# Spatiotemporal analysis of TDP-43 toxicity and endolysosomal turnover mechanisms

> **NIH NIH R56** · UNIVERSITY OF ARIZONA · 2022 · $393,027

## Abstract

Amyotrophic Lateral Sclerosis (ALS) is a devastating disease in which progressive degeneration of motor
neurons leads to paralysis, usually resulting in death 2-5 years after diagnosis. No therapies exist that
significantly increase quality of life or life expectancy. Mutations in >30 genes are linked to ALS onset, albeit
>90% of all ALS cases are sporadic. However, a unifying cellular hallmark in >95% of all ALS cases is the
cytoplasmic mis-localization, accumulation, and aggregation of the nuclear RNA binding protein TDP-43 (TAR
DNA/RNA binding protein 43) in motor neurons and support cells. Similar TDP-43 pathology is observed in other
neurodegenerative diseases, including in forebrain neurons of ~50% of Frontotemporal dementia patients (FTD).
TDP-43 pathology confers a toxicity to neurons, but the nature of this toxicity remains fiercely debated. Loss of
nuclear function (LOF) and gain of cytoplasmic function (GOF) mechanisms have been proposed, though
separating such mechanisms and identifying the earliest impacts of TDP-43 pathology has remained elusive.
Regardless, a therapeutic strategy that has shown promise in some ALS models is promoting the degradation
of cytoplasmic TDP-43. Recently, cytoplasmic TDP-43 was shown to be degraded via a novel endolysosomal
pathway, which, when induced, suppresses TDP-43 toxicity. However, understanding of this degradation
pathway remains limited. Key gaps in understanding include determining, in an ALS-relevant neuronal model,
the earliest and most disease-relevant impacts of TDP-43 pathology and defining how endolysosomal TDP-43
degradation occurs. The aims of this grant are: 1.) Establish a novel endogenous TDP-43 reporter system in
neurons that allows precise control of TDP-43 abundance and cellular localization via small molecule and
optogenetic means. Using this system, the impacts of altered TDP-43 levels and localization on TDP-43 itself,
ALS phenotypes and gene expression, focusing on RNA abundance and translation, will be examined. 2.) Test
an endolysosomal degradation model involving TDP-43 ubiquitination and endosomal membrane invagination
in neurons using an optical pulse labelling approach. TDP-43 degradation mechanisms will also be defined in
patient-derived ALS models using similar means. Finally, a novel high throughput yeast dot-blot assay will be
used to identify genetic and chemical regulators of TDP-43 and Fused in Sarcoma (FUS) abundance, which is
also implicated in ALS and FTD pathology. This grant is innovative in that a novel approach to exert
spatiotemporal control of TDP-43 expression, which promises separation of TDP43 LOF and GOF toxicity
effects, and a means to identify regulators of TDP-43 and FUS abundance via dot blot, are proposed. Finally,
mechanistically defining endolysosomal-based means of cytoplasmic TDP-43 degradation promises new basic
insight into proteostasis for TDP-43 and other substrates. In summary, TDP-43 and FUS are logical entry points
for the study of ALS...

## Key facts

- **NIH application ID:** 10626673
- **Project number:** 1R56NS128110-01
- **Recipient organization:** UNIVERSITY OF ARIZONA
- **Principal Investigator:** Sami Barmada
- **Activity code:** R56 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $393,027
- **Award type:** 1
- **Project period:** 2022-07-01 → 2024-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10626673

## Citation

> US National Institutes of Health, RePORTER application 10626673, Spatiotemporal analysis of TDP-43 toxicity and endolysosomal turnover mechanisms (1R56NS128110-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10626673. Licensed CC0.

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