# SGK Regulation of Epithelial Sodium Transport

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2024 · $475,634

## Abstract

PROJECT SUMMARY/ABSTRACT
Regulated transport of Na+ and K+ in the kidney tubules is central to blood pressure regulation and fluid and
electrolyte homeostasis. The mTORC2-SGK1-ENaC axis is a well-established component of this regulatory
machinery, however, key mechanistic features remain poorly characterized, and in vivo data is limited. We have
used in vivo and in vitro approaches to identify novel features of this signaling system, particularly its role in K+
homeostasis. Our recent data suggest that: 1) K+ acts through WNK kinases to activate mTORC2; 2) tubule cell
mTORC2 is central to the rapid response to a K+ load in mice; 3) Structural features of mTORC2, elucidated
using cryo-EM, play an essential role in mTORC2 activity and specificity. To explore these hypotheses, we will:
Aim 1: Assess the temporal sequence of responses to K+ in mice with tubule-specific knockout of
mTORC2. We have generated a rapidly inducible kidney tubule-specific Rictor KO (TRKO) mouse model.
Preliminary data show that these mice fail to respond to KCl normally; differences between WT and KO mice
manifest in <3h following KCl gavage, and are striking by 48 h of high K diet. To characterize these mice, we will
determine: (A) Time course of response to acute KCl load. Mice will be treated with an oral K+ load and urinary
and plasma parameters will be assessed. Kidneys will be harvested and assessed for tissue signaling
parameters and ion transporter expression and modification. ENaC, ROMK, and BK channel activities will be
measured using patch clamp. Cytoplasm from patched cells will be captured and mRNA subjected to RNA-seq
to identify single-cell gene expression patterns. (B) Time course of gene deletion: Mice will be adapted to a high
K+ diet prior to initiating gene deletion. Balance experiments will be performed on adrenal-intact and ADX + aldo
mice at time points prior to and during emergence of phenotype. Patch clamp will be performed and cytoplasmic
RNA harvested and analyzed as in (A). (C) Evaluate PKC role and identify novel targets. We will identify key
PKC substrates using LC/MS, and functionally characterize them in vivo and in cultured cells. Aim 2: mTORC2
molecular signaling mechanisms and specificity: structural features and effects of WNK kinases. We will
establish the cellular and molecular features that underlie mTORC2 regulation of SGK1 using cultured cells and
cryo-EM. (A) Investigate WNK1 and WNK4 as scaffolds that promote mTORC2-dependent
phosphorylation of SGK1 in a K+-dependent fashion. We will examine effects of WNK kinases on physical
interactions and phosphorylation of SGK1, and functional effects on ENaC and ROMK in cultured mpkCCD and
HEK-293 cells. (B) Cryo-EM structural analysis of mTORC2 core complex, and with SGK1 and WNK1.
mTORC2 core components will be subjected to cryo-EM in the presence and absence of SGK1 and WNK
kinases. Aim 3: Assess the roles of WNK1 and WNK4 and functional interactions with mTORC2 in vivo
using inducible KO ...

## Key facts

- **NIH application ID:** 10782443
- **Project number:** 5R01DK056695-21
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** DAVID PEARCE
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $475,634
- **Award type:** 5
- **Project period:** 2000-08-15 → 2027-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10782443

## Citation

> US National Institutes of Health, RePORTER application 10782443, SGK Regulation of Epithelial Sodium Transport (5R01DK056695-21). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10782443. Licensed CC0.

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