title: The Lin28 cold-shock domain remodels pre-let-7 microRNA.
abstract: The RNA-binding protein Lin28 regulates the processing of a developmentally important group of microRNAs, the let-7 family. Lin28 blocks the biogenesis of let-7 in embryonic stem cells and thereby prevents differentiation. It was shown that both RNA-binding domains (RBDs) of this protein, the cold-shock domain (CSD) and the zinc-knuckle domain (ZKD) are indispensable for pri- or pre-let-7 binding and blocking its maturation. Here, we systematically examined the nucleic acid-binding preferences of the Lin28 RBDs and determined the crystal structure of the Lin28 CSD in the absence and presence of nucleic acids. Both RNA-binding domains bind to single-stranded nucleic acids with the ZKD mediating specific binding to a conserved GGAG motif and the CSD showing only limited sequence specificity. However, only the isolated Lin28 CSD, but not the ZKD, can bind with a reasonable affinity to pre-let-7 and thus is able to remodel the terminal loop of pre-let-7 including the Dicer cleavage site. Further mutagenesis studies reveal that the Lin28 CSD induces a conformational change in the terminal loop of pre-let-7 and thereby facilitates a subsequent specific binding of the Lin28 ZKD to the conserved GGAG motif.
authors.affiliation: Crystallography, Max-Delbrück Center for Molecular Medicine, Robert-Rössle Straße 10, 13125 Berlin, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Mayr
authors.forename: Anja
authors.initials: A
authors.lastname: Schütz
authors.forename: Nadine
authors.initials: N
authors.lastname: Döge
authors.forename: Udo
authors.initials: U
authors.lastname: Heinemann
mesh_terms: D000818:Animals; D001665:Binding Sites; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D035683:MicroRNAs / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D009711:Nucleotides / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D012322:RNA Precursors / Q000737:chemistry* / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D014981:Xenopus
chemical_list: D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; C516326:LIN28B protein, human; D035683:MicroRNAs; D009711:Nucleotides; D012322:RNA Precursors; D016601:RNA-Binding Proteins; C576137:let-7 microRNA, Xenopus; D012313:RNA