Determine how telomere dysfunction impacts neuronal function

在这里，我们将：i)定义为什么端粒功能障碍选择性地影响齿状回（DG）的神经发生，ii)我们将定义DG端粒功能障碍时神经元损失的机制。为了确定为什么端粒功能障碍选择性地影响齿状回的神经发生，我们将测试端粒功能障碍选择性地影响成年出生的颗粒细胞GCs的假设，因为它们的成熟阶段延长。为此，我们制备了条件敲除TRF2的小鼠胚胎干细胞，该干细胞可分化为神经元细胞。这个系统将允许我们测试分化的时间在端粒功能障碍反应中起主要作用的假设。同时，我们将采用在DG分化过程中发生改变的小鼠遗传学模型。为了确定DG中端粒功能障碍导致神经元丢失的机制，我们将利用我们之前旨在剖析细胞对TRF2耗竭反应的工作，评估DNA损伤激活和端到端染色体融合的作用。在这个系统中，DNA损伤反应的激活可以通过耗尽ATM来消除，在较小程度上，也可以通过耗尽p53来消除。抑制nhej通路完全消除端到端染色体融合的发生。总的来说，这些实验将更好地理解端粒在中枢神经系统中的作用，以及更深入地理解神经元DNA损伤激活导致神经元损失的分子机制。

Here we will: i) define why telomere dysfunction selectively affects neurogenesis in the dentate gyrus (DG) and, ii) we will define the mechanism of neuronal loss upon telomere dysfunction in the DG. To define why telomere dysfunction selectively affects neurogenesis in the dentate gyrus we will test the hypothesis that telomere dysfunction selectively affects adult-born granule cells GCs due to their prolonged maturation phase. To this end, we generated conditional knockout TRF2 mouse embryonic stem cells that can be differentiated into neuronal cells. This system will allow us to test the hypothesis that the timing of differentiation plays a major role in response to telomere dysfunction. In parallel, we will employ mouse genetics models with alteration in the process of DG differentiation. To define the mechanism of neuronal loss upon telomere dysfunction in the DG we will assess the role of the DNA damage activation and onset of end-to-end chromosome fusions taking advantage of our previous work aimed at dissecting the cellular response to TRF2 depletion. In this system, activation of the DNA damage response can be abolished by depletion of ATM and, to a lesser extent, by depletion of p53. Suppression of the NHEJ-pathway completely abolishes the onset of end-to-end chromosome fusions. Collectively, these experiments will provide a better understanding of the role of telomeres in the CNS as well as a deeper understanding of the molecular mechanism that leads to neuronal loss upon DNA damage activation in neurons.