title: TIA-1 RRM23 binding and recognition of target oligonucleotides.
abstract: TIA-1 (T-cell restricted intracellular antigen-1) is an RNA-binding protein involved in splicing and translational repression. It mainly interacts with RNA via its second and third RNA recognition motifs (RRMs), with specificity for U-rich sequences directed by RRM2. It has recently been shown that RRM3 also contributes to binding, with preferential binding for C-rich sequences. Here we designed UC-rich and CU-rich 10-nt sequences for engagement of both RRM2 and RRM3 and demonstrated that the TIA-1 RRM23 construct preferentially binds the UC-rich RNA ligand (5΄-UUUUUACUCC-3΄). Interestingly, this binding depends on the presence of Lys274 that is C-terminal to RRM3 and binding to equivalent DNA sequences occurs with similar affinity. Small-angle X-ray scattering was used to demonstrate that, upon complex formation with target RNA or DNA, TIA-1 RRM23 adopts a compact structure, showing that both RRMs engage with the target 10-nt sequences to form the complex. We also report the crystal structure of TIA-1 RRM2 in complex with DNA to 2.3 Å resolution providing the first atomic resolution structure of any TIA protein RRM in complex with oligonucleotide. Together our data support a specific mode of TIA-1 RRM23 interaction with target oligonucleotides consistent with the role of TIA-1 in binding RNA to regulate gene expression.
authors.affiliation: Monash Biomedicine Discovery Institute, Department of Biochemistry & Molecular Biology, Monash University, Victoria 3800, Australia.
authors.forename: Saboora
authors.initials: S
authors.lastname: Waris
authors.affiliation: Instituto de Investigaciones Químicas (IIQ)-Centro de Investigaciones Científicas Isla de la Cartuja (cicCartuja), Universidad de Sevilla-Consejo Superior de Investigaciones Científicas (CSIC), Sevilla 41092, Spain.
authors.forename: Sofía M
authors.initials: SM
authors.lastname: García-Mauriño
authors.affiliation: Monash Biomedicine Discovery Institute, Department of Biochemistry & Molecular Biology, Monash University, Victoria 3800, Australia.
authors.forename: Andrew
authors.initials: A
authors.lastname: Sivakumaran
authors.affiliation: Monash Biomedicine Discovery Institute, Department of Biochemistry & Molecular Biology, Monash University, Victoria 3800, Australia.
authors.forename: Simone A
authors.initials: SA
authors.lastname: Beckham
authors.affiliation: Monash Biomedicine Discovery Institute, Department of Biochemistry & Molecular Biology, Monash University, Victoria 3800, Australia.
authors.forename: Fionna E
authors.initials: FE
authors.lastname: Loughlin
authors.affiliation: Laboratory of Genetics and Genomics, National Institute on Aging-Intramural Research Program, National Institutes of Health, Baltimore, MD 21224, USA.
authors.forename: Myriam
authors.initials: M
authors.lastname: Gorospe
authors.affiliation: Instituto de Investigaciones Químicas (IIQ)-Centro de Investigaciones Científicas Isla de la Cartuja (cicCartuja), Universidad de Sevilla-Consejo Superior de Investigaciones Científicas (CSIC), Sevilla 41092, Spain.
authors.forename: Irene
authors.initials: I
authors.lastname: Díaz-Moreno
authors.affiliation: Monash Biomedicine Discovery Institute, Department of Biochemistry & Molecular Biology, Monash University, Victoria 3800, Australia.
authors.forename: Matthew C J
authors.initials: MCJ
authors.lastname: Wilce
authors.affiliation: Monash Biomedicine Discovery Institute, Department of Biochemistry & Molecular Biology, Monash University, Victoria 3800, Australia.
authors.forename: Jacqueline A
authors.initials: JA
authors.lastname: Wilce
mesh_terms: D018360:Crystallography, X-Ray; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D005786:Gene Expression Regulation; D006801:Humans; D009841:Oligonucleotides / Q000737:chemistry; D039102:Poly(A)-Binding Proteins / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding / Q000235:genetics; D060066:Protein Interaction Maps / Q000235:genetics; D000071377:RNA Recognition Motif / Q000235:genetics; D012262:Ribonucleoside Diphosphate Reductase / Q000737:chemistry* / Q000235:genetics; D000074019:T-Cell Intracellular Antigen-1
chemical_list: D004268:DNA-Binding Proteins; D009841:Oligonucleotides; D039102:Poly(A)-Binding Proteins; D000074019:T-Cell Intracellular Antigen-1; C071558:TIA1 protein, human; D004247:DNA; C093713:ribonucleotide reductase M2; D012262:Ribonucleoside Diphosphate Reductase