Abstract
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 in vivo and in vitro 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.
Highlights
CFTR, a plasma membrane chloride channel with a main role in epithelial cells, is mutated in cystic fibrosis (CF), one of the most frequent genetic diseases (Stoltz et al, 2015; Castellani and Assael, 2017)
USP13 appears as an important protein regulating the fate of mutant CFTR while spautin-1 may become an interesting probe for mechanistic studies and the search of new therapeutic agents
The decrease in CFTR function by spautin-1 was paralleled by an altered pattern of F508del-CFTR maturation as indicated by immunoblot experiments
Summary
CFTR, a plasma membrane chloride channel with a main role in epithelial cells, is mutated in cystic fibrosis (CF), one of the most frequent genetic diseases (Stoltz et al, 2015; Castellani and Assael, 2017). It has been shown that more marked levels of F508del-CFTR rescue can be obtained with combinations of correctors having complementary mechanisms of action (Farinha et al, 2013; Okiyoneda et al, 2013). Such other correctors may work by binding to a second site in the CFTR protein or by modulation of the cell machinery responsible for CFTR processing and degradation. We found that spautin-1 antagonizes the rescue by VX-809 causing a rapid rundown of F508del-CFTR at the functional and molecular level This effect may involve USP13 inhibition but is independent from autophagy block. USP13 appears as an important protein regulating the fate of mutant CFTR while spautin-1 may become an interesting probe for mechanistic studies and the search of new therapeutic agents
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