AKT蛋白激酶介导的组蛋白修饰变化对晶状体上皮细胞间充质转化的调控作用

Posterior capsule opacification (PCO) is the most common complication after cataract surgery, causing vision impairment. EMT (Epithelial-Mesenchymal Transition) of the residual lens epithelial cells in the lens capsular bag is a major pathologic change during the development of PCO. Even though a few studies have pointed out the correlation between AKT kinase activation and TGFβ-induced EMT of lens epithelial cells, the underlying regulatory mechanisms remains unknown. In this project, we focus on the phosphorylation substrates of AKT kinases: EZH2 and p300, two important histone modification enzymes, and investigate the function of histone modifications controlled by AKT kinases in TGFβ-induced EMT of lens epithelial cells. Specifically, we aim to elucidate that 1) AKT-EZH2-H3K27me3 axis and AKT-p300-H3/H4Ac axis are important for the regulation of EMT gene expression. 2) The molecular mechanisms by which AKT kinase isoforms AKT1 and AKT2 differentially phosphorylate EZH2 and p300, as well as the consequence of differential H3K27me3 and H3/H4Ac modifications on EMT gene expression. 3) Evaluate the clinical significance of small molecules targeting H3K27me3 and H3/H4Ac modifications in PCO prevention and treatment. The obtained results will guide the application of the available epigenetic drugs for PCO, and provide novel information for the development of PCO therapeutic strategy.

晶体后囊膜混浊（PCO）是人工晶体植入术后造成视力下降的最普遍后遗症，晶状体上皮细胞间充质转化（EMT）是其病理过程中的重要分子事件。AKT蛋白激酶参与转化生长因子TGFβ 诱导的晶状体上皮细胞EMT过程，但具体调控机制不明确。本项目以AKT蛋白激酶对组蛋白修饰酶EZH2和p300的磷酸化修饰为切入点，探索其介导的组蛋白修饰变化调控TGFβ诱导晶状体上皮细胞EMT的机制，将确立1）AKT-EZH2-H3K27me3和AKT-p300-H3/H4Ac通路对EMT基因转录调控的重要作用；2）AKT蛋白激酶异构体AKT1和AKT2差异性磷酸化EZH2和p300的分子机制，及由其介导的差异性H3K27me3和H3/H4Ac对EMT基因表达调控的影响；3）评介靶向H3K27me3和H3/H4Ac的小分子药物在PCO预防和治疗中的临床意义。项目的研究结论将为临床PCO药物的选择使用和研发提供新思路。

晶体后囊膜混浊（PCO）是人工晶体植入术后造成视力下降的最普遍后遗症，晶状体上皮细胞间充质转化（EMT）是其病理过程中的重要分子事件。AKT蛋白激酶参与转化生长因子TGFβ 诱导的晶状体上皮细胞EMT过程，但具体调控机制不明确。本项目旨在剖析其分子机制，为药物的选择使用和新药研发提供新思路。项目总体上按预期计划进行了研究工作。（1）利用分子生细胞生物学实验充分论证了AKT-EZH2-H3K27me3表观遗传途经是调控TGFβ诱导晶状体上皮细胞 EMT基因表达的重要机制；（2）利用质谱分析、细胞生物学等实验鉴定了MYPT1-PP1是负责组蛋白甲基转移酶EZH2 S21位点去磷酸化的重要磷酸酶；（3）利用EZH2 S21A/D磷酸化位点突变体细胞及MYPT1敲除细胞论证了EZH2 S21磷酸化是控制人类晶状体上皮细胞EMT基因表达的关键节点；（4）RNA-seq鉴定出与肿瘤转移侵袭密切相关的多个重要胞外基质（extracellular matrix， ECM）蛋白基因在晶状体上皮细胞EMT进程中的激活表达，并受AKT-EZH2-H3K27me3通路控制。（5）充分利用了损伤诱导前囊下白内障小鼠模型和依托单位临床晶状体囊膜白内障样本资源，在体内验证了EMT基因表达调控的分子机制。研究成果明确了AKT蛋白激酶通过对组蛋白甲基转移酶EZH2的磷酸化进而控制晶状体上皮细胞EMT进程的分子机制，并首次证明肿瘤转移侵袭密切相关ECM基因在晶状体上皮细胞EMT进程中的关键作用；指出AKT-EZH2通路阻断是抑制晶状体上皮细胞EMT进程延缓术后PCO药物开发的潜在策略。

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