![]() Finally, we show that slicer activity is required for the siRNA-dependent association of Ago1 with chromatin and for the spreading of histone H3-K9 methylation. Argonaute proteins found in animals are classified into three major groups, based on their interaction with specific small RNA species. ![]() elegans contains 27 AGO genes, raising the question of what roles these genes play in RNAi and related gene-silencing. In humans, Argonaute-encoding genes are phylogenetically grouped into two different clades called the Argonaute-like and the Piwi-like proteins that are, respectively, required for the miRNA and PIWI-interacting RNA (piRNA) pathways (reviewed in ( 2 )). Consistent with this observation, purified Arb1 inhibits the slicer activity of Ago1 in vitro, and purified catalytically inactive Ago1 contains only double-stranded siRNA. Argonaute (AGO) proteins interact with small RNAs to mediate gene silencing. (A) Deeper knockdown is achieved by 24 h compared to 3 h in all Rab5c siRNA treated cells, except MEF Ago2 / treated with CDS-targeting siRNA, where no significant knockdown is detected at either time point. Furthermore, whereas siRNAs in the RITS complex are mostly single-stranded, siRNAs associated with ARC are mostly double-stranded, indicating that Arb1 and Arb2 inhibit the release of the siRNA passenger strand from Ago1. In addition, it shown that the siRNA-class AGO lineage is characterized by high rates of molecular evolution, suggesting a role in the arms race with viruses, while the miRNA-class AGOs display. Figure S2: Features of siRNA-mediated degradation of mRNA in the absence of Ago2. We also find that ALG-1/2 can bind and use a Dicer-processed miRNA duplex to target mRNAs, suggesting an ability to displace RNA strands. Structural determinants of Argonaute proteins for small RNA sorting Argonaute (AGO) proteins provide numerous possibilities for RNAprotein interactions that might underlie the proposed determinants of small RNA strand sorting. ![]() Here, we identify a second fission yeast Argonaute complex (Argonaute siRNA chaperone, ARC), which contains, in addition to Ago1, two previously uncharacterized proteins, Arb1 and Arb2, both of which are required for histone H3 Lys9 (H3-K9) methylation, heterochromatin assembly and siRNA generation. Here, we report that miRNA-specific ALG-1 and ALG-2 Argonautes from Caenorhabditis elegans possess the slicing activity normally implicated in the siRNA-silencing pathway. The RNA-induced transcriptional silencing (RITS) complex, containing Ago1, Chp1, Tas3 and centromeric small interfering RNAs (siRNAs), is required for heterochromatic gene silencing at centromeres. ![]()
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