase Z, human; D058727:RNA, Small Untranslated