circ-AFF2调控PRC2复合物介导的AML发生发展机制研究

Acute myeloid leukemia (AML) is a kind of malignant neoplasm with severe heterogeneity that seriously affects human health. Our high-throughput sequencing result indicated that some circular RNAs (such as circ-AFF2 and circ-RNF220) established higher expression in the peripheral blood mononuclear cells of AML patients than that in healthy volunteers, which may serve as potential regulators of AML. By RNA-purified chromatin separation (ChIRP) and mass spectrometry, we found that circ-AFF2 binds to EED and EZH2 which are components of PRC2 protein complex. Overexpression of circ-AFF2 in AML cells significantly promotes the proliferation, and upregulates the expression level of target genes of PRC2 such as HOXA7 and HOXA9. Therefore, this project intends to explain the mechanism by which circ-AFF2 regulates PRC2 complex-mediated AML development from the aspects of bioinformatics, molecular and cell biology. The project will provide a theoretical basis for the development of AML diagnosis and treatment strategies.

急性髓性白血病(acute myeloid leukemia, AML)是一种异质性很强的恶性肿瘤疾病，严重影响人类的健康。课题组高通量测序分析发现环状RNA如circ-AFF2、circ-RNF220等在AML患者外周血单个核细胞中高表达，可能作为AML的潜在调控因子。通过RNA纯化的染色质分离（ChIRP）技术联合质谱鉴定技术，我们发现circ-AFF2与PRC2蛋白复合物组分EED和EZH2蛋白的结合，过表达circ-AFF2会显著促进AML细胞的增殖，并上调PRC2调控的靶基因如HOXA7、HOXA9等的表达。因此，本项目拟从生物信息学、分子及细胞生物学的角度着重阐释circ-AFF2调控PRC2复合物介导的AML发生发展机制。项目的开展，将为AML诊断和治疗策略的开拓提供理论基础。

circRNA是一类环状非编码RNA，被证实可以调控多种癌症的发生发展。本研究中，我们通过收集临床样本进行高通量测序，利用生物信息学工具对急性髓系白血病（AML）中差异表达的circRNA进行分析。通过差异分析我们在AML中筛选出差异表达的circRNA的391个，其中高表达的有256个，低表达的有135个。通过验证发现circAFF2在AML样本和AML细胞系中均显著高表达（p<0.05），并且在细胞质和细胞核中均有分布。在AML细胞THP-1和KG-1a中通过用siRNA干扰实验发现circAFF2的表达可以显著敲降（p<0.05），而母本基因的表达不受影响。同时与对照组相比，敲低circAFF2的表达可以抑制AML细胞的增殖，同时诱导AML细胞的凋亡，表明circAFF2有促进AML发生发展的作用。小鼠体内实验则证明了敲低circAFF2细胞系对白血病细胞的影响。相对于空载细胞，移植敲低circAFF2的细胞能够显著增加小鼠的生存时间，并且抑制白血病细胞的数量。最后，通过RNA pull down和质谱检测了可能与circAFF2作用的蛋白。