Pesce Emanuela, Sondo Elvira, Ferrera Loretta, Tomati Valeria, Caci Emanuela, Scudieri Paolo, Musante Ilaria, Renda Mario, Baatallah Nesrine, Servel Nathalie, Hinzpeter Alexandre, di Bernardo Diego, Pedemonte Nicoletta, Galietta Luis J V
U.O.C. Genetica Medica, IRCCS Istituto Giannina Gaslini, Genoa, Italy.
Telethon Institute of Genetics and Medicine, Pozzuoli, Italy.
Front Pharmacol. 2018 Dec 13;9:1464. doi: 10.3389/fphar.2018.01464. eCollection 2018.
The mutation F508del, responsible for a majority of cystic fibrosis cases, provokes the instability and misfolding of the CFTR chloride channel. Pharmacological recovery of F508del-CFTR may be obtained with small molecules called correctors. However, treatment with a single corrector and only leads to a partial rescue, a consequence of cell quality control systems that still detect F508del-CFTR as a defective protein causing its degradation. We tested the effect of spautin-1 on F508del-CFTR since it is an inhibitor of USP10 deubiquitinase and of autophagy, a target and a biological process that have been associated with cystic fibrosis and mutant CFTR. We found that short-term treatment of cells with spautin-1 downregulates the function and expression of F508del-CFTR despite the presence of corrector VX-809, a finding obtained in multiple cell models and assays. In contrast, spautin-1 was ineffective on wild type CFTR. Silencing and upregulation of USP13 (another target of spautin-1) but not of USP10, had opposite effects on F508del-CFTR expression/function. In contrast, modulation of autophagy with known activators or inhibitors did not affect F508del-CFTR. Our results identify spautin-1 as a novel chemical probe to investigate the molecular mechanisms that prevent full rescue of mutant CFTR.
导致大多数囊性纤维化病例的F508del突变会引发CFTR氯离子通道的不稳定和错误折叠。使用称为校正剂的小分子可以实现F508del-CFTR的药理学恢复。然而，单一校正剂治疗仅能带来部分挽救效果，这是由于细胞质量控制系统仍将F508del-CFTR检测为有缺陷的蛋白质并导致其降解。我们测试了spautin-1对F508del-CFTR的影响，因为它是USP10去泛素酶和自噬的抑制剂，而自噬是一个与囊性纤维化和突变型CFTR相关的靶点和生物学过程。我们发现，尽管存在校正剂VX-809，但用spautin-1对细胞进行短期处理会下调F508del-CFTR的功能和表达，这一发现在多个细胞模型和实验中均得到证实。相比之下，spautin-1对野生型CFTR无效。USP13（spautin-1的另一个靶点）的沉默和上调而非USP10，对F508del-CFTR的表达/功能有相反的影响。相比之下，用已知的自噬激活剂或抑制剂调节自噬并不影响F508del-CFTR。我们的研究结果确定spautin-1是一种新型化学探针，可用于研究阻止突变型CFTR完全挽救的分子机制。
