Inhibitory Tract Traps the Epithelial Na+ Channel in a Low Activity Conformation

Kashlan, Ossama B.; Blobner, Brandon M.; Zuzek, Zachary; Carattino, Marcelo D.; Kleyman, Thomas R.

doi:10.1074/jbc.m112.358218

Cited by 23 publications

(36 citation statements)

References 21 publications

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“…The absence of an effect with Cu 2+ alone is consistent with previous work (22). Notably, extracellular DTT treatment did not affect wild type currents, consistent with previous studies (20,23,24). Wild type channel insensitivity to DTT suggests either that native extracellular disulfide bonds are not susceptible to reduction by DTT, or that reduction of those bonds does not affect channel function.…”

Section: Finger Domain Cysteines In the A Subunit Are Near The Fingersupporting

confidence: 92%

“…1A). We previously reported evidence of crosslinking between this site and Y474C at the top of the thumb domain in the a subunit (20). When we perfused bifunctional MTS-2-MTS for 1 min, we observed an increase in channel currents that remained elevated after washout of the crosslinker (Fig.…”

Section: Crosslinking Across the G Subunit Finger-thumb Domain Interfmentioning

confidence: 60%

“…1A) and the thumb domain (blue circle in Fig. 1A) of the a subunit using short MTS-based crosslinkers (20). Inhibition by MTS crosslinkers was length-dependent.…”

Section: Finger Domain Cysteines In the A Subunit Are Near The Fingermentioning

confidence: 97%

“…For example, the a and g subunits are subject to activating proteolysis, while the b and d subunits are not (12,26,27). We hypothesized that the conformations of the channel pore and the g subunit's finger-thumb domain interface were coupled, analogous to the a subunit (20). We introduced a cysteine at position 115 in the g subunit (gD115C), which is equivalent to site 171 in helix 1 of the a subunit at the fingerthumb domain interface (see Fig.…”

Section: Crosslinking Across the G Subunit Finger-thumb Domain Interfmentioning

confidence: 99%

“…1A). Indeed, directly crosslinking the finger-thumb interface inhibited the channel in a crosslinker-length dependent manner (20).…”

Section: Introductionmentioning

confidence: 99%

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

Blobner¹,

Wang²,

Kashlan

2018

Journal of Biological Chemistry

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The epithelial Na + channel (ENaC) is a member of the ENaC/Degenerin family of ion channels. In the structure of a related family member, the 'thumb' domain's base interacts with the pore, and its tip interacts with the divergent 'finger' domain. Between the base and tip, the thumb domain is characterized by a conserved 5-rung disulfide ladder holding together two anti-parallel a helices. The ENaC a and g subunits' finger domains harbor autoinhibitory tracts that can be proteolytically liberated to activate the channel, and also host an ENaC-specific pair of cysteines. Using a crosslinking approach, we show that one of the finger domain cysteines in the a subunit (aC263) and both of the finger domain cysteines in the g subunit (gC213 and gC220) lie near the dynamic finger-thumb domain interface. Our data suggest that the aC256/aC263 pair is not disulfide bonded. In contrast, we found that the gC213/gC220 pair is disulfide bonded. Our data also suggest the g subunit lacks the terminal rung in the thumb domain disulfide ladder, suggesting asymmetry between the subunits. We also observed functional asymmetry between the a and g subunit finger-thumb domain interfaces: crosslinks bridging the a subunit fingerthumb interface only inhibited ENaC currents, while crosslinks bridging the g subunit fingerthumb interface activated or inhibited currents dependent on the length of the crosslinker. Our data suggest that reactive cysteines lie at the dynamic finger-thumb interfaces of the a and g subunits, and may play a yet undefined role in channel regulation.

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Section: Finger Domain Cysteines In the A Subunit Are Near The Fingersupporting

confidence: 92%

Section: Crosslinking Across the G Subunit Finger-thumb Domain Interfmentioning

confidence: 60%

“…1A) and the thumb domain (blue circle in Fig. 1A) of the a subunit using short MTS-based crosslinkers (20). Inhibition by MTS crosslinkers was length-dependent.…”

Section: Finger Domain Cysteines In the A Subunit Are Near The Fingermentioning

confidence: 97%

Section: Crosslinking Across the G Subunit Finger-thumb Domain Interfmentioning

confidence: 99%

“…1A). Indeed, directly crosslinking the finger-thumb interface inhibited the channel in a crosslinker-length dependent manner (20).…”

Section: Introductionmentioning

confidence: 99%

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

Blobner¹,

Wang²,

Kashlan

2018

Journal of Biological Chemistry

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Epithelial Na ⁺ Channels Function as Extracellular Sensors

Kashlan,

Wang,

Sheng

et al. 2024

Comprehensive Physiology

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The epithelial Na + channel (ENaC) resides on the apical surfaces of specific epithelia in vertebrates and plays a critical role in extracellular fluid homeostasis. Evidence that ENaC senses the external environment emerged well before the molecular identity of the channel was reported three decades ago. This article discusses progress toward elucidating the mechanisms through which specific external factors regulate ENaC function, highlighting insights gained from structural studies of ENaC and related family members. It also reviews our understanding of the role of ENaC regulation by the extracellular environment in physiology and disease. After familiarizing the reader with the channel's physiological roles and structure, we describe the central role protein allostery plays in ENaC's sensitivity to the external environment. We then discuss each of the extracellular factors that directly regulate the channel: proteases, cations and anions, shear stress, and other regulators specific to particular extracellular compartments. For each regulator, we discuss the initial observations that led to discovery, studies investigating molecular mechanism, and the physiological and pathophysiological implications of regulation. © 2024 American Physiological Society. Compr Physiol 14:5407‐5447, 2024.

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Epithelial Sodium Channels (ENaC)

Song¹,

Ma²,

Eaton³

2020

Studies of Epithelial Transporters and Ion Channels

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Inhibitory Tract Traps the Epithelial Na+ Channel in a Low Activity Conformation

Cited by 23 publications

References 21 publications

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

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

Epithelial Na ⁺ Channels Function as Extracellular Sensors

Epithelial Sodium Channels (ENaC)

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Inhibitory Tract Traps the Epithelial Na+ Channel in a Low Activity Conformation

Cited by 23 publications

References 21 publications

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

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

Epithelial Na + Channels Function as Extracellular Sensors

Epithelial Sodium Channels (ENaC)

Contact Info

Product

Resources

About

Epithelial Na ⁺ Channels Function as Extracellular Sensors