Alu-linked hairpins efficiently mediate RNA interference with less toxicity than do H1-expressed short hairpin RNAs.
RNA interference has become a powerful tool for specific inhibition of gene expression in mammalian cells. Expression constructs allow for the long-term delivery of short interfering RNAs, usually through the expression of Pol III-transcribed hairpins. In some instances, these expression systems have been shown to have side effects, including induction of the interferon response and cytotoxicity. Here we demonstrate that H1-expressed hairpins, as well as the cloning vector, reduce the plating efficiency of HeLa cells. This toxicity is abrogated by coexpression of the hairpin in the same transcript as a human Alu repetitive element. These Alu-linked hairpins retain the ability to knock down expression of target mRNAs. This modification, which we term SINE (short interspersed repetitive element)-enhanced short hairpin RNA, provides an alternative expression system for hairpins with reduced side effects.
Gasior
Stephen L
SL
Tulane Cancer Center and Department of Epidemiology, Tulane University Health Sciences Center, New Orleans, LA 70112, USA.
Palmisano
Melanie
M
Deininger
Prescott L
PL
D020087:Alu Elements / Q000502:physiology*; D001483:Base Sequence; D049452:Green Fluorescent Proteins / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008156:Luciferases / Q000235:genetics; D012313:RNA / Q000737:chemistry / Q000502:physiology* / Q000633:toxicity*; D034622:RNA Interference
C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D012313:RNA; D008156:Luciferases