title: Structural features of Argonaute-GW182 protein interactions.
abstract: MicroRNAs (miRNAs) guide Argonaute (Ago) proteins to target mRNAs, leading to gene silencing. However, Ago proteins are not the actual mediators of gene silencing but interact with a member of the GW182 protein family (also known as GW proteins), which coordinates all downstream steps in gene silencing. GW proteins contain an N-terminal Ago-binding domain that is characterized by multiple GW repeats and a C-terminal silencing domain with several globular domains. Within the Ago-binding domain, Trp residues mediate the direct interaction with the Ago protein. Here, we have characterized the interaction of Ago proteins with GW proteins in molecular detail. Using biochemical and NMR experiments, we show that only a subset of Trp residues engage in Ago interactions. The Trp residues are located in intrinsically disordered regions, where flanking residues mediate additional weak interactions, that might explain the importance of specific tryptophans. Using cross-linking followed by mass spectrometry, we map the GW protein interactions with Ago2, which allows for structural modeling of Ago-GW182 interaction. Our data further indicate that the Ago-GW protein interaction might be a two-step process involving the sequential binding of two tryptophans separated by a spacer with a minimal length of 10 aa.
authors.affiliation: Biochemistry Center Regensburg, Laboratory for RNA Biology, University of Regensburg, 93053 Regensburg, Germany.
authors.forename: Janina
authors.initials: J
authors.lastname: Pfaff
authors.forename: Janosch
authors.initials: J
authors.lastname: Hennig
authors.forename: Franz
authors.initials: F
authors.lastname: Herzog
authors.forename: Ruedi
authors.initials: R
authors.lastname: Aebersold
authors.forename: Michael
authors.initials: M
authors.lastname: Sattler
authors.forename: Dierk
authors.initials: D
authors.lastname: Niessing
authors.forename: Gunter
authors.initials: G
authors.lastname: Meister
mesh_terms: D060565:Argonaute Proteins / Q000737:chemistry* / Q000378:metabolism; D001324:Autoantigens / Q000737:chemistry* / Q000378:metabolism; D016367:Baculoviridae; D002942:Circular Dichroism; D005454:Fluorescence Polarization; D005786:Gene Expression Regulation / Q000235:genetics*; D005822:Genetic Vectors; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D034622:RNA Interference; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism
chemical_list: C466422:AGO2 protein, human; D060565:Argonaute Proteins; D001324:Autoantigens; D046912:Multiprotein Complexes; D016601:RNA-Binding Proteins; C465663:TNRC6A protein, human
keywords: RNA interference; RNAi; gene regulation; small RNA–mediated gene silencing
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