Division of Cell and Developmental Biology, College of Life Sciences, University of Dundee, Dundee, United Kingdom.
PLoS One. 2010 Dec 17;5(12):e15264. doi: 10.1371/journal.pone.0015264.
The Drosophila argonaute2 (ago2) gene plays a major role in siRNA mediated RNA silencing pathways. Unlike mammalian Argonaute proteins, the Drosophila protein has an unusual amino-terminal domain made up largely of multiple copies of glutamine-rich repeats (GRRs). We report here that the ago2 locus produces an alternative transcript that encodes a putative short isoform without this amino-terminal domain. Several ago2 mutations previously reported to be null alleles only abolish expression of the long, GRR-containing isoform. Analysis of drop out (dop) mutations had previously suggested that variations in GRR copy number result in defects in RNAi and embryonic development. However, we find that dop mutations genetically complement transcript-null alleles of ago2 and that ago2 alleles with variant GRR copy numbers support normal development. In addition, we show that the assembly of the central RNAi machinery, the RISC (RNA induced silencing complex), is unimpaired in embryos when GRR copy number is altered. In fact, we find that GRR copy number is highly variable in natural D. melanogaster populations as well as in laboratory strains. Finally, while many other insects share an extensive, glutamine-rich Ago2 amino-terminal domain, its primary sequence varies drastically between species. Our data indicate that GRR variation does not modulate an essential function of Ago2 and that the amino-terminal domain of Ago2 is subject to rapid evolution.
果蝇 Argonaute2（ago2）基因在 siRNA 介导的 RNA 沉默途径中发挥主要作用。与哺乳动物 Argonaute 蛋白不同，果蝇蛋白具有不寻常的氨基末端结构域，主要由多个富含谷氨酰胺的重复序列（GRR）组成。我们在此报告ago2 基因座产生另一种转录本，该转录本编码一种没有该氨基末端结构域的假定短同工型。先前报道的几种被认为是无效等位基因的 ago2 突变仅会使长的、含有 GRR 的同工型的表达失活。对dropout（dop）突变的分析先前表明，GRR 拷贝数的变化导致 RNAi 和胚胎发育缺陷。然而，我们发现 dop 突变在遗传上可以与 ago2 的转录本缺失等位基因互补，并且具有不同 GRR 拷贝数的 ago2 等位基因支持正常发育。此外，我们表明中央 RNAi 机制 RISC（RNA 诱导沉默复合物）的组装在改变 GRR 拷贝数时在胚胎中不受影响。事实上，我们发现 GRR 拷贝数在自然的黑腹果蝇种群以及实验室品系中高度可变。最后，虽然许多其他昆虫共享广泛的富含谷氨酰胺的 Ago2 氨基末端结构域，但它们的一级序列在物种之间差异很大。我们的数据表明，GRR 变异不会调节 Ago2 的基本功能，并且 Ago2 的氨基末端结构域受到快速进化的影响。
