自噬相关因子Atg7对DNA修复蛋白RAD51的调控及其对肿瘤形成的影响机制研究

Defects in the cellular recycling process of autophagy can lead to genomic instability, which is associated with DNA damage responses (DDR) and greatly increases the threat of neoplastic transformation in humans. The RAD51 recombinase plays key roles in homologous pairing and strand transfer of DNA and is essential in the homologous recombination and repair of DNA. In our recent work, we identified a novel protein interaction between the essential autophagy gene Atg7 and the recombinase RAD51, suggesting a crosstalk of the autophagy and DDR processes. We found that the Atg7-dependent autophagy activity regulates the posttranslational modification of RAD51 protein and also controls the formation of RAD51 nuclear foci, which indicates the involvement of the activation of DNA repair pathways. Moreover, our further functional analysis of Atg7-deficent cells suggests that the lack of Atg7 increases DNA damage and promotes neoplastic growth. Therefore, we hypothesize that Atg7 may regulate RAD51 protein modification and function and play an important role in tumor growth. In present study we will investigate the interaction of Atg7 and RAD51 and characterize the RAD51 protein modification and functions that are regulated by Atg7-dependent autophagy activity. We will use Atg7 knockout animal model and also develop xenograft tumor model in nude mice to analyze the influence of Atg7-mediated protein modification of RAD51 in the activation of DNA repair pathways and tumor growth. Our study will elucidate the novel protein modification and functions of RAD51 that are regulated by Atg7 and suggest a strategy for improving targeted therapy for neoplastic growth.

细胞自噬与DNA损伤修复是维系细胞稳态的重要细胞行为。RAD51蛋白是DNA损伤修复途径中的重要一员，其参与DNA损伤修复或稳定DNA复制，从而影响基因组的稳定性，与肿瘤形成有紧密关联。我们的新近研究结果表明自噬相关因子Atg7通过与RAD51相结合并调控RAD51蛋白的修饰，这种依赖于Atg7细胞自噬反应的RAD51蛋白修饰影响RAD51蛋白在细胞核内foci的形成，从而参与DNA损伤修复，影响肿瘤发生发展。本项目将从Atg7与RAD51的蛋白交互和协同作用入手，以Atg7基因敲除动物模型为背景，结合基因敲除肿瘤细胞裸鼠种植动物模型，动态观察Atg7介导RAD51蛋白及其DNA损伤应答反应对肿瘤形成的影响，解析Atg7调控RAD51蛋白修饰的类型和功能，并且探索其作为新的药物靶点的可能，为设计合理的治疗方案提供依据。

自噬及相关因子是否协同参与细胞在不同情况和不同程度应对DNA复制压力的DNA损伤修复反应，目前研究尚不明确。在实验中，我们首次发现ATG7是以E1-like泛素激活酶的身份参与蛋白RAD51的泛素化修饰，调控其所介导的DNA损伤修复反应，从而协同影响肿瘤发生发展。本研究针对ATG7泛素化修饰RAD51蛋白，参与DNA损伤修复的机制和功能进行探讨，从多方面证实了ATG7作为E1-like泛素激活酶，调控RAD51蛋白的泛素化修饰；这一修饰调控是RAD51参与DNA损伤修复反应的必要条件，解析了细胞自噬参与DNA损伤修复反应的分子机制。与此同时，针对ATG7泛素化修饰RAD51的机制所设计的功能多肽Rap，在体内和体外实验中，能够抑制DNA损伤修复机制，提高肿瘤细胞的药物敏感性，促进肿瘤细胞凋亡。上述研究，为寻找安全有效的DNA修复抑制剂进行抗肿瘤治疗，提供了新的理论依据和研究基础。