The prion-like domain of Fused in Sarcoma is phosphorylated by multiple kinases affecting liquid- and solid-phase transitions.
The prion-like domain of Fused in Sarcoma is phosphorylated by multiple kinases affecting liquid- and solid-phase transitions.
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DOI：
10.1091/mbc.e20-05-0290
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发表时间：
2020-11-01
影响因子：
3.3
通讯作者：
Shewmaker F
中科院分区：
文献类型：
--
作者：
Owen I;Rhoads S;Yee D;Wyne H;Gery K;Hannula I;Sundrum M;Shewmaker F
融合肉瘤（FUS）是一种广泛表达的蛋白质，可以从核质和细胞质中相分离成不同的液滴结构。它主要是核，其大部分功能与RNA和DNA代谢有关。细胞质相分离组件内FUS的过度持久性与肌萎缩性侧索硬化症和额颞叶痴呆症有关。在细胞模型和体外实验中，细胞核磷脂酰肌醇3-激酶相关激酶（PIKK）家族激酶在DNA损伤后磷酸化FUS的朊病毒样结构域（PrLD）可改变FUS的液相和固相转变。然而，蛋白质组学数据表明FUS的PrLD在许多其他位点被磷酸化，并且尚不清楚其他非PIKK和非核激酶是否可能影响FUS的相变。在这里，我们评估疾病突变和压力条件下，增加FUS积累到细胞质相分离的结构。我们观察到，细胞质液相结构含有FUS磷酸化的新网站，这发生独立的PIKK家族激酶。我们在FUS的PrLD中设计了磷酸模拟物取代，并观察到模拟一些磷酸化位点强烈抑制FUS固相聚集，同时最小限度地改变液相冷凝。这些作用的发生独立于磷酸化模拟物取代的确切位置，这表明调节PrLD磷酸化可能提供对疾病中观察到的固相聚集具有特异性的治疗策略。
Fused in Sarcoma (FUS) is a ubiquitously expressed protein that can phase-separate from nucleoplasm and cytoplasm into distinct liquid-droplet structures. It is predominantly nuclear and most of its functions are related to RNA and DNA metabolism. Excessive persistence of FUS within cytoplasmic phase-separated assemblies is implicated in the diseases amyotrophic lateral sclerosis and frontotemporal dementia. Phosphorylation of FUS’s prion-like domain (PrLD) by nuclear phosphatidylinositol 3-kinase-related kinase (PIKK)-family kinases following DNA damage was previously shown to alter FUS’s liquid-phase and solid-phase transitions in cell models and in vitro. However, proteomic data suggest that FUS’s PrLD is phosphorylated at numerous additional sites, and it is unknown if other non-PIKK and nonnuclear kinases might be influencing FUS’s phase transitions. Here we evaluate disease mutations and stress conditions that increase FUS accumulation into cytoplasmic phase-separated structures. We observed that cytoplasmic liquid-phase structures contain FUS phosphorylated at novel sites, which occurred independent of PIKK-family kinases. We engineered phosphomimetic substitutions within FUS’s PrLD and observed that mimicking a few phosphorylation sites strongly inhibited FUS solid-phase aggregation, while minimally altering liquid-phase condensation. These effects occurred independent of the exact location of the phosphomimetic substitutions, suggesting that modulation of PrLD phosphorylation may offer therapeutic strategies that are specific for solid-phase aggregation observed in disease.