Multiple small RNA pathways regulate the silencing of repeated and foreign genes in C. elegans

Abstract

Gene segments from other organisms, such as viruses, are detected as foreign and targeted for silencing by RNAi pathways. A deep-sequencing map of the small RNA response to repeated transgenes introduced to Caenorhabditis elegans revealed that specific segments are targeted by siRNAs. Silencing of the foreign gene segments depends on an antiviral response that involves changes in active and silent chromatin modifications and altered levels of antisense siRNAs. Distinct Argonaute proteins target foreign genes for silencing or protection against silencing. We used a repeated transgene in a genome-wide screen to identify gene disruptions that enhance silencing of foreign genetic elements and identified 69 genes. These genes cluster in four groups based on overlapping sets of coexpressed genes, including a group of germline-expressed genes that are likely coregulated by the E2F transcription factor. Many of the gene inactivations enhance exogenous RNAi. About half of the 69 genes have roles in endogenous RNAi pathways that regulate diverse processes, including silencing of duplicated genes and transposons and chromosome segregation. Of these newly identified genes, several are required for siRNA biogenesis or stability in the oocyte-specific ERGO-1 pathway, including eri-12, encoding an interactor of the RNAi-defective protein RDE-10, and ntl-9/CNOT9, one of several CCR4/NOT complex genes that we identified. The conserved ARF-like small GTPase ARL-8 is required specifically for primary siRNA biogenesis or stability in the sperm-specific ALG-3/4 endogenous RNAi pathway.

Early Developmental Exposure to dsRNA Is Critical for Initiating Efficient Nuclear RNAi in C. elegans

“…Consistent with the known role of these four genes in transgene silencing (Yigit et al, 2006, Grishok et al, 2005, Fischer et al, 2013, Yang et al, 2014, Shirayama et al, 2014, Shiu et al, 2014), expression of the GFP reporter was significantly enhanced in all four mutants, but in no mutant was significant pharyngeal silencing in the first-generation observed (Figure 1B). RDE-1, an Argonaute essential for primary siRNA maturation (Steiner et al, 2009, Sijen et al, 2001), is required for second-generation pharyngeal muscle silencing (Figure 1C).…”

Enhancer of Rudimentary Cooperates with Conserved RNA-Processing Factors to Promote Meiotic mRNA Decay and Facultative Heterochromatin Assembly

“…As HOOD assembly depends on RNAi, Pab2, cryptic introns and splicing factors (Lee et al, 2013; Yamanaka et al, 2013), it is plausible that CCR4-NOT and Pir2 are recruited by splicing factors (and EMC at certain loci) and cooperate with RNAi to assemble HOODs. Supporting this idea, it has been shown that: (1) ARS2 interacts with Dicer in Drosophila and Arabidopsis and is involved in RNAi (Sabin et al, 2009; Yang et al, 2006); (2) C. elegans ntl-9 , a subunit of CCR4-NOT, is required to generate siRNAs from its endogenous targets (Fischer et al, 2013); and (3) human ARS2 co-purifies with the spliceosome (Rappsilber et al, 2002). Alternatively, Pir2 may facilitate HOOD assembly through non-canonical transcription termination.…”

Accumulation of RNA on chromatin disrupts heterochromatic silencing

“…In fission yeast, the Ccr4-Not complex was recently shown also to be involved in deposition of H3K9 methylation at rDNA and meiotic genes (Cotobal et al 2015;Sugiyama et al 2016). Additionally, the Ccr4-Not complex was shown to play a role in silencing of subtelomeric transposons in Caenorhabditis elegans and Drosophila melanogaster (Fischer et al 2013;Morgunova et al 2015). Transposons are targeted by piRNAs in animal germline cells that establish heterochromatin in a similar way to siRNAs in fission yeast (Hirakata and Siomi 2016).…”