miR-125b/PML-RARα反馈通路在APL发生发展中的作用及机制

The molecular feature of APL is an aberrant chromosomal translocation that juxtaposes the PML gene with the RARα gene, and the subsequent expression of the PML-RARα fusion protein blocks the differentiation of promyelocytes and leads to an accumulation of leukemic promyelocytes. Although the pathogenetic importance of fusion protein in APL has been recognized for over a decade, the mechanism by which PML-RARα generates the disease is far from clear, in particular gene activation by fusion protein and the regulation of itself. Our privious study show that the t(15;17) translocation leads to elevated microRNA-125b (miR-125b) expression; Interestingly,miR-125b also increased PML-RARα expression, suggesting that PML/RARα and miR-125b may form a positive feedback loop and play an important role in APL leukemogenesis. This study will investigate the fuction and machanism of miR-125b/PML-RARα feedback loop in APL leukemogenesis. Here,we will explore the molecular mechanism of PML-RARα activates transcriptional regulation of miR-125b through chromatin immunoprecipitation assay, electrophoretic mobility shift assay and promoter luciferase assay.We will perform 3’-UTR luciferase activity assay, western blotting, RNAi and quantitative real-time PCR to demonstrate that miR-125b regulates the expression of PML-RARα through altered interaction of the PML-RARα promoter with transcription factors affected by target genes of miR-125b.Further,we will investigated the effect of miR-125b in APL cell proliferation, and leukemia progression in vivo. Understanding the mechanisms of miR-125b/PML-RARα feedback loop and their roles in APL pathogenesis will provide a novel potential target for diagnosis and therapy.

急性早幼粒细胞白血病（APL）的主要遗传学特征是PML-RARα融合基因的产生，尽管研究已显示PML-RARα在APL发病中的重要性，但其调控机制仍未完全明确，尤其是PML-RARα自身表达的调控及其激活部分基因表达的方式仍不甚清楚。我们研究发现，miR-125b与PML-RARα形成正反馈回路，提示miR-125b可能协同PML-RARα在APL发病中起重要作用。本研究拟在此基础上，探讨PML-RARα与miR-125b形成的正反馈回路的调控机制，及其可能在APL发生发展中的作用。①明确PML-RARα激活miR-125b表达的机制。②验证miR-125b靶向的转录因子对PML-RARα的调控作用。③在体内及体外观察miR-125b对APL的恶性特征的影响。从而阐明miR-125b与PML-RARα之间相互调控的机制，探讨其对APL发生的影响，为APL的治疗提供新的生物标志物和治疗靶点。

PML-RARα融合基因是急性早幼粒细胞白血病（APL）的主要致病基因，但其调控机制仍未完全明确，对PML-RARα在诱导APL发病中的作用及其机制的深入研究，对了解APL的发病机理有重要的意义。本项目研究发现APL的致病因子PML-RARα融合蛋白和转录因子NRF2的表达能促进miR-125b的表达，PML-RARα主要通过激活NRF2这个转录因子促进miR-125b的表达；我们研究还发现， miR-125b的靶基因STAT3可调控PML-RARα的表达，说明miR-125b与PML-RARα可能形成正反馈回路。miR-125b的表达进一步激活AKT及MAPK信号通路，进一步发现miR-125b通过抑制BTG2、MAP3K11、RPS6KA1 和 PRDM1等靶基因的表达促进APL细胞的增殖。我们进一步检测了BTG2、MAP3K11、RPS6KA1 和 PRDM1等miR-125b靶基因的在初发APL细胞中的表达。我们的研究成果丰富了APL发生机制和miR-125b在疾病发生发展中的作用机制的认识。