title: GW182-Free microRNA Silencing Complex Controls Post-transcriptional Gene Expression during Caenorhabditis elegans Embryogenesis.
abstract: MicroRNAs and Argonaute form the microRNA induced silencing complex or miRISC that recruits GW182, causing mRNA degradation and/or translational repression. Despite the clear conservation and molecular significance, it is unknown if miRISC-GW182 interaction is essential for gene silencing during animal development. Using Caenorhabditis elegans to explore this question, we examined the relationship and effect on gene silencing between the GW182 orthologs, AIN-1 and AIN-2, and the microRNA-specific Argonaute, ALG-1. Homology modeling based on human Argonaute structures indicated that ALG-1 possesses conserved Tryptophan-binding Pockets required for GW182 binding. We show in vitro and in vivo that their mutations severely altered the association with AIN-1 and AIN-2. ALG-1 tryptophan-binding pockets mutant animals retained microRNA-binding and processing ability, but were deficient in reporter silencing activity. Interestingly, the ALG-1 tryptophan-binding pockets mutant phenocopied the loss of alg-1 in worms during larval stages, yet was sufficient to rescue embryonic lethality, indicating the dispensability of AINs association with the miRISC at this developmental stage. The dispensability of AINs in miRNA regulation is further demonstrated by the capacity of ALG-1 tryptophan-binding pockets mutant to regulate a target of the embryonic mir-35 microRNA family. Thus, our results demonstrate that the microRNA pathway can act independently of GW182 proteins during C. elegans embryogenesis.
authors.affiliation: St-Patrick Research Group in Basic Oncology, CHU de Québec-Université Laval Research Centre (Hôtel-Dieu de Québec), Quebec City, Québec, Canada.|Laval University Cancer Research Centre, Quebec City, Québec, Canada.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Jannot
authors.affiliation: St-Patrick Research Group in Basic Oncology, CHU de Québec-Université Laval Research Centre (Hôtel-Dieu de Québec), Quebec City, Québec, Canada.|Laval University Cancer Research Centre, Quebec City, Québec, Canada.
authors.forename: Pascale
authors.initials: P
authors.lastname: Michaud
authors.affiliation: St-Patrick Research Group in Basic Oncology, CHU de Québec-Université Laval Research Centre (Hôtel-Dieu de Québec), Quebec City, Québec, Canada.|Laval University Cancer Research Centre, Quebec City, Québec, Canada.
authors.forename: Miguel
authors.initials: M
authors.lastname: Quévillon Huberdeau
authors.affiliation: St-Patrick Research Group in Basic Oncology, CHU de Québec-Université Laval Research Centre (Hôtel-Dieu de Québec), Quebec City, Québec, Canada.|Laval University Cancer Research Centre, Quebec City, Québec, Canada.
authors.forename: Louis
authors.initials: L
authors.lastname: Morel-Berryman
authors.affiliation: Department of Chemistry & Biochemistry, The Ohio State University, Columbus, Ohio, United States of America.
authors.forename: James A
authors.initials: JA
authors.lastname: Brackbill
authors.affiliation: St-Patrick Research Group in Basic Oncology, CHU de Québec-Université Laval Research Centre (Hôtel-Dieu de Québec), Quebec City, Québec, Canada.|Laval University Cancer Research Centre, Quebec City, Québec, Canada.
authors.forename: Sandra
authors.identifier: http://orcid.org/0000-0002-5745-9862
authors.initials: S
authors.lastname: Piquet
authors.affiliation: Program in Molecular Medicine, RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, Massachusetts, United States of America.
authors.forename: Katherine
authors.initials: K
authors.lastname: McJunkin
authors.affiliation: Department of Chemistry & Biochemistry, The Ohio State University, Columbus, Ohio, United States of America.
authors.forename: Kotaro
authors.initials: K
authors.lastname: Nakanishi
authors.affiliation: St-Patrick Research Group in Basic Oncology, CHU de Québec-Université Laval Research Centre (Hôtel-Dieu de Québec), Quebec City, Québec, Canada.|Laval University Cancer Research Centre, Quebec City, Québec, Canada.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Simard
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics / Q000254:growth & development; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000235:genetics*; D002352:Carrier Proteins / Q000235:genetics*; D047108:Embryonic Development / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D035683:MicroRNAs / Q000096:biosynthesis / Q000235:genetics*; D009154:Mutation; D034622:RNA Interference; D016601:RNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics*; D034743:RNA-Induced Silencing Complex / Q000235:genetics
chemical_list: C504319:AIN-1 protein, C elegans; C525939:AIN-2 protein, C elegans; C558782:ALG-1 protein, C elegans; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; C000618258:GW182 protein, C elegans; C570417:MIRN35 microRNA, C elegans; D035683:MicroRNAs; D016601:RNA-Binding Proteins; D034743:RNA-Induced Silencing Complex