Transcriptome analysis of C. elegans embryos lacking ADARs and the 26G pathway

Adenosine deaminases that act on RNA (ADARs) catalyze the conversion of adenosine to inosine in dsRNA. C. elegans ADARs, ADR-1 and ADR-2, promote the expression of genes containing dsRNA structures by preventing their processing into siRNAs and silencing by RNAi. The 26G endogenous siRNA (endo-siRNA) pathway generates a subset of siRNAs distinct from those made in adr-1;adr-2 mutants, but using many of the same factors. We found that adr-1;adr-2;rrf-3 mutants, lacking both ADARs and the RNA-dependent RNA polymerase RRF-3 required for the 26G pathway, display a bursting phenotype rescued by the RNAi factors RDE-1 and RDE-4. To determine what gene expression changes underlie the synthetic phenotype of adr-1;adr-2;rrf-3 mutants, we sequenced poly(A)+ RNA from adr-1;adr-2;rrf-3 embryos, their parent strains, and strains rescued with mutations in rde-1 and rde-4. We found that genes associated with edited structures were robustly downregulated in adr-1;adr-2;rrf-3 mutants in a manner partially dependent on rde-1 and rde-4. Additionally, genes induced during Orsay virus infections were induced in rrf-3 mutants and further upregulated in adr-1;adr-2;rrf-3 mutants, again dependent in part on rde-1 and rde-4. Overall design: RNAseq of poly(A)+ RNA from C.elegans mixed-stage embryos, four biological replicates per genotype, six genotypes: wildtype (Bristol N2), adr-1(uu49);adr-2(uu28), rrf-3(uu56), adr-1(uu49);adr-2(uu28);rrf-3(uu56), adr-1(uu49);adr-2(uu28);rrf-3(uu56);rde-1(uu51), and adr-1(uu49);adr-2(uu28);rrf-3(uu56);rde-4(uu53).