This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.MicroRNAs (miRNAs) regulate gene expression for diverse functions, but only a limited number of mRNA targets have been experimentally identified. We show that GW182 family proteins AIN-1 and AIN-2 act redundantly to regulate the expression of miRNA targets, but not miRNA biogenesis. Immunoprecipitation (IP) and mass spectrometry indicate that AIN-1 and AIN-2 interact only with miRNA-specific Argonaute proteins ALG-1 and ALG-2 and with components of the core translational initiation complex. Known miRNA targets are enriched in AIN-2 complexes, correlating with the expression of corresponding miRNAs. Combining IP with pyrosequencing and microarray analysis of RNAs associated with AIN-1/AIN-2, we identified 106 previously annotated miRNAs plus nine new candidate miRNAs, but nearly no siRNAs, and more than 3500 potential miRNA targets, including nearly all known ones. Our results demonstrate an effective biochemical approach to systematically identify miRNA targets and provide valuable insights regarding the properties of miRNA effector complexes.
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