# Defining protein:protein interactions for the regulation of renal V-ATPase function: role in expression, assembly and trafficking.

> **NIH NIH R01** · MASSACHUSETTS GENERAL HOSPITAL · 2022 · $539,372

## Abstract

PROJECT SUMMARY
 Despite its central role in extracellular acidification in the kidney and other organs, as well as in critical
intracellular processes, the regulation of proton-pumping ATPase (V-ATPase) activity at the molecular level is poorly
understood. During the prior funding period, two proteins that associate strongly with the V-ATPase to regulate its
function were identified – Ncoa7 and Dmxl1. The overall objective of this proposal is to determine the mechanisms
by which they interact with the V-ATPase to regulate proton secretion by the kidney, thereby maintaining systemic
acid/base balance. The long-term goal is to develop strategies (including drug and peptide design) for the regulation
of acidification processes that are inappropriately up- or downregulated not only in the kidney, but also in diseases
affecting other cells and organs. Ncoa7 null mice have markedly decreased V-ATPase subunit expression in collecting
duct intercalated cells (ICs) resulting in distal RTA, while Dmxl1 knockdown in renal epithelial cells in vitro causes
deficient intracellular vesicle acidification to the same degree as knockdown of bone-fide V-ATPase subunits. Aim 1
will determine the mechanism by which Ncoa7 regulates V-ATPase subunit expression, focusing on translational,
transcriptional and degradation pathways in WT and Ncoa7 knockout mice. The V-ATPase-binding sequence of Ncoa7
will be identified by protein interaction and mutagenesis studies, and its role in V-ATPase-dependent acidification
events will be determined. Aim 2 will address the novel hypothesis that V-ATPase exocytosis and accumulation on
the plasma membrane of ICs in response to systemic acid/base cues requires, counter-intuitively, a partial disassembly
of the large, sterically hindering V-ATPase holoenzymes that coat intracellular transport vesicles. The working
hypothesis is that Dmxl1, a homolog of the Rav1p yeast V-ATPase assembly protein, coordinates V-ATPase
assembly/disassembly with V-ATPase recycling to and from the plasma membrane, which together regulate V-ATPase
activity and proton secretion in kidney ICs and other cells. The functionally important V-ATPase-binding sequence of
Dmxl1 will also be identified by protein interaction and mutagenesis studies. Thus, a major innovative aspect of our
proposed studies is the concept that two newly-identified V-ATPase interacting proteins are involved in the
regulation of V-ATPase function at the “upstream” expression level and the “downstream” assembly level. Both
Aims 1 and 2 make use of integrated cell and molecular techniques in conjunction with genetically modified mouse
models, isolated ICs and renal cell cultures in vitro, and interaction domain studies using purified proteins and specific
subdomains. The proposed research is significant: a) because it will allow the field to move forward not at the most
basic cellular level by elucidating new V-ATPase dependent acidification regulatory pathways, and b) because the
interaction ...

## Key facts

- **NIH application ID:** 10454931
- **Project number:** 5R01DK121848-04
- **Recipient organization:** MASSACHUSETTS GENERAL HOSPITAL
- **Principal Investigator:** Dennis Brown
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $539,372
- **Award type:** 5
- **Project period:** 2019-09-01 → 2024-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10454931

## Citation

> US National Institutes of Health, RePORTER application 10454931, Defining protein:protein interactions for the regulation of renal V-ATPase function: role in expression, assembly and trafficking. (5R01DK121848-04). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10454931. Licensed CC0.

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