拟南芥HYL1互作蛋白HYC1在microRNA加工代谢中的作用机制研究

微小RNA（miRNA）是一类在动植物中保守的重要的小RNA调控分子，广泛调控发育、代谢和逆境胁迫等各种生物过程。miRNA以碱基互补配对识别其并负调的其目标mRNA，因而miRNA序列直接决定了其靶标mRNA及miRNA本身的功能。miRNA序列是生物合成决定的，所以miRNA的生物合成一直是该领域研究的重要方向。拟南芥中，miRNA前体高效而准确的切割需要HYL1等蛋白对RNA酶III/DCL1的精细调控。为了克服遗传学方法在发现关键基因的局限性，我们利用蛋白免疫沉淀方法鉴定了HYL1结合的新蛋白HYC1，发现了HYC1沉默的植株中成熟miRNA水平比对照植株中的下降，因而推测HYC1参与miRNA加工代谢。本项目将利用我们建立的miRNA体外重组加工系统，结合遗传、细胞等手段，阐明HYC1的具体作用方式，在miRNA加工代谢方面，有望深入揭示通过遗传学方法无法发现的新分子机制。

microRNA is a class of conserved small regulatory RNA. microRNAs take part in many physiologic processes such as development, metabolism, and stress response etc., by regulating thousands of target mRNAs. microRNAs inhibit gene expression by translational repression or to promote their cleavage by pairing with their target mRNAs. Hence the regulating targets and specific function of one microRNA are determined by the microRNA sequence itself. Since, the sequence of microRNA is determined by its coding MIRNA genes and how it is generated, the molecular mechanisms of microRNA biogenesis and metabolism is of intensive interests in the field. The results of genetic studies in Arabidopsis indicate that the RNase III DICER-Like1 (DCL1) is required for the effective and accurate processing of microRNA precursors with the help of co-factor HYPONASTIC LEAVES1 (HYL1), in the Arabidopsis cell nucleus. In order to overcome the limitations of genetic approaches in the discovery of essential genes' function, we identified an unknown HYL1 interacting protein HYC1 (HYL1 COMPLEX1) by immunoprecipitation. The microRNA level is decreased in the HYC1 RNAi plants, indicating that HYC1 may be involved in microRNA biogenesis or metabolism. Therefore, we propose to elucidate the detailed mechanism on how HYC1 affects microRNA biogenesis by in vitro microRNA processing assay in combination with genetic, cell biology, and bioinformatics approaches. Hopefully, our work will shed light on the microRNA biogenesis mechanism in plant.