title: Structural analysis of the yeast Dhh1-Pat1 complex reveals how Dhh1 engages Pat1, Edc3 and RNA in mutually exclusive interactions.
abstract: Translational repression and deadenylation of eukaryotic mRNAs result either in the sequestration of the transcripts in a nontranslatable pool or in their degradation. Removal of the 5' cap structure is a crucial step that commits deadenylated mRNAs to 5'-to-3' degradation. Pat1, Edc3 and the DEAD-box protein Dhh1 are evolutionary conserved factors known to participate in both translational repression and decapping, but their interplay is currently unclear. We report the 2.8 Å resolution structure of yeast Dhh1 bound to the N-terminal domain of Pat1. The structure shows how Pat1 wraps around the C-terminal RecA domain of Dhh1, docking onto the Phe-Asp-Phe (FDF) binding site. The FDF-binding site of Dhh1 also recognizes Edc3, revealing why the binding of Pat1 and Edc3 on Dhh1 are mutually exclusive events. Using co-immunoprecipitation assays and structure-based mutants, we demonstrate that the mode of Dhh1-Pat1 recognition is conserved in humans. Pat1 and Edc3 also interfere and compete with the RNA-binding properties of Dhh1. Mapping the RNA-binding sites on Dhh1 with a crosslinking-mass spectrometry approach shows a large RNA-binding surface around the C-terminal RecA domain, including the FDF-binding pocket. The results suggest a model for how Dhh1-containing messenger ribonucleoprotein particles might be remodeled upon Pat1 and Edc3 binding.
authors.affiliation: Structural Cell Biology Department, Max Planck Institute of Biochemistry, Am Klopferspitz 18, Martinsried/Munich, D-82152 Germany and Cellular Biochemistry Department, Max Planck Institute of Biophysical Chemistry, Am Faßberg 11, 37077 Göttingen, Germany.
authors.forename: Humayun
authors.initials: H
authors.lastname: Sharif
authors.forename: Sevim
authors.initials: S
authors.lastname: Ozgur
authors.forename: Kundan
authors.initials: K
authors.lastname: Sharma
authors.forename: Claire
authors.initials: C
authors.lastname: Basquin
authors.forename: Henning
authors.initials: H
authors.lastname: Urlaub
authors.forename: Elena
authors.initials: E
authors.lastname: Conti
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D053487:DEAD-box RNA Helicases / Q000737:chemistry* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D012313:RNA / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000378:metabolism; D016415:Sequence Alignment
chemical_list: C488957:EDC3 protein, S cerevisiae; C103266:PAT1 protein, S cerevisiae; D016601:RNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D012313:RNA; C445083:DHH1 protein, S cerevisiae; D053487:DEAD-box RNA Helicases