# Regulation of ENaC expression by paraoxonase-2

> **NIH NIH R03** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2020 · $117,375

## Abstract

Project Abstract
The epithelial sodium channel (ENaC) mediates the rate-limiting step of Na+ uptake across the
apical membrane of specific epithelia. ENaC-dependent Na+ absorption in the kidney has
important roles in regulating extracellular fluid volume, blood pressure and extracellular [K+].
Functional ENaC complexes in the kidney consist of three homologous subunits, namely α, β, and
γ. ENaC functional expression is tightly regulated by multiple intracellular and exogenous factors.
Several molecular chaperones have been implicated in key steps during ENaC biogenesis. The
major goal of this proposal is to investigate the mechanism by which PON-2 regulates ENaC
functional expression. PON-2 is a membrane-bound protein that shares structural features with
MEC-6, a ER-resident chaperone of C. elegans. MEC-6 is required for the proper assembly and
surface expression of the touch-sensing MEC-4/MEC-10 channel in worm touch-receptor
neurons. We recently reported that PON-2 is expressed in principal cells of the distal nephron,
where ENaC resides. PON-2 co-immunoprecipitates with ENaC subunits when co-expressed in
HEK293 cells, and reduces ENaC activity when co-expressed in Xenopus oocytes. The PON-2–
dependent reduction of ENaC activity is associated with a reduction in the number of
Na+ channels at the cell surface. PON-2 lactonase activity is not required for its inhibitory effect
on ENaC activity, suggesting that PON-2, like MEC-6, may function as a molecular chaperone to
regulate ENaC expression. In this application, we propose to examine whether PON-2 reduces
ENaC surface expression by impeding ENaC biogenesis and/or its forwarding trafficking, or by
accelerating ENaC degradation via proteasome or lysosome pathways, by pharmacologically
blocking specific steps of ENaC turnover or by employing mutant αβγ channels that are defective
in gating or endocytosis (Liddle mutations). Another goal is to determine the physiological role of
PON-2 in ENaC expression in Fisher rat thyroid (FRT) cell monolayers. We will examine whether
ENaC expression and/or channel activity in FRT cells is altered under conditions in which PON-
2 is either overexpressed or knocked out. As a proof of concept, we propose to generate a Pon-2
knockout FRT cell line with CRISPR/Cas9 technology to study the physiological role of
endogenous PON-2 in ENaC expression. These studies will provide key preliminary data for future
studies of ENaC regulation by PON-2 in murine Pon-2 knockout models. Understanding
mechanisms by which PON-2 regulates ENaC expression will provide novel insights into both
normal and altered ENaC functional states, such as hypertension.

## Key facts

- **NIH application ID:** 9857596
- **Project number:** 5R03DK119752-02
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** Shujie Shi
- **Activity code:** R03 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $117,375
- **Award type:** 5
- **Project period:** 2019-02-01 → 2022-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9857596

## Citation

> US National Institutes of Health, RePORTER application 9857596, Regulation of ENaC expression by paraoxonase-2 (5R03DK119752-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9857596. Licensed CC0.

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