Volloch V, Schweitzer B, Rits S
Department of Metabolic Regulation, Boston Biomedical Research Institute, MA 02114.
Biochem Biophys Res Commun. 1991 Sep 30;179(3):1600-5. doi: 10.1016/0006-291x(91)91757-4.
The objective of the experiments described in this paper was to test the potential of antisense RNAs complementary to the internal portion of an intron to inhibit the splicing process and to determine the mechanism of such inhibition. The results obtained indicate that RNA fragments complementary to the internal portion of an intron can effectively inhibit the splicing of pre-mRNA. Inhibition was observed only with antisense RNA complementary to pre-mRNA suggesting that the inhibitory effect was due to the formation of a hybrid with the corresponding portion of the pre-mRNA's intron. The observed inhibition was not due to interference with possible intron elements essential for the splicing process, for the deletion of the sequences complementary to inhibitory antisense RNA from the corresponding pre-mRNA molecule did not affect the efficiency of a splicing reaction, and the addition of antisense RNA to pre-mRNA mutants carrying such deletions did not result in any inhibition. Our results indicate that the observed inhibition is a function of the length of the antisense RNA expressed as a fraction of an intron with which it interacts when antisense RNA is modified by incorporation of a "hinge" element, it loses its inhibitory potential suggesting that the inhibitory effect is probably due to limitation of conformational flexibility of an intron.
本文所述实验的目的是测试与内含子内部区域互补的反义RNA抑制剪接过程的潜力，并确定这种抑制作用的机制。所获得的结果表明，与内含子内部区域互补的RNA片段能够有效抑制前体mRNA的剪接。仅在与前体mRNA互补的反义RNA中观察到抑制作用，这表明抑制作用是由于与前体mRNA内含子的相应部分形成了杂交体。观察到的抑制作用并非由于干扰剪接过程中可能必需的内含子元件，因为从相应的前体mRNA分子中删除与抑制性反义RNA互补的序列并不影响剪接反应的效率，并且将反义RNA添加到携带此类缺失的前体mRNA突变体中也不会导致任何抑制作用。我们的结果表明，观察到的抑制作用是反义RNA长度的函数，当反义RNA通过掺入“铰链”元件进行修饰时，它失去了抑制潜力，这表明抑制作用可能是由于内含子构象灵活性的限制。
