# Megalin Traffic in Dent Disease

> **NIH NIH F31** · UNIVERSITY OF PITTSBURGH AT PITTSBURGH · 2021 · $31,647

## Abstract

Project Summary/ Abstract
The polarized epithelial cells that comprise the proximal tubule (PT) have a specialized and high capacity apical
endocytic pathway that is necessary to recover essential nutrients and to maintain a protein-free urine. Proteins
in the ultrafiltrate bind to the multiligand receptors megalin and cubilin on the apical surface of PT cells and are
internalized via receptor-mediated endocytosis. Despite the critical role of this pathway in maintaining PT
function, the molecular identities of the compartments involved in sorting and recycling in PT cells, and thus the
mechanisms by which they are regulated, are largely unknown.
 Impaired PT endocytosis results in urinary excretion of filtered proteins [termed low molecular weight (LMW)
proteinuria]. Dent disease is an X-linked disorder caused by mutations in the CLCN5 gene that encodes CLC-5,
an electrogenic 2Cl-/H+ exchanger. Patients with Dent disease present with LMW proteinuria and typically
progress to renal failure. Loss of CLC-5 has been shown to decrease the endocytic uptake of filtered ligands in
mouse models of Dent disease. PTs from CLCN5 knockout mice exhibit a marked reduction in megalin protein
expression (without altered RNA levels) that likely accounts for the reduced endocytic capacity of these cells. It
has been suggested that loss of CLC-5 causes enhanced degradation of megalin, but this has not been directly
tested and the mechanism by which this might occur is unclear.
 A significant barrier to understanding the regulation of receptor-mediated apical endocytosis of filtered proteins
has been the lack of a highly differentiated cell culture model that retains the organization and high capacity of
the PT apical endocytic pathway. The Weisz lab has optimized a cell culture model of differentiated PT cells that
develops morphological specializations, metabolism, and endocytic capacity similar to the PT in vivo. Using this
system, we can now determine the itinerary, kinetics, and regulation of PT apical endocytic traffic. To this end, I
will develop a model to describe the route and kinetics of megalin traffic in PT cells using data acquired from
imaging and biochemical approaches, and the itinerary of megalin trafficking will be verified in mouse proximal
tubules. This model will be used to make testable predictions for how changes in megalin trafficking result in
LMW proteinuria in Dent disease. I will test these predictions in a Dent disease cell culture model, generated
using CRISPR/Cas9 technology. I hypothesize that loss of CLC-5 alters megalin trafficking, by impairing
recycling and shifting the receptors toward degradative pathways, leading to reduced megalin expression and to
LMW proteinuria. Completing the proposed studies will enhance our understanding of how disease can lead to
LMW proteinuria and provide insight into new approaches to manipulate PT endocytic capacity to preserve
kidney function in proteinuric diseases.

## Key facts

- **NIH application ID:** 10200802
- **Project number:** 5F31DK121394-03
- **Recipient organization:** UNIVERSITY OF PITTSBURGH AT PITTSBURGH
- **Principal Investigator:** Katherine E Shipman
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $31,647
- **Award type:** 5
- **Project period:** 2019-07-01 → 2022-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10200802

## Citation

> US National Institutes of Health, RePORTER application 10200802, Megalin Traffic in Dent Disease (5F31DK121394-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10200802. Licensed CC0.

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