title: Molecular basis for improved gene silencing by Dicer substrate interfering RNA compared with other siRNA variants.
abstract: The canonical exogenous trigger of RNA interference (RNAi) in mammals is small interfering RNA (siRNA). One promising application of RNAi is siRNA-based therapeutics, and therefore the optimization of siRNA efficacy is an important consideration. To reduce unfavorable properties of canonical 21mer siRNAs, structural and chemical variations to canonical siRNA have been reported. Several of these siRNA variants demonstrate increased potency in downstream readout-based assays, but the molecular mechanism underlying the increased potency is not clear. Here, we tested the performance of canonical siRNAs and several sequence-matched variants in parallel in gene silencing, RNA-induced silencing complex (RISC) assembly, stability and Argonaute (Ago) loading assays. The commonly used 19mer with two deoxythymidine overhangs (19merTT) variant performed similarly to canonical 21mer siRNA. A shorter 16mer variant (16merTT) did not perform comparably in our assays. Dicer substrate interfering RNA (dsiRNA) demonstrated better gene silencing by the guide strand (target complementary strand), better RISC assembly, persistence of the guide strand and relatively more loading of the guide strand into Ago. Hence, we demonstrate the advantageous properties of dsiRNAs at upstream, intermediate and downstream molecular steps of the RNAi pathway.
authors.affiliation: Irell and Manella Graduate School of Biological Sciences, Beckman Research Institute of City of Hope, 1500 E. Duarte Rd., Duarte, CA 91010, USA.
authors.forename: Nicholas M
authors.initials: NM
authors.lastname: Snead
authors.forename: Xiwei
authors.initials: X
authors.lastname: Wu
authors.forename: Arthur
authors.initials: A
authors.lastname: Li
authors.forename: Qi
authors.initials: Q
authors.lastname: Cui
authors.forename: Kumi
authors.initials: K
authors.lastname: Sakurai
authors.forename: John C
authors.initials: JC
authors.lastname: Burnett
authors.forename: John J
authors.initials: JJ
authors.lastname: Rossi
mesh_terms: D060565:Argonaute Proteins / Q000378:metabolism; D001483:Base Sequence; D057809:HEK293 Cells; D006801:Humans; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000737:chemistry*; D043244:Ribonuclease III / Q000378:metabolism*
chemical_list: D060565:Argonaute Proteins; D034741:RNA, Small Interfering; D043244:Ribonuclease III