Department of Biochemistry and Biophysics, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, USA.
Mol Cell Biol. 2010 Sep;30(18):4368-78. doi: 10.1128/MCB.00384-10. Epub 2010 Jul 20.
Telomerase RNA is an essential component of telomerase, a ribonucleoprotein enzyme that maintains chromosome ends in most eukaryotes. Here we employ a novel approach, namely, RNA-guided RNA modification, to assess whether introducing 2'-O methylation into telomerase RNA can influence telomerase activity in vivo. We generate specific 2'-O methylation sites in and adjacent to the triple helix (within the conserved pseudoknot structure) of Saccharomyces cerevisiae telomerase RNA (TLC1). We show that 2'-O methylation at U809 reduces telomerase activity, resulting in telomere shortening, whereas 2'-O methylation at A804 or A805 leads to moderate telomere lengthening. Importantly, we also show that targeted 2'-O methylation does not affect TLC1 levels and that 2'-O-methylated TLC1 appears to be efficiently assembled into telomerase ribonucleoprotein. Our results demonstrate that RNA-guided RNA modification is a highly useful approach for modulating telomerase activity.
端粒酶 RNA 是端粒酶的一个重要组成部分，端粒酶是一种核糖核蛋白酶，在大多数真核生物中维持染色体末端。在这里，我们采用了一种新的方法，即 RNA 引导的 RNA 修饰，来评估在端粒酶 RNA 中引入 2'-O 甲基化是否会影响体内的端粒酶活性。我们在酿酒酵母端粒酶 RNA（TLC1）的三螺旋（保守假结结构内）内和附近产生特定的 2'-O 甲基化位点。我们表明，U809 的 2'-O 甲基化降低了端粒酶活性，导致端粒缩短，而 A804 或 A805 的 2'-O 甲基化导致适度的端粒延长。重要的是，我们还表明，靶向 2'-O 甲基化不会影响 TLC1 水平，并且 2'-O-甲基化的 TLC1 似乎能够有效地组装成端粒酶核糖核蛋白。我们的结果表明，RNA 引导的 RNA 修饰是一种非常有用的调节端粒酶活性的方法。
