TY - JOUR
T1 - The Caenorhabditis elegans RDE-10/RDE-11 complex regulates RNAi by promoting secondary siRNA amplification
AU - Zhang, Chi
AU - Montgomery, Taiowa A.
AU - Fischer, Sylvia E.J.
AU - Garcia, Susana M.D.A.
AU - Riedel, Christian G.
AU - Fahlgren, Noah
AU - Sullivan, Christopher M.
AU - Carrington, James C.
AU - Ruvkun, Gary
N1 - Funding Information:
We are very grateful to Huan Yang and Ho Yi Mak for rde-10 and rde-11 mutant and transgenic strains, as well as communicating unpublished data. We thank Darryl Conte, Weifeng Gu, Pedro Batista, and Daniel Chaves for providing protocols for immunoprecipitation and generation of small RNA cDNA libraries for deep sequencing; Xiaoyun Wu for protocols for large-scale immunoprecipitation; John Asara for mass spectrometry analysis; Christian Daly, Jiangwen Zhang, Mark Borowsky, Tammy Gillis, and Kaleena Shirley for Illumina deep sequencing and computational assistance; and the Caenorhabditis Genetics Center and Shohei Mitani for strains. This work was supported by a grant to G.R. from National Institutes of Health (NIH) (GM44619), grants to J.C.C. from the National Science Foundation (MCB-1231726) and NIH (AI43288), fellowships to C.Z. and S.E.J.F. from the MGH ECOR Fund for Medical Discovery, fellowship to T.A.M. from the Damon Runyon Cancer Research Foundation (DRG-2029-09), fellowship to S.M.D.A.G. from the American Heart Association (0825938D), and fellowships to C.G.R. from the European Molecular Biology Organization and the Human Frontier Science Program.
PY - 2012/5/22
Y1 - 2012/5/22
N2 - Background: In nematodes, plants, and fungi, RNAi is remarkably potent and persistent due to the amplification of initial silencing signals by RNA-dependent RNA polymerases (RdRPs). In Caenorhabditis elegans (C. elegans), the interaction between the RNA-induced silencing complex (RISC) loaded with primary small interfering RNAs (siRNAs) and the target messenger RNA (mRNA) leads to the recruitment of RdRPs and synthesis of secondary siRNAs using the target mRNA as the template. The mechanism and genetic requirements for secondary siRNA accumulation are not well understood. Results: From a forward genetic screen for C. elegans genes required for RNAi, we identified rde-10, and through proteomic analysis of RDE-10-interacting proteins, we identified a protein complex containing the new RNAi factor RDE-11, the known RNAi factors RSD-2 and ERGO-1, and other candidate RNAi factors. The RNAi defective genes rde-10 and rde-11 encode a novel protein and a RING-type zinc finger domain protein, respectively. Mutations in rde-10 and rde-11 genes cause dosage-sensitive RNAi deficiencies: these mutants are resistant to low dosage but sensitive to high dosage of double-stranded RNAs. We assessed the roles of rde-10, rde-11, and other dosage-sensitive RNAi-defective genes rsd-2, rsd-6, and haf-6 in both exogenous and endogenous small RNA pathways using high-throughput sequencing and qRT-PCR. These genes are required for the accumulation of secondary siRNAs in both exogenous and endogenous RNAi pathways. Conclusions: The RDE-10/RDE-11 complex is essential for the amplification of RNAi in C. elegans by promoting secondary siRNA accumulation.
AB - Background: In nematodes, plants, and fungi, RNAi is remarkably potent and persistent due to the amplification of initial silencing signals by RNA-dependent RNA polymerases (RdRPs). In Caenorhabditis elegans (C. elegans), the interaction between the RNA-induced silencing complex (RISC) loaded with primary small interfering RNAs (siRNAs) and the target messenger RNA (mRNA) leads to the recruitment of RdRPs and synthesis of secondary siRNAs using the target mRNA as the template. The mechanism and genetic requirements for secondary siRNA accumulation are not well understood. Results: From a forward genetic screen for C. elegans genes required for RNAi, we identified rde-10, and through proteomic analysis of RDE-10-interacting proteins, we identified a protein complex containing the new RNAi factor RDE-11, the known RNAi factors RSD-2 and ERGO-1, and other candidate RNAi factors. The RNAi defective genes rde-10 and rde-11 encode a novel protein and a RING-type zinc finger domain protein, respectively. Mutations in rde-10 and rde-11 genes cause dosage-sensitive RNAi deficiencies: these mutants are resistant to low dosage but sensitive to high dosage of double-stranded RNAs. We assessed the roles of rde-10, rde-11, and other dosage-sensitive RNAi-defective genes rsd-2, rsd-6, and haf-6 in both exogenous and endogenous small RNA pathways using high-throughput sequencing and qRT-PCR. These genes are required for the accumulation of secondary siRNAs in both exogenous and endogenous RNAi pathways. Conclusions: The RDE-10/RDE-11 complex is essential for the amplification of RNAi in C. elegans by promoting secondary siRNA accumulation.
UR - https://www.scopus.com/pages/publications/84861532359
U2 - 10.1016/j.cub.2012.04.011
DO - 10.1016/j.cub.2012.04.011
M3 - Article
C2 - 22542102
AN - SCOPUS:84861532359
SN - 0960-9822
VL - 22
SP - 881
EP - 890
JO - Current Biology
JF - Current Biology
IS - 10
ER -