Vickers Timothy A, Crooke Stanley T
Department of Core Antisense Research, ISIS Pharmaceuticals, Inc., Carlsbad, California, United States of America.
PLoS One. 2014 Oct 9;9(10):e108625. doi: 10.1371/journal.pone.0108625. eCollection 2014.
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.
反义寡核苷酸（ASO）最常用于通过核糖核酸酶H1依赖性降解来减少靶向RNA。在本文中，我们证明细胞蛋白可以竞争核糖核酸酶H1依赖性ASO靶向的位点。我们进一步表明，一些旨在介导核糖核酸酶H1切割的ASO在某些情况下，既能通过核糖核酸酶H1介导的切割促进靶标减少，也能通过空间位阻抑制剪接因子在加工前体mRNA所需位点的结合来促进靶标减少。在后一种情况下，第二种蛋白质HSPA8与ASO/前体mRNA异源双链体结合，阻止了核糖核酸酶H的切割。此外，我们使用精确控制的小基因系统直接证明，靶向内含子中位点的ASO活性受到剪接效率的强烈影响。
