Journal
AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY
Volume 321, Issue 2, Pages L308-L320Publisher
AMER PHYSIOLOGICAL SOC
DOI: 10.1152/ajplung.00298.2020
Keywords
CFTR; cystic fibrosis; ENaC; FLIM; Liddle's syndrome
Categories
Funding
- National Institutes of Health (NIH) [R21HL085112]
- GMS [R01GM123971]
- NIH [DK37206, P30DK072482, R474-CR11, R01 DK059600]
- National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) [T32 DK007545]
- UAB-UCSD O'Brien Center for Acute Kidney Injury Research - NIDDK [P30 DK079337]
- UAB Health Services Foundation General Endowment Fund [2004622]
- MBRU-COM Internal Grant Awards [MBRU-CM-RG2018-04, MBRU-CMRG2018-05]
- Sandooq Al Watan Research & Development Grant [SWARD-F2018-002]
- Al Mahmeed Collaborative Research Award
- Al Jalila Foundation Grant [AJF201763]
- MBRU Post-Doctoral Fellow Award [MBRU-PD-2020-04]
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The study demonstrates a close physical association between CFTR and ENaC in the pathophysiology of cystic fibrosis, even in the presence of Liddle's syndrome mutations, suggesting a direct intermolecular interaction between these proteins. The findings are supported by coimmunoprecipitation and fluorescence resonance energy transfer measurements, indicating a potential therapeutic target for CF treatment.
The association of the cystic fibrosis transmembrane conductance regulator (CFTR) and epithelial sodium channel (ENaC) in the pathophysiology of cystic fibrosis (CF) is controversial. Previously, we demonstrated a close physical association between wild-type (WT) CFTR and WT ENaC. We have also shown that the F508del CFTR fails to associate with ENaC unless the mutant protein is rescued pharmacologically or by low temperature. In this study, we present the evidence for a direct physical association between WT CFTR and ENaC subunits carrying Liddle's syndrome mutations. We show that all three ENaC subunits bearing Liddle's syndrome mutations (both point mutations and the complete truncation of the carboxy terminus), could be coimmunoprecipitated with WT CFTR. The biochemical studies were complemented by fluorescence lifetime imaging microscopy (FLIM), a distance-dependent approach that monitors protein-protein interactions between fluorescently labeled molecules. Our measurements revealed significantly increased fluorescence resonance energy transfer between CFTR and all tested ENaC combinations as compared with controls (ECFP and EYFP cotransfected cells). Our findings are consistent with the notion that CFTR and ENaC are within reach of each other even in the setting of Liddle's syndrome mutations, suggestive of a direct intermolecular interaction between these two proteins.
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