A comprehensive RNAi-based screen for the DExD/H-box RNA helices involved in C. elegans microRNA function
Yude Chu, Shih-Peng Chan
Graduate Institute of Microbiology, College of Medicine, National Taiwan University, Taiwan
In eukaryotes, RNA helicases generally belong to the superfamily 2 (SF2) in helicase classification, especially the DExD/H-box helicase family. The DExD/H-box proteins have been shown in association with almost all processes of RNA metabolism, including transcription, pre-mRNA splicing, rRNA processing, translation regulation and RNA degradation. However, little is known about their roles in microRNA function. To better understand the involvement of the DExD/H-box proteins with miRNAs, we employed RNAi to reduce the expression of 48 reported DEAD-box RNA helicases in C. elegans and examined the impacts on the function of the let-7 miRNA. In C. elegans, the let-7 miRNA controls the timing of cell cycle exit and terminal differentiation. The temperature-sensitive allele let-7(n2853) causes heterochronic phenotypes, reiterated lateral seam cell division and lethal vulval bursting at the larva-to-adult switch. Reverse genetics screen for suppression of let-7(n2853) phenotypes has been broadly used to detect let-7-interacting factors. In addition, a weak allele let-7(mg279), without severe vulval bursting phenotype, could be used as a sensitized background in which to screen for synthetic lethality and also search for effectors in the let-7 miRNA pathway. Here, we report a comprehensive reverse genetics screen for the DExD/H-box proteins that may be involved in the let-7 miRNA pathway. We found reduced expression of several RNA helicases functioning in ribosomal RNA processing and pre-mRNA splicing altered let-7 mutant phenotypes and/or the expression of let-7, suggesting they may also play roles in the miRNA pathway.