拟南芥中PAF1c调控mRNA 3’末端形成的机制研究

Gene expression in eukaryotes is tightly controlled through regulation at both the transcriptional and post-transcriptional levels. RNA processing, which includes the 5' capping, splicing, and 3' end formation of pre-mRNA molecules, is a key step in the post-transcriptional regulation of gene expression. PAF1c is an evolutionarily conserved complex that has been proposed to be associated with RNA polymerase II during transcriptional elongation and termination in yeast and animal cells. In plants, PAF1c is reportedly involved in the regulation of flowering time by promoting the histone modification of chromatin and the transcription of FLC/MAFs in Arabidopsis. However, whether PAF1c is involved in the regulation of gene expression at the post-transcriptional level is unknown. We previously showed that PAF1c regulates 3' end formation for several genes. The proposed work will examine the role of PAF1c in the regulation of 3' end formation at the whole-genome scale, and reveal how PAF1c regulates 3' end formation during transcription termination in Arabidopsis. We will also examine whether PAF1c mediates a specific developmental process by regulating the 3' end formation of genes involved in that process. The results of the proposed work will not only uncover a new mechanism whereby PAF1c regulates gene expression at the post-transcriptional level, they will also provide new insight into the regulatory mechanism of mRNA 3' end formation in plants.

真核生物基因表达在转录及转录后水平受到严格调控，从而精确控制mRNA及蛋白的水平来调节机体发育。RNA加工是基因转录后调控的关键步骤，包括对初始转录产物5’加帽，剪接和3’末端形成。PAF1c在进化过程中保守，与RNA聚合酶II一起负责基因转录延伸和终止。植物体系中已有结果表明PAF1c通过影响组蛋白修饰调控FLC转录进而调节拟南芥开花。但PAF1c是否参与转录后基因表达调控还未见报道。我们前期结果检测到拟南芥PAF1c可影响一些基因的3’末端形成，暗示PAF1c可能参与植物前体mRNA 3’末端形成。本项目拟进一步检测拟南芥PAF1c对基因组范围内mRNA 3’末端形成影响，探讨其调控基因3’末端形成的分子机制，并证实PAF1c是否能够通过调控基因的3’末端形成影响植物某个或某些发育过程。研究结果将完善PAF1c调控植物基因表达的分子机制，还将丰富前体mRNA 3’末端形成的分子机制。

PAF1c在进化上保守，酵母和动物中PAF1c与RNA聚合酶II一同促进基因转录的延伸和终止。本项目对拟南芥中PAF1c调控基因3’末端形成的作用及分子机制进行了探讨。.明确了拟南芥中PAF1c在基因组范围内调控mRNA 3’末端形成中的作用。PAF1c促进基因3’末端polyA位点的使用并促进全长转录本的形成，抑制基因内部polyA位点的使用从而抑制截短形式RNA的产生；PAF1c还可以抑制基因3’末端通读形式RNA的产生。.揭示了PAF1c调控基因3’末端形成的机制，PAF1c可与植物特异的RNA 结合蛋白FCA和FPA相互作用，并通过FCA（或FPA）与保守的3’polyA因子FY（或PA4）在一个大的复合体中，调控特定基因的3’末端形成；PAF1c还可以与PA4直接相互作用。由此，PAF1c将基因的转录和RNA加工过程联系起来。.阐述了RNA结合蛋白FPA调控3’末端形成的机制。筛选到FPA可以与保守的3’polyA因子的PA4直接相互作用，并稳定位于一个复合体中。FPA-PA4可共同抑制基因远端polyA位点的使用。推测FPA通过结合特定基因的3’polyA位点，招募PA4在内的3’末端形成因子，促进这些位点的3’末端形成。.分析了PAF1c组分ELF7的N端具有无序结构，且N端介导ELF7与FCA、FPA和PA4的相互作用。体内这些蛋白呈现相分离状态。暗示PAF1c在基因转录过程中提供一个平台的作用，它可以通过ELF7 N端的无序区域招募RNA结合蛋白和3’加工因子，并通过与这些加工因子共在同一个相促进转录与转录后3’末端加工的高效进行。.综上，项目研究完善了PAF1c通过RNA加工调控基因表达的分子机制，并补充了RNA 3’末端形成调控的分子机制。主要结果在分析整理准备撰写论文。发表SCI论文两篇，均标注了该项目的资助。