Epithelial sodium channels (ENaCs) are members of the ubiquitous ENaC/DEG superfamily of trimeric voltage-independent, Na+-selective and amiloride-sensitive ion channels. Highly expressed in the kidneys, ENaCs assemble as heterotrimers that harbor an exquisitely Na+-selective pore that is critical in the fine tuning of Na+ and K+ balance. ENaC function is regulated at multiple levels from transcription of the ENaC genes, to the trafficking and the proteolytic activation of the ion channel. Abnormal regulation in the functional activity of ENaCs contributes importantly to human disease, and especially to hypertension (high blood pressure), a condition that affects about 1 billion people worldwide. Furthermore, although proteolysis is integral in the activation of ENaCs, aberrant proteolysis of ENaCs contributes to NaCl retention that characterizes nephrotic syndrome. Despite such clinical importance, detail into the mechanism of ENaC function at atomic resolution has remained elusive and the lack of x-ray crystal structures of ENaC has been a barrier to progress in the field. The objective of this research application is to resolve molecula mechanisms underlying ENaC assembly, gating, ion permeation, and allosteric modulation utilizing methods of x-ray crystallography, electrophysiology, and other biochemical and biophysical assays. At present, there are no methods for producing large quantities of heterotrimeric ENaCs for biochemical and biophysical experiments.
The aim of this research application is to develop the technology to express milligram quantities of ENaC that demonstrate a homogenous population suitable for functional and structural assays. Central to this application is to determine the x-ray crystal structures of ENaC at different physiological states and these studies will provide the first atomic-level presentation of these Na+-selective channels in their resting, closed and open states, thus contributing to the understanding of the molecular basis of ENaC function. More importantly, these structural studies will serve as blueprints for future therapeutic strategies.

Public Health Relevance

Epithelial sodium channels (ENaCs) are heteromeric integral membrane proteins that play crucial roles in regulating extracellular fluid volume and blood pressure, thus ENaCs are implicated in hypertension, a condition that affects about 1 billion people worldwide. The role of ENaCs are exemplified by disorders such as Liddle's syndrome and pseudohypoaldosteronism type I (PHA-1), characterized by hypertension (high blood pressure) and hypotension (low blood pressure), respectively, resulting from abnormal ENaC activity. Thus, understanding how ENaCs work at the atomic level is the main goal of this research application.

National Institute of Health (NIH)
Office of The Director, National Institutes of Health (OD)
Early Independence Award (DP5)
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Special Emphasis Panel (ZRG1)
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Basavappa, Ravi
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Oregon Health and Science University
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United States
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Noreng, Sigrid; Bharadwaj, Arpita; Posert, Richard et al. (2018) Structure of the human epithelial sodium channel by cryo-electron microscopy. Elife 7: