Canonical small interfering RNAs (siRNAs) are processed from double-stranded RNA (dsRNA) by Dicer and associate with Argonautes to direct RNA silencing. In Caenorhabditis elegans, 22G-RNAs and 26G-RNAs are often referred to as siRNAs but display distinct characteristics. For example, 22G-RNAs do not originate from dsRNA and do not depend on Dicer, whereas 26G-RNAs require Dicer but derive from an atypical RNA duplex and are produced exclusively antisense to their messenger RNA (mRNA) templates. To identify canonical siRNAs in C. elegans, we first characterized the siRNAs produced via the exogenous RNA interference (RNAi) pathway. During RNAi, dsRNA is processed into ∼23 nt duplexes with ∼2 nt, 3′-overhangs, ultimately yielding siRNAs devoid of 5′G-containing sequences that bind with high affinity to the Argonaute RDE-1, but also to the microRNA (miRNA) pathway Argonaute, ALG-1. Using these characteristics, we searched for their endogenous counterparts and identified thousands of endogenous loci representing dozens of unique elements that give rise to mostly low to moderate levels of siRNAs, called 23H-RNAs. These loci include repetitive elements, putative coding genes, pseudogenes, noncoding RNAs, and unannotated features, many of which adopt hairpin (hp) structures reminiscent of the hpRNA/RNAi pathway in flies and mice. RDE-1 competes with other Argonautes for binding to 23H-RNAs. When RDE-1 is depleted, these siRNAs are enriched in ALG-1 and ALG-2 complexes. Our results expand the known repertoire of C. elegans small RNAs and their Argonaute interactors, and demonstrate that key features of the endogenous siRNA pathway are relatively unchanged in animals.

中文翻译：

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一类低丰度的 Dicer 依赖性 siRNA，由秀丽隐杆线虫中的多种特征产生


经典小干扰 RNA （siRNA） 由 Dicer 从双链 RNA （dsRNA） 加工而成，并与 Argonautes 结合以指导 RNA 沉默。在秀丽隐杆线虫中，22G-RNA 和 26G-RNA 通常被称为 siRNA，但表现出不同的特征。例如，22G-RNA 不是源自 dsRNA 且不依赖于 Dicer，而 26G-RNA 需要 Dicer，但源自非典型 RNA 双链体，并且完全与其信使 RNA （mRNA） 模板产生反义。为了鉴定秀丽隐杆线虫中的经典 siRNA，我们首先表征了通过外源性 RNA 干扰 （RNAi） 途径产生的 siRNA。在 RNAi 过程中，dsRNA 被加工成具有 ∼2 nt、3′-突出端的 ∼23 nt 双链体，最终产生不含 5′G 序列的 siRNA，这些序列以高亲和力结合 Argonaute RDE-1，但也与 microRNA （miRNA） 通路 Argonaute， ALG-1 结合。利用这些特征，我们搜索了它们的内源性对应物，并确定了数千个内源性基因座，这些基因座代表了数十种独特元素，这些元素产生了大多数低到中等水平的 siRNA，称为 23H-RNA。这些基因座包括重复元件、推定编码基因、假基因、非编码 RNA 和未注释的特征，其中许多采用让人想起果蝇和小鼠的 hpRNA/RNAi 通路的发夹 （hp） 结构。RDE-1 与其他 Argonautes 竞争结合 23H-RNA。当 RDE-1 耗尽时，这些 siRNA 在 ALG-1 和 ALG-2 复合物中富集。我们的结果扩展了秀丽隐杆线虫小 RNA 及其 Argonaute 相互作用物的已知库，并表明内源性 siRNA 通路的关键特征在动物中相对没有变化。