CGG repeat associated translation in Fragile X-associated Tremor/Ataxia Syndrome

脆性X相关震颤/共济失调综合征(FXTAS)是人类神经系统疾病的一大类 这是由于核苷酸重复序列的不稳定和扩张造成的。在FXTAS中，CGG核苷酸重复序列在 脆性X基因的5‘非翻译区，FMR1，并触发聚集性蛋白质包涵体的形成 在病人的大脑里。我们的团队发现，FXTAS CGG重复序列被翻译成有毒的均聚体 蛋白质，尽管它位于规范的开放阅读框架之外，通过一种称为重复的过程 关联的非8月启动(RAN)转换。RAN翻译的蛋白在患者组织中积累并 在多个模型系统中导致CGG重复相关毒性。在这份提案中，我们将探讨如何 这种可供选择的翻译启动是机械地发生的。我们的初步数据显示 CGG重复序列的翻译被细胞压力选择性地增强，这通常会阻止蛋白质的合成。 同时，CGG重复直接引发细胞应激并触发应激颗粒的形成。我们的中央 假说是CGG RAN翻译和细胞应力参与了一个前馈循环，推动 神经退行性变。我们的合作团队将通过使用生化技术直接验证这一假设 以及果蝇、小鼠和人类诱导的FXTAS多能干细胞模型。在目标1中，我们将 确定细胞应激如何选择性地激活RAN翻译，特别关注启动因素 这是这一过程的基础。在目标2中，我们将阐明CGG重复如何引起细胞压力和影响 应激颗粒动力学以及对这一过程的干预是否影响重复毒性。在目标3中，我们将测试 选择性阻断细胞应激通路能否破坏这一前馈环路并缓解CGG 在疾病模型中重复相关的毒性。总而言之，这些研究将阐明 FXTAS和其他核苷酸重复扩增障碍的发病机制同时严格检测两个 对选择性RAN翻译封锁的补充和创新方法。

Fragile X-associated tremor/Ataxia syndrome (FXTAS) is one of a large class of human neurological disorders that result from instability and expansion of nucleotide repeats. In FXTAS, a CGG nucleotide repeat expands in the 5’ untranslated region of the fragile X gene, FMR1, and triggers formation of aggregated protein inclusions in the patient brains. Our group found that the FXTAS CGG repeat gets translated into toxic homopolymeric proteins despite its location outside of a canonical open reading frame through a process known as repeat associated Non-AUG initiated (RAN) translation. RAN translated proteins accumulate in patient tissues and contribute to CGG repeat associated toxicity in multiple model systems. In this proposal, we will explore how this alternative translational initiation occurs mechanistically. Our preliminary data suggests that RAN translation at CGG repeats is selectively enhanced by cellular stress, which typically blocks protein synthesis. In parallel, CGG repeats directly elicit cellular stress and trigger stress granule formation. Our central hypothesis is that CGG RAN translation and cellular stress participate in a feed-forward loop that drives neurodegeneration. Our collaborative team will directly test this hypothesis by using biochemical techniques as well as drosophila, mouse and human induced pluripotent stem cell models of FXTAS. In Aim 1 we will determine how cellular stress selectively activates RAN translation, focusing specifically on initiation factors that underlie this process. In Aim 2, we will elucidate how CGG repeats elicit cellular stress and influence stress granule dynamics and whether interventions in this process impact repeat toxicity. In Aim 3, we will test whether selective blockade of cellular stress pathways can disrupt this feed forward loop and alleviate CGG repeat associated toxicity across disease models. Together, these studies will illuminate critical events in the pathogenesis of FXTAS and other nucleotide repeat expansion disorders while rigorously testing two complementary & innovative approaches to selective RAN translation blockade.