TIA-1 RRM23 binding and recognition of target oligonucleotides

• Published in: Nucleic acids research 2017-05, Vol.45 (8), p.4944-4957
• Main Authors: Waris, Saboora, García-Mauriño, Sofía M, Sivakumaran, Andrew, Beckham, Simone A, Loughlin, Fionna E, Gorospe, Myriam, Díaz-Moreno, Irene, Wilce, Matthew C J, Wilce, Jacqueline A
• Format: Article
• Language: English
• Subjects: Crystallography, X-Ray; DNA - chemistry; DNA - genetics; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; Gene Expression Regulation; Humans; Oligonucleotides - chemistry; Poly(A)-Binding Proteins - chemistry; Poly(A)-Binding Proteins - genetics; Protein Binding - genetics; Protein Interaction Maps - genetics; Ribonucleoside Diphosphate Reductase - chemistry; Ribonucleoside Diphosphate Reductase - genetics; RNA Recognition Motif - genetics; Structural Biology; T-Cell Intracellular Antigen-1
• ISSN: 0305-1048; 1362-4962
• DOI: 10.1093/nar/gkx102

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.