Mutant alleles of the Caenorhabditis elegans rde-1 gene identified through chemical mutagenesis of an snRNA misprocessing reporter.

The processing of small nuclear RNA post transcription involves endolytic cleavage of a 3' tail to produce a mature transcript that is incorporated into the spliceosome to regulate RNA splicing. We previously reported in the Caenorhabditis elegans model several novel genetic regulators including those functioning in RNAi processing to be required for snRNA cleavage through a genome-wide RNAi screen using an in vivo snRNA misprocessing reporter. Here, we conducted a forward genetic screen using the mutagen ethyl methanesulfonate to screen for viable mutants that exhibit constitutive snRNA misprocessing. This screen generated three new recessive rde-1 mutant alleles (cww1, cww4, cww9) identified via WGS SNP mapping, which encode the primary Argonaute protein involved in the processing of exogenous RNAi. The three rde-1 alleles failed to complement each other and rde-1(cww1) which contains a premature stop codon in exon 3 also failed to be complemented by the classic rde-1(ne219) allele. We show that the three rde-1 mutants display a varying degree of snRNA misprocessing reporter activation, but are all fully resistant to various RNAi that are known to cause larval arrest or an abnormal vulva phenotype. Thus, the screen has reinforced a connection between RNAi processing and snRNA cleavage and generated mutants that are useful for future studies of the rde-1 Argonaute gene.