GSK3b mediates radiation-induced cytotoxicity in hippocampal neurons

描述（由申请人提供）：治疗原发性和转移性脑癌（尤其是儿童）的一个巨大挑战是由颅脑照射（IR）诱导的海马神经元凋亡导致的长期神经认知缺陷。我们的实验室发现了代谢激酶GSK 3 β与修复DNA双链断裂（DSB）的非同源末端连接（NHEJ）途径之间的新功能联系。此外，我们的初步数据显示，NHEJ介体53 BP 1直接被GSK 3 β磷酸化;同时，经典GSK 3 β底物β-连环蛋白表达的增加与IR诱导的DSB的修复增强和海马神经元的存活有关。因此，我们假设GSK 3 β通过抑制53 BP 1和β-catenin功能调节NHEJ介导的DSB修复，并决定IR后神经元的细胞毒性。此外，含有异常GSK 3 β活性的肿瘤细胞不会表现出GSK 3 β介导的对IR诱导的细胞毒性的保护作用。本研究拟通过一系列的体内外实验来验证我们的假设：目的1将确定53 BP 1中的GSK 3 β磷酸化位点，并确定GSK 3 β特异性磷酸化是否指导53 BP 1在NHEJ中的功能和受辐射海马神经元的存活。目的2将确定GSK 3 β是否通过抑制β- catenin来调节53 BP 1，β- catenin可能通过增加53 BP 1转录或直接与53 BP 1相互作用来促进NHEJ活性。目的3将确定是否异常的GSK 3 β活性决定脑肿瘤细胞对预防性GSK 3 β抑制介导的辐射诱导细胞毒性保护的抗性。

DESCRIPTION (provided by applicant): A formidable challenge in the treatment of primary and metastatic brain cancers, especially in children, is the long-term neurocognitive deficiencies resulting from cranial irradiation (IR)-induced hippocampal neuronal apoptosis. Our laboratory has discovered a novel functional connection between the metabolic kinase GSK3� and the Non- homologous End-joining (NHEJ) pathway that repair DNA double-strand breaks (DSBs). Furthermore, our preliminary data revealed that the NHEJ mediator 53BP1 is directly phosphorylated by GSK3�; meanwhile, increased expression of the classic GSK3� substrate �-catenin is associated with enhanced repair of IR-induced DSBs and survival in hippocampal neurons. Thus, we hypothesize that GSK3� regulates NHEJ-mediated repair of DSBs and determines neuron cytotoxicity following IR via suppression of 53BP1 and �-catenin function. In addition, tumor cells which contain abnormal GSK3 � activity will not exhibit GSK3�-mediated protection from IR-induced cytotoxicity. A series of in vitro and in vivo experiments are proposed to test our hypotheses: Aim 1 will identify the GSK3 � phosphorylation sites in 53BP1 and determine whether GSK3 � -specific phosphorylation direct 53BP1 function in NHEJ and in survival of irradiated hippocampal neurons. Aim 2 will determine whether GSK3 � regulates 53BP1 through suppressing � - catenin that may promote NHEJ activity by increasing 53BP1 transcription, or by directly interacts with 53BP1. Aim 3 will determine if abnormal GSK3 � activity determine brain tumor cell resistance to the prophylactic GSK3 � -inhibition mediated protection from radiation induced cytotoxicity.