circ-MYBL2通过调控母基因MYBL2蛋白表达抑制白血病细胞分化及分子机制研究

Compelling evidences demonstrate that circRNAs play important roles in tumorigenesis through interacting with miRNAs and RNA-binding proteins (RBPs). In the pilot studies, we did reveal that circ-MYBL2 suppresses acute myeloid leukemia (AML) cells differentiation via activating the MYBL2 transcription. Multiple layers of evidence from our first-hand bioinformatics analyses show that 1) both circ-MYBL2 and MYBL2 mRNA carry miR-149-3p binding sites; and 2) circMYBL2 contains 5 binding sites of RBPs, among of which are three functional proteins for transcriptional regulation, namely EIF4A3, IGF2BP1 and IGF2BP3. All these point to the role of circ-MYBL2 in the mRNA stability and translation. Thus, we hypothesize that circ-MYBL2 cis-regulated MYBL2 through 1) sequestering miR-149-3p and 2) stabilizing MYBL2 mRNA, which finally promotes MYBL2 transcription and translation by interacting with EIF4A3, IGF2BP1 and IGF2BP3. Herein, we are setting out to apply a series of molecular methods (i.e., RIP, CRISPR, luciferase etc.) to address two aims: 1) systemically get insight of the clinical significance of circ-MYBL2 and its translational potential (i.e., a prognostic marker), and 2) characterize the mechanisms of circ-MYBL2/MYBL2 regulation. By doing so, the whole page of circ-MYBL2 will definitely broaden our knowledge about AML leukemogenesis and provide a new therapeutic option for AML treatment.

研究表明circRNA主要通过结合miRNA及RNA结合蛋白（RBP）影响恶性肿瘤的重要生物学特征。本课题组前期证明circ-MYBL2 通过上调母基因MYBL2的蛋白表达，抑制白血病细胞分化。生物信息学分析发现①circ-MYBL2与MYBL2 mRNA均具有miR-149-3p结合位点；②circ-MYBL2具有5个RBP的结合位点，其中EIF4A3、 IGF2BP1和 IGF2BP3具有促进mRNA翻译及稳定的功能。提出假说：circ-MYBL2上调MYBL2蛋白表达的可能机制：①与miR-149-3p结合，释放MYBL2 mRNA，促进其翻译；②与上述三个RBP结合，增加MYBL2 mRNA稳定性并促进翻译。本课题组拟采用多种分子生物学（RIP, CRISPR等）技术对circ-MYBL2在AML中的表达、功能及其调控分化的机制进行研究。此项目的完成可能为白血病治疗提供新靶点。

白血病是由未成熟白细胞分化异常和过度增殖所致。因此，发现白血病中导致分化异常的关键因子以及机制至为重要。本项目通过对急性髓系白血病（AML）细胞分化过程中的异常表达的环状RNA（circRNA）进行高通量测序，筛选出了差异性circRNA，即circ-MYBL2。本项目进一步通过功能试验和机制研究，检测了circ-MYBL2在AML细胞分化、增殖和凋亡中的作用，并通过检测circMYBL2是否与miR-149-3p以及RNA结合蛋白EIF4A3、 IGF2BP1和 IGF2BP3相互作用来研究其潜在的机制。本项目研究结果发现，circMYBL2的主要作用是抑制AML细胞的分化，而对细胞增殖和凋亡的影响不大。而circMYBL2抑制细胞分化的作用是通过与IGF2BP1结合后增强MYBL2 mRNA的翻译水平促进MYBL2蛋白的表达来实现的。本项目对揭示AML发生发展的分子机制具有重要意义，有望为白血病的诊断和治疗提供新的思路和理论依据。本项目标注下发表SCI论文4篇。此外，申请并获得国家发明专利2项，培养博士后1名。