Abasic pivot substitution harnesses target specificity of RNA interference

Hye Sook Lee, Heeyoung Seok, Dong Ha Lee, Juyoung Ham, Wooje Lee, Emilia Moonkyung Youm, Jin Seon Yoo, Yong Seung Lee, Eun Sook Jang, Sung Wook Chi

Research output: Contribution to journalArticlepeer-review

39 Citations (Scopus)

Abstract

Gene silencing via RNA interference inadvertently represses hundreds of off-target transcripts. Because small interfering RNAs (siRNAs) can function as microRNAs, avoiding miRNA-like off-target repression is a major challenge. Functional miRNA-target interactions are known to pre-require transitional nucleation, base pairs from position 2 to the pivot (position 6). Here, by substituting nucleotide in pivot with abasic spacers, which prevent base pairing and alleviate steric hindrance, we eliminate miRNA-like off-target repression while preserving on-target activity at ∼80-100%. Specifically, miR-124 containing dSpacer pivot substitution (6pi) loses seed-mediated transcriptome-wide target interactions, repression activity and biological function, whereas other conventional modifications are ineffective. Application of 6pi allows PCSK9 siRNA to efficiently lower plasma cholesterol concentration in vivo, and abolish potentially deleterious off-target phenotypes. The smallest spacer, C3, also shows the same improvement in target specificity. Abasic pivot substitution serves as a general means to harness the specificity of siRNA experiments and therapeutic applications.

Original languageEnglish
Article number10154
JournalNature communications
Volume6
DOIs
Publication statusPublished - 2015 Dec 18

Bibliographical note

Funding Information:
We thank the members of the Chi laboratories for helpful discussions, K.-K. Kim for initiation and help with bioinformatics analyses. Thanks to S. Hohng for help with in vitro Ago2 cleavage assay. This work was supported by Korea University Grant, a grant from the Korean Health Technology R&D Project, Ministry of Health and Welfare, Republic of Korea (HI11C1931, HI14C2353), grants from the Basic Science Research Program (NRF-2013R1A1A1008579) and the Science Research Center Program (NRF-2015R1A5A1009024) through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning. H.S. was supported, in part, by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (NRF-2014R1A1A2A16055016).

ASJC Scopus subject areas

  • General Chemistry
  • General Biochemistry,Genetics and Molecular Biology
  • General Physics and Astronomy

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