title: A conserved three-nucleotide core motif defines Musashi RNA binding specificity.
abstract: Musashi (MSI) family proteins control cell proliferation and differentiation in many biological systems. They are overexpressed in tumors of several origins, and their expression level correlates with poor prognosis. MSI proteins control gene expression by binding RNA and regulating its translation. They contain two RNA recognition motif (RRM) domains, which recognize a defined sequence element. The relative contribution of each nucleotide to the binding affinity and specificity is unknown. We analyzed the binding specificity of three MSI family RRM domains using a quantitative fluorescence anisotropy assay. We found that the core element driving recognition is the sequence UAG. Nucleotides outside of this motif have a limited contribution to binding free energy. For mouse MSI1, recognition is determined by the first of the two RRM domains. The second RRM adds affinity but does not contribute to binding specificity. In contrast, the recognition element for Drosophila MSI is more extensive than the mouse homolog, suggesting functional divergence. The short nature of the binding determinant suggests that protein-RNA affinity alone is insufficient to drive target selection by MSI family proteins.
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: N Ruth
authors.initials: NR
authors.lastname: Zearfoss
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Laura M
authors.initials: LM
authors.lastname: Deveau
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Carina C
authors.initials: CC
authors.lastname: Clingman
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Eric
authors.initials: E
authors.lastname: Schmidt
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Emily S
authors.initials: ES
authors.lastname: Johnson
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605.
authors.forename: Francesca
authors.initials: F
authors.lastname: Massi
authors.affiliation: From the Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605 Sean.Ryder@umassmed.edu.
authors.forename: Sean P
authors.initials: SP
authors.lastname: Ryder
mesh_terms: D000465:Algorithms; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D017124:Conserved Sequence / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D005454:Fluorescence Polarization; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D059372:Nucleotide Motifs / Q000235:genetics*; D011485:Protein Binding; D012313:RNA / Q000235:genetics* / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid
chemical_list: D029721:Drosophila Proteins; C438151:MSI2 protein, human; C104786:Msi1h protein, mouse; C499059:Msi2h protein, mouse; D009419:Nerve Tissue Proteins; D016601:RNA-Binding Proteins; C088623:msi protein, Drosophila; D012313:RNA
keywords: Binding Specificity; Cell Proliferation; Fluorescence Anisotropy; RNA; RNA Recognition; RNA-Protein Interaction; RNA-binding Protein; RRM