Antisense oligonucleotides capable of promoting specific target mRNA reduction via competing RNase H1-dependent and independent mechanisms

PLoS One. 2014 Oct 9;9(10):e108625. doi: 10.1371/journal.pone.0108625. eCollection 2014.

Abstract

Antisense oligonucleotides (ASOs) are most commonly designed to reduce targeted RNA via RNase H1-dependent degradation. In this paper we demonstrate that cellular proteins can compete for sites targeted by RNase H1-dependent ASOs. We further show that some ASOs designed to mediate RNase H1 cleavage can, in certain instances, promote target reduction both by RNase H1-mediated cleavage and by steric inhibition of binding of splicing factors at a site required for efficient processing of the pre-mRNA. In the latter case, RNase H cleavage was prevented by binding of a second protein, HSPA8, to the ASO/pre-mRNA heteroduplex. In addition, using a precisely controlled minigene system, we directly demonstrated that activity of ASOs targeting sites in introns is strongly influenced by splicing efficiency.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Line, Tumor
  • HeLa Cells
  • Humans
  • Introns / genetics
  • Oligonucleotides, Antisense / genetics*
  • RNA Precursors / genetics
  • RNA, Messenger / genetics*
  • Ribonuclease H / metabolism*

Substances

  • Oligonucleotides, Antisense
  • RNA Precursors
  • RNA, Messenger
  • Ribonuclease H
  • ribonuclease HI

Grants and funding

This work was supported by ISIS Pharmaceuticals, Inc. The funder provided support in the form of salaries for authors TAV and STC but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.