In a screen for C. elegans systemic RNAi deficient mutants (Winston et al., 2001), two alleles of
sid-5 were identified. The mutants were isolated as defective in silencing GFP expressed in body wall muscles when fed bacteria expressing GFP dsRNA.
sid-5 worms injected into either the gonad or intestine with
mex-3 or
unc-22 dsRNA produce affected progeny, indicating that
sid-5 is not strictly required for general or systemic RNAi. Therefore, we explored the requirement for
sid-5 in environmental RNAi, where the dsRNA is introduced via feeding or soaking.
sid-5 worms fed bacteria expressing dsRNA that corresponds to endogenous targets (
unc-22,
unc-54, or
pal-1 ) are less affected than wild-type controls but the longer the worms are exposed to the food, the stronger the silencing becomes. In contrast,
sid-5 worms soaked in dsRNA (
unc-22 or
pal-1 ) display a strong RNAi effect which, relative to wild-type controls, diminishes over time. These results indicate that
sid-5 is needed for robust environmental RNAi. The
sid-5 gene was mapped to a small interval on linkage group X and rescued by injection of a genomic fragment that encompassed two genes, one of which was mutated in both
sid-5 alleles.
sid-5 is predicted to encode a 67 amino acid protein with a single transmembrane domain and no signal sequence. Preliminary analysis of a full-length SID-5::GFP fusion indicates that SID-5 is expressed in the excretory cell, spermatheca, and rectum. Current experiments are focused on refining the RNAi phenotype and SID-5 expression pattern more fully. Winston, W. M., C. M., Molodowitch, C. P. Hunter. 2002. Systemic RNAi in C. elegans requires the putative transmembrane domain protein SID-1. Science. 295:2456-59.