Department of Developmental Neurobiology, St. Jude Children's Research Hospital, 262 Danny Thomas Place, Memphis, TN 38105, USA.
Center for Proteomics and Metabolomics, St. Jude Children's Research Hospital, 262 Danny Thomas Place, Memphis, TN 38105, USA.
Cell Rep Methods. 2024 Oct 21;4(10):100875. doi: 10.1016/j.crmeth.2024.100875. Epub 2024 Oct 8.
Loss of proteostasis is a hallmark of aging that underlies many age-related diseases. Different cell compartments experience distinctive challenges in maintaining protein quality control, but how aging regulates subcellular proteostasis remains underexplored. Here, by targeting the misfolding-prone Fluc luciferase to the cytoplasm, mitochondria, and nucleus, we established transgenic sensors to examine subcellular proteostasis in Drosophila. Analysis of detergent-insoluble and -soluble levels of compartment-targeted Fluc variants indicates that thermal stress, cold shock, and pro-longevity inter-organ signaling differentially affect subcellular proteostasis during aging. Moreover, aggregation-prone proteins that cause different neurodegenerative diseases induce a diverse range of outcomes on Fluc insolubility, suggesting that subcellular proteostasis is impaired in a disease-specific manner. Further analyses with Fluc and mass spectrometry indicate that pathogenic tau produces an unexpectedly complex regulation of solubility for different Fluc variants and protein subsets. Altogether, compartment-targeted Fluc sensors pinpoint a diverse modulation of subcellular proteostasis by aging regulators.
蛋白质稳态的丧失是衰老的一个标志，是许多与年龄相关疾病的基础。不同的细胞区室在维持蛋白质质量控制方面都面临着独特的挑战，但衰老如何调节亚细胞蛋白质稳态仍未得到充分探索。在这里，我们通过将易错误折叠的 Fluc 荧光素酶靶向细胞质、线粒体和细胞核，建立了转基因传感器来检测果蝇的亚细胞蛋白质稳态。对靶向 Fluc 变体的去污剂不溶性和可溶性水平的分析表明，热应激、冷休克和延长寿命的器官间信号在衰老过程中对亚细胞蛋白质稳态有不同的影响。此外，导致不同神经退行性疾病的聚集倾向蛋白对 Fluc 不溶性产生不同的影响，这表明亚细胞蛋白质稳态以特定疾病的方式受到损害。使用 Fluc 和质谱的进一步分析表明，致病性 tau 对不同的 Fluc 变体和蛋白质亚群的溶解度产生了出乎意料的复杂调节。总之，靶向 Fluc 的传感器突出了衰老调节剂对亚细胞蛋白质稳态的多样化调节。
