# Single molecule study of C9ORF72 repeat RNA metabolisms in ALS/FTD

> **NIH NIH RF1** · JOHNS HOPKINS UNIVERSITY · 2020 · $3,156,336

## Abstract

The goal of this proposal is to investigate the pathogenic mechanism of amyotrophic
lateral sclerosis (ALS) and frontotemporal degeneration (FTD) induced by hexanucleotide repeat
expansion in the C9ORF72 gene. The expansion of GGGGCC repeats in the first intron of
C9ORF72 is the most common genetic cause of both ALS and FTD. The disease has been
predominantly attributed to the RNA repeats induced toxicity, including RNA granules that
sequester essential RNA binding proteins (RBP), and the toxic poly-dipeptides produced through
repeat-associated non-AUG (RAN) translation. Previous biochemical and genetic studies have
revealed cellular pathways impaired by the repeat expansion. However, the biophysical properties
of RNA granules and DPR biogenesis are still poorly understood at the molecular level. Here we
combine cutting edge in vivo single molecule imaging approach with biochemical analysis and
high-throughput screening to study the repeat RNA metabolisms at every RNA processing step,
including transcription and splicing, turnover and aggregation properties, RAN translation, RNA
localization and trafficking in reporter cell lines, patient derived cells and mouse models. We will
fluorescently label both sense and antisense RNA with different colors and study the biophysical
property of RNA granules and their interactions in live cells. We will measure the translation
efficiency, initiation and elongation dynamics, and possible frameshift in live cells for each reading
frame of both sense and antisense repeats. We will label C9ORF72 intron and exon with different
colors and examine the splicing and repeat-mediated intron export. We will test candidate genetic
modifiers of both RNA granules and RAN translation, as well as screen for candidates involved in
the nuclear export of repeat-containing intron. All these pathways will be examined at single
molecule level in live cells for the dynamic properties. We will also determine whether the repeat
RNA metabolisms have cell type-specific features and how stress stimuli and neuronal activities
could influence these properties. The approach proposed here allows us to address the
mechanistic problems intractable by any other techniques. The molecular insights resulting from
this study will help understand the etiology of the disease and develop novel therapeutic strategy.

## Key facts

- **NIH application ID:** 9859796
- **Project number:** 1RF1NS113820-01
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Shuying Sun
- **Activity code:** RF1 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $3,156,336
- **Award type:** 1
- **Project period:** 2020-03-01 → 2025-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9859796

## Citation

> US National Institutes of Health, RePORTER application 9859796, Single molecule study of C9ORF72 repeat RNA metabolisms in ALS/FTD (1RF1NS113820-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9859796. Licensed CC0.

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