# High-Resolution Mapping of Subcellular RNA Dynamics Using Photocatalytic Proximity Labeling

> **NIH NIH F32** · PRINCETON UNIVERSITY · 2021 · $65,610

## Abstract

Project Abstract
RNA is a key functional biomolecule across all domains of life, and intracellular movement of different transcripts
is a widespread strategy employed by cells to regulate biological events, including development, metabolism,
cell migration, and neurological function. Given this broad role, it is unsurprising that dysregulated RNA
localization is also linked to a host of diseases, including autism, fragile X syndrome, Alzheimer’s disease,
Huntington’s disease, and several types of cancer. Despite the critical biological importance of intracellular RNA
transport, our understanding of the molecular mechanisms, scale, and impact of these events is significantly
limited and confined by the inherent shortcomings in currently available methods for mapping subcellular RNA
movement. High-throughput tracking of transcript distribution is vital to understanding how RNA contributes to
cellular function and causes disease, and one of the most effective approaches for achieving these goals
employs proximity labeling, whereby a catalyst is embedded into different subcellular locations to biotinylate
nearby molecules. Engineered biotin ligases and peroxidases have shown utility in these applications, but these
techniques also suffer from poor spatial and temporal control over labeling and exhibit in vivo toxicity. To
overcome these limitations, the proposed research will leverage the MacMillan group’s photocatalytic proximity
labeling approach. In particular, this “micromapping” (µMap) method utilizes an iridium photocatalyst to activate
nearby diazirines and label biomolecules of interest. In contrast to enzymatic approaches, this method utilizes a
non-toxic blue light trigger, providing high spatiotemporal control over labeling. In addition, activated diazirines
are very short-lived (T1/2 ~1 ns) and quenched by water, resulting in a small labeling radius (~2 nm) to generate
high-resolution maps of biomolecule localization and interaction networks. Building off these exciting results, this
proposal seeks to leverage this photocatalytic approach toward high-resolution mapping of subcellular RNA
localization and trafficking. Together, this method will enable researchers to map the intracellular transcriptome
with higher spatial resolution, in turn providing better understanding of how these events contribute to cellular
function. In addition, these experiments will help elucidate disease-causing mechanisms related to RNA
transport, and facilitate identification of new diagnostic and therapeutic targets. Lastly, the methods developed
here can be applied to other biological questions surrounding RNA trafficking, including epitranscriptomic
modifications, RNA splicing, and metabolism/turnover, in turn providing an impactful technology that is of broad
utility to the RNA and cell biology community.

## Key facts

- **NIH application ID:** 10232477
- **Project number:** 1F32GM142206-01
- **Recipient organization:** PRINCETON UNIVERSITY
- **Principal Investigator:** Steven Douglas Knutson
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $65,610
- **Award type:** 1
- **Project period:** 2021-05-01 → 2024-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10232477

## Citation

> US National Institutes of Health, RePORTER application 10232477, High-Resolution Mapping of Subcellular RNA Dynamics Using Photocatalytic Proximity Labeling (1F32GM142206-01). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10232477. Licensed CC0.

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