Elucidation of a C-rich signature motif in target mRNAs of RNA-binding protein TIAR

The RNA-binding protein TIAR [related to TIA(T-cell intracellular antigen)-1] was shown to associate with subsets of mRNAs bearing U-rich sequences in their 3 -untranslated regions (UTRs). TIAR can function as a translational repressor, particularly in response to cytotoxic agents. Using unstressed colon cancer cells, collections of mRNAs associated with TIAR were isolated by immunoprecipitation (IP) of [TIAR-RNA] ribonucleoprotein (RNP) complexes, identified by microarray analysis, and used to elucidate a common signature motif present among TIAR target transcripts. The predicted TIAR motif was an unexpectedly cytosine-rich, 28- to 32-nucleotide long element forming a stem and a loop of variable size with an additional side loop. The ability of TIAR to bind an RNA oligonucleotide with a representative C-rich TIAR motif sequence was verified in vitro using surface plasmon resonance. By this analysis, TIAR containing 2 or 3 RNA recognition domains (TIAR12, TIAR123) showed low but significant binding to the C-rich sequence. In vivo, insertion of the C-rich motif into a heterologous reporter strongly suppressed its translation in cultured cells. Using this signature motif, an additional approximately 2,500 UniGene targets were identified (3.4 of the total UniGene database). A subset of specific mRNAs were validated by RNP IP analysis. Interestingly, in response to treatment with short-wavelength ultraviolet light (UVC), a stress agent causing DNA damage, each of these target mRNAs bearing C-rich motifs dissociated from TIAR. In turn, expression of the encoded proteins was elevated in a TIAR-dependent manner. In sum, we report the identification of a C-rich signature motif present in TIAR target mRNAs whose association with TIAR decreases following exposure to a stress-causing agent.