Gesnel Marie-Claude, Del Gatto-Konczak Fabienne, Breathnach Richard
INSERM, U892, 8 quai Moncousu, BP 70721, 44007 Nantes Cedex 1, France.
J Biomed Biotechnol. 2009;2009:104853. doi: 10.1155/2009/104853. Epub 2010 Jan 14.
Splicing of the FGFR2 K-SAM exon is repressed by hnRNP A1 bound to the exon and activated by TIA-1 bound to the downstream intron. Both proteins are expressed similarly by cells whether they splice the exon or not, so it is important to know which one is dominant. To answer this question, we used bacteriophage PP7 and bacteriophage MS2 coat fusions to tether hnRNP A1 and TIA-1 to distinct sites on the same pre-mRNA molecule. hnRNP A1 fused to one coat protein was tethered to a K-SAM exon containing the corresponding coat protein's binding site. TIA-1 fused to the other coat protein was tethered to the downstream intron containing that coat protein's binding site. This led to efficient K-SAM exon splicing. Our results show that TIA-1 is dominant for K-SAM exon splicing control and validate the combined use of PP7 and MS2 coat proteins for studying posttranscriptional events.
与该外显子结合的hnRNP A1会抑制FGFR2 K-SAM外显子的剪接，而与下游内含子结合的TIA-1则会激活它。无论细胞是否剪接该外显子，这两种蛋白的表达都相似，因此了解哪种蛋白起主导作用很重要。为了回答这个问题，我们使用噬菌体PP7和噬菌体MS2外壳融合蛋白，将hnRNP A1和TIA-1拴系到同一前体mRNA分子的不同位点上。与一种外壳蛋白融合的hnRNP A1被拴系到一个含有相应外壳蛋白结合位点的K-SAM外显子上。与另一种外壳蛋白融合的TIA-1被拴系到含有该外壳蛋白结合位点的下游内含子上。这导致了K-SAM外显子的高效剪接。我们的结果表明，TIA-1在K-SAM外显子剪接控制中起主导作用，并验证了PP7和MS2外壳蛋白在研究转录后事件中的联合应用。
