Novel DNA-binding properties of the RNA-binding protein TIAR.
TIA-1 related protein binds avidly to uridine-rich elements in mRNA and pre-mRNAs of a wide range of genes, including interleukin (IL)-8 and vascular endothelial growth factor (VEGF). The protein has diverse regulatory roles, which in part depend on the locus of binding within the transcript, including translational control, splicing and apoptosis. Here, we observed selective and potent inhibition of TIAR-RNP complex formation with IL-8 and VEGF 3'-untranslated regions (3'-UTRs) using thymidine-rich deoxyoligonucleotide (ODN) sequences derived from the VEFG 3'-UTR. We show by ultraviolet crosslinking and electrophoretic mobility shift assays that TIAR can bind directly to single-stranded, thymidine-rich ODNs but not to double-stranded ODNs containing the same sequence. TIAR had a nearly 6-fold greater affinity for DNA than RNA (K(d)app = 1.6x10(-9) M versus 9.4 x 10(-9) M). Truncation of TIAR indicated that the high affinity DNA-binding site overlaps with the RNA-binding site involving RNA recognition motif 2 (RRM2). However, RRM1 alone could also bind to DNA. Finally, we show that TIAR can be displaced from single-stranded DNA by active transcription through the binding site. These results provide a potential mechanism by which TIAR can shuttle between RNA and DNA ligands.
Suswam
Esther A
EA
Department of Neurology, University of Alabama Birmingham, AL 35295, USA.
Li
Yan Yan
YY
Mahtani
Harry
H
King
Peter H
PH
D020413:3' Untranslated Regions / Q000378:metabolism; D001665:Binding Sites; D045744:Cell Line, Tumor; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D016209:Interleukin-8 / Q000235:genetics; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D013936:Thymidine / Q000032:analysis; D014158:Transcription, Genetic; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics
D020413:3' Untranslated Regions; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D016209:Interleukin-8; D020319:Oligodeoxyribonucleotides, Antisense; D016601:RNA-Binding Proteins; D042461:Vascular Endothelial Growth Factor A; C076751:TIAL1 protein, human; D013936:Thymidine