Abstract
Tumor necrosis factor (TNF) is known to activate the epithelial Na+ channel (ENaC) in A549 cells. A549 cells are widely used model for ENaC research. The role of δ-ENaC subunit in TNF-induced activation has not been studied. In this study we hypothesized that δ-ENaC plays a major role in TNF-induced activation of ENaC channel in A549 cells which are widely used model for ENaC research. We used CRISPR/Cas 9 approach to knock down (KD) the δ-ENaC in A549 cells. Western blot and immunofluorescence assays were performed to analyze efficacy of δ-ENaC protein KD. Whole-cell patch clamp technique was used to analyze the TNF-induced activation of ENaC. Overexpression of wild type δ-ENaC in the δ-ENaC KD of A549 cells restored the TNF-induced activation of whole-cell Na+ current. Neither N-linked glycosylation sites nor carboxyl terminus domain of δ-ENaC was necessary for the TNF-induced activation of whole-cell Na+ current in δ-ENaC KD of A549 cells. Our data demonstrated that in A549 cells the δ-ENaC plays a major role in TNF-induced activation of ENaC.
Original language | English |
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Article number | 1858 |
Pages (from-to) | 1-10 |
Number of pages | 10 |
Journal | International Journal of Molecular Sciences |
Volume | 22 |
Issue number | 4 |
DOIs | |
Publication status | Published - 2 Feb 2021 |
Austrian Fields of Science 2012
- 106002 Biochemistry
Keywords
- A549 Cells
- CRISPR-Cas Systems
- Epithelial Sodium Channels/genetics
- Humans
- Tumor Necrosis Factor-alpha/genetics
- CRISPR/Cas9
- Tumor necrosis factor (TNF)
- Epithelial sodium channel (ENaC)