Determine how telomere dysfunction impacts neuronal function

在这里，我们将：i)确定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中端粒功能障碍导致神经元丢失的机制。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，我们将检验这样的假设：由于成熟期延长，端粒功能障碍选择性地影响成体出生的颗粒细胞GC。为此，我们产生了条件基因敲除的TRF2小鼠胚胎干细胞，可以分化为神经细胞。这个系统将使我们能够检验这样的假设，即分化的时机在端粒功能障碍的反应中起着主要作用。同时，我们将使用在DG分化过程中发生改变的小鼠遗传学模型。为了明确DG中神经元丢失导致端粒功能障碍的机制，我们将利用我们先前旨在剖析TRF2缺失的细胞反应的工作，评估DNA损伤、激活和端到端染色体融合的开始所起的作用。在这个系统中，DNA损伤反应的激活可以通过ATM的耗尽而取消，在较小程度上也可以通过p53的耗尽来取消。抑制NHEJ途径完全消除了端到端染色体融合的开始。总之，这些实验将使我们更好地理解端粒在中枢神经系统中的作用，以及更深入地理解在神经元DNA损伤激活时导致神经元丢失的分子机制。

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.