Single-molecule imaging reveals the oligomeric state of functional TNFα-induced plasma membrane TNFR1 clusters in cells
Single-molecule imaging reveals the oligomeric state of functional TNFα-induced plasma membrane TNFR1 clusters in cells
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DOI：
10.1126/scisignal.aax5647
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发表时间：
2020-01-14
影响因子：
7.3
通讯作者：
Heilemann, Mike
中科院分区：
文献类型：
--
作者：
Karathanasis, Christos;Medler, Juliane;Heilemann, Mike
配体诱导的肿瘤坏死因子受体1（TNFR 1）激活控制核因子。活化B细胞（NF-κ B）信号传导、细胞增殖、程序性细胞死亡和存活的轻链增强子，并且在炎症、自身免疫性疾病和癌症进展中至关重要。尽管TNFR 1聚集与信号传导相关，但无配体和配体活化的TNFR 1的寡聚化仍然存在争议。目前，模型的范围从配体非依赖性受体预二聚化到配体诱导的寡聚化。在这里，我们使用定量，单分子超分辨率显微镜直接在天然细胞环境和生理细胞表面丰度研究TNFR 1组装。在缺乏其配体TNF α的情况下，TNFR 1组装成单体和二聚体受体单位。在结合TNF α后，TNFR 1不仅主要聚集成三聚体，而且聚集成更高级的寡聚体。TNFR 1的preligand装配结构域中的功能突变仅导致单体TNFR 1，其表现出受损的配体结合。相比之下，在配体结合CRD 2亚结构域中具有突变的TNFR 1形式保留了未配体的野生型TNFR 1的单体与二聚体比率，但没有表现出配体结合。这些结果强调了配体非依赖性TNFR 1二聚化在NF-κ B信号传导中的重要性。
Ligand-induced tumor necrosis factor receptor 1 (TNFR1) activation controls nuclear factor.-light-chain-enhancer of activated B cells (NF-kappa B) signaling, cell proliferation, programmed cell death, and survival and is crucially involved in inflammation, autoimmune disorders, and cancer progression. Despite the relevance of TNFR1 clustering for signaling, oligomerization of ligand-free and ligand-activated TNFR1 remains controversial. At present, models range from ligand-independent receptor predimerization to ligand-induced oligomerization. Here, we used quantitative, single-molecule superresolution microscopy to study TNFR1 assembly directly in native cellular settings and at physiological cell surface abundance. In the absence of its ligand TNF alpha, TNFR1 assembled into monomeric and dimeric receptor units. Upon binding of TNF alpha, TNFR1 clustered predominantly not only into trimers but also into higher-order oligomers. A functional mutation in the preligand assembly domain of TNFR1 resulted in only monomeric TNFR1, which exhibited impaired ligand binding. In contrast, a form of TNFR1 with a mutation in the ligand-binding CRD2 subdomain retained the monomer-to-dimer ratio of the unliganded wild-type TNFR1 but exhibited no ligand binding. These results underscore the importance of ligand-independent TNFR1 dimerization in NF-kappa B signaling.