Coordination of DNA damage response and aging by ubiquitin signaling

DNA双链断裂(DSB)的修复受到泛素修饰的严格调控。已知泛素依赖的蛋白分解作用随着年龄的增长而减少，这与受损蛋白聚集的增加有关。相比之下，泛素/蛋白酶体系统(UPS)在与年龄相关的DNA损伤积累和基因组不稳定中的生理作用仍很不清楚。我们最近的工作发现了保守的泛素连接酶UFD-2在DSB修复中的核心作用，这对线虫的生殖系功能、繁殖力和寿命都是重要的。UFD-2是一种特殊的酶，它通过将不可降解的泛素信号转化为26S蛋白酶体底物周转的信号来触发蛋白水解酶。DNA损伤后，UFD-2在胚系中形成局部堆积，持续到同源重组完成，反映了泛素依赖的调控和DSB修复之间的复杂协调。UFD-2位点还含有泛素选择性分离酶CDC-48/p97和去泛素化酶ATX-3/Aaxin-3，两者在染色质相关蛋白的降解、DSB修复和寿命中都具有保守的功能。这项研究的主要目的是了解泛素信号在发育和衰老过程中对基因组稳定性的生理作用。该项目将致力于并系统地分析依赖泛素的DSB修复的时空协调：UFD-2功能的组织特异性调节以及泛素信号和DNA损伤反应(DDR)在衰老中的生理作用。为此，体外和体内泛素化分析、基于生物素化的蛋白质-蛋白质相互作用图谱将结合大规模蛋白质组学、CRISPR/Cas9基因编辑、时间推移显微镜和寿命测量进行。

Repair of DNA double-strand breaks (DSBs) is tightly regulated by ubiquitin modification. Ubiquitin-dependent proteolysis is known to decrease with age, correlating with increased aggregation of damaged proteins. In contrast, the physiological role of the ubiquitin/proteasome-system (UPS) in age-related DNA damage accumulation and genome instability remains largely unclear. Our recent work has uncovered a central role of the conserved ubiquitin ligase UFD-2 in DSB repair, which is important for germline function, fecundity, and lifespan in C. elegans. UFD-2 is a specialized enzyme that triggers proteolysis by converting a nondegradable ubiquitin signal into a signal destined for substrate turnover by the 26S proteasome. Following DNA damage, UFD-2 forms focal accumulations in the germline that persist until homologous recombination is complete, reflecting the intricate coordination between ubiquitin-dependent regulation and DSB repair. The UFD-2 foci also contain the ubiquitin-selective segregase CDC-48/p97 and the deubiquitylation enzyme ATX-3/Ataxin-3, both of which have conserved functions in chromatin-associated protein degradation, DSB repair, and longevity. The main goal of the proposed research is to understand the physiological role of ubiquitin signaling in genome stability during development and aging. The project will address and systematically analyze the spatiotemporal coordination of ubiquitin-dependent DSB repair: the tissue-specific regulation of UFD-2 function and the physiological role of ubiquitin signaling and DNA damage response (DDR) in aging. To this end, in vitro and in vivo ubiquitylation assays, biotinylation-based protein-protein interaction mapping will be performed in combination with large-scale proteomics, CRISPR/Cas9 gene editing, time-lapse microscopy, and lifespan measurements will be performed.