ESCRT复合物介导的泛素化蛋白分选降解对于植物蛋白稳态的精确调控以及植物及时调整个体生长发育状态以应对复杂多变的外界环境至关重要。其中重要的科学问题是植物细胞内有哪些蛋白以及它们如何选择性地被ESCRT捕获而进入液泡降解。我们发现植物特有ESCRT组分蛋白FREE1与两个植物miRNA形成的核心调控蛋白HYL1和CPL1互作，且FREE1影响HYL1磷酸化和蛋白稳定性。HYL1直接调控miRNA形成的过程，CPL1则通过去磷酸化提高HYL1活性，但目前尚不清楚HYL1蛋白的降解途径及其分子调控机制。本研究项目拟从生化、分子细胞和遗传等多个角度研究FREE1影响HYL1磷酸化的功能，重点探究FRRE1调控HYL1稳定性的动态过程，最终阐明FREE1影响HYL1磷酸化和蛋白稳定性调控植物miRNA形成的分子机制，为深入理解真核生物ESCRT复合物功能和miRNA形成调控机理提供理论基础。

Tight control of protein homeostasis by ESCRT complexes that mediates ubiquitinated protein sorting into vacuole for degradation, is crucial for plants to adjust the growth and development status and to adapt to the ever-changing environment. An important question is which proteins and how they are selectively captured by the ESCRT complex to sort into vacuole for degradation. In the early stage of this project, it was found that unique plant ESCRT component FREE1 interacts with HYL1 and CPL1, the core regulatory proteins of plant miRNA biosynthesis. Furthermore, the FREE1 affecting the phosphorylation and protein stability of HYL1. HYL1 is directly involved in the process of pri-miRNA shearing to mature miRNA and CPL1 enhances the activity of HYL1 by dephosphorylation, but the molecular mechanism of protein degradation of HYL1 is still unclear. In this study, using a combination of molecular, cellular, biochemical and genetic approaches, we aim to study the function of FREE1 affects the phosphorylation of HYL1. We will focus on the molecular mechanism that FREE1 affects the protein stability of HYL1, and finally clarify the molecular mechanism of FREE1 in the regulation of miRNA biogenesis through affecting the phosphorylation and protein stability of HYL1. This study will reveal the new function of FREE1 and elucidates the cross-regulation between vesicle transport and miRNA biosynthesis, in which provides a theoretical basis for further understanding the function of ESCRT complex and the regulation mechanism of miRNA biosynthesis in eukaryotes.