Conserved cysteines in the finger domain of the epithelial Na+ channel α and γ subunits are proximal to the dynamic finger–thumb domain interface

Blobner, Brandon M.; Wang, Xue‐Ping; Kashlan, Ossama B.

doi:10.1074/jbc.m117.819367

Cited by 8 publications

(6 citation statements)

References 44 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…1A). Furthermore, we have shown that the finger-thumb domain interfaces of both the ␣ and ␥ subunits can be manipulated independently of peptide addition to modulate channel function (21,22). These data suggest that inhibitory peptide binding occurs at dynamic domain interfaces whose conformations are functionally coupled to the conformation of the channel pore.…”

Section: ␥ Cleavage Relieves Inhibition At the Finger-thumb Interfacementioning

confidence: 78%

“…5A). We and others have reported that longer exposures to bifunctional methanethiosulfonate (MTS) compounds irreversibly inhibit ENaC (21)(22)(23)(24). This step labels accessible cysteines on the channel with MTS-4-MTS, potentially leaving an unreacted MTS group tethered to the mutated site.…”

Section: ␥-11 Derivatives Cross-link To Sites In the ␥ Subunit Fingermentioning

confidence: 99%

See 1 more Smart Citation

The epithelial Na+ channel γ subunit autoinhibitory tract suppresses channel activity by binding the γ subunit's finger–thumb domain interface

Balchak

Thompson

Kashlan

2018

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Epithelial Na channel (ENaC) maturation and activation require proteolysis of both the α and γ subunits. Cleavage at multiple sites in the finger domain of each subunit liberates their autoinhibitory tracts. Synthetic peptides derived from the proteolytically released fragments inhibit the channel, likely by reconstituting key interactions removed by the proteolysis. We previously showed that a peptide derived from the α subunit's autoinhibitory sequence (α-8) binds at the α subunit's finger-thumb domain interface. Despite low sequence similarity between the α and γ subunit finger domains, we hypothesized that a peptide derived from the γ subunit's autoinhibitory sequence (γ-11) inhibits the channel through an analogous mechanism. Using oocytes, we found here that channels lacking a γ subunit thumb domain were no longer sensitive to γ-11, but remained sensitive to α-8. We identified finger domain sites in the γ subunit that dramatically reduced γ-11 inhibition. Using cysteines and sulfhydryl reactive cross-linkers introduced into both the peptide and the subunit, we also could cross-link γ-11 to both the finger domain and the thumb domain of the γ subunit. Our results suggest that α-8 and γ-11 occupy similar binding pockets within their respective subunits, and that proteolysis of the α and γ subunits activate the channel through analogous mechanisms.

show abstract

Section: ␥ Cleavage Relieves Inhibition At the Finger-thumb Interfacementioning

confidence: 78%

Section: ␥-11 Derivatives Cross-link To Sites In the ␥ Subunit Fingermentioning

confidence: 99%

The epithelial Na+ channel γ subunit autoinhibitory tract suppresses channel activity by binding the γ subunit's finger–thumb domain interface

Balchak

Thompson

Kashlan

2018

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…In our ENaC FL structure, the P3 and P4 strands are linked by a loop containing a predicted glycosylation site adjacent to the α5 helix of the thumb domain in all three subunits. Additionally, the important residue αGly225 is adjacent to αThr240 and forms hydrogen bonds with the C-terminal end of the αP1 peptide ( Figure 5b ; Blobner et al, 2018 ).…”

Section: Resultsmentioning

confidence: 99%

Molecular principles of assembly, activation, and inhibition in epithelial sodium channel

Noreng

Posert

Bharadwaj

et al. 2020

eLife

View full text Add to dashboard Cite

The molecular bases of heteromeric assembly and link between Na+ self-inhibition and protease-sensitivity in epithelial sodium channels (ENaCs) are not fully understood. Previously, we demonstrated that ENaC subunits – α, β, and γ – assemble in a counterclockwise configuration when viewed from outside the cell with the protease-sensitive GRIP domains in the periphery (Noreng et al., 2018). Here we describe the structure of ENaC resolved by cryo-electron microscopy at 3 Å. We find that a combination of precise domain arrangement and complementary hydrogen bonding network defines the subunit arrangement. Furthermore, we determined that the α subunit has a primary functional module consisting of the finger and GRIP domains. The module is bifurcated by the α2 helix dividing two distinct regulatory sites: Na+ and the inhibitory peptide. Removal of the inhibitory peptide perturbs the Na+ site via the α2 helix highlighting the critical role of the α2 helix in regulating ENaC function.

show abstract

“…The extracellular domains represent ~70% of the sequence of subunits, with the highest homology between ASICs and ENaC found in the palm and β-ball regions [ 44 , 48 ]. In contrast, the finger domain is the least conserved domain in the ENAC/degenerin family, as evidenced by an ASIC1 subunit that shares only 8% of homology with the α subunit of ENaC in the finger domain [ 48 , 49 ].…”

Section: Tlp Activation Of the Epithelial Sodium Channel (Enac)mentioning

confidence: 99%

Trypsin-Like Proteases and Their Role in Muco-Obstructive Lung Diseases

Carroll

Bailo

Reihill

et al. 2021

IJMS

View full text Add to dashboard Cite

Trypsin-like proteases (TLPs) belong to a family of serine enzymes with primary substrate specificities for the basic residues, lysine and arginine, in the P1 position. Whilst initially perceived as soluble enzymes that are extracellularly secreted, a number of novel TLPs that are anchored in the cell membrane have since been discovered. Muco-obstructive lung diseases (MucOLDs) are characterised by the accumulation of hyper-concentrated mucus in the small airways, leading to persistent inflammation, infection and dysregulated protease activity. Although neutrophilic serine proteases, particularly neutrophil elastase, have been implicated in the propagation of inflammation and local tissue destruction, it is likely that the serine TLPs also contribute to various disease-relevant processes given the roles that a number of these enzymes play in the activation of both the epithelial sodium channel (ENaC) and protease-activated receptor 2 (PAR2). More recently, significant attention has focused on the activation of viruses such as SARS-CoV-2 by host TLPs. The purpose of this review was to highlight key TLPs linked to the activation of ENaC and PAR2 and their association with airway dehydration and inflammatory signalling pathways, respectively. The role of TLPs in viral infectivity will also be discussed in the context of the inhibition of TLP activities and the potential of these proteases as therapeutic targets.

show abstract

Conserved cysteines in the finger domain of the epithelial Na+ channel α and γ subunits are proximal to the dynamic finger–thumb domain interface

Cited by 8 publications

References 44 publications

The epithelial Na+ channel γ subunit autoinhibitory tract suppresses channel activity by binding the γ subunit's finger–thumb domain interface

The epithelial Na+ channel γ subunit autoinhibitory tract suppresses channel activity by binding the γ subunit's finger–thumb domain interface

Molecular principles of assembly, activation, and inhibition in epithelial sodium channel

Trypsin-Like Proteases and Their Role in Muco-Obstructive Lung Diseases

Contact Info

Product

Resources

About