# Chemical Biology Approach for Validating and Manipulating Cellular RNA-Protein Interactions

> **NIH NIH R01** · UNIVERSITY OF MICHIGAN AT ANN ARBOR · 2021 · $78,169

## Abstract

ABSTRACT
Recent studies have shown that RNAs are invariably bound to and often modified by RNA-binding proteins
(RBPs). Thus, it is no surprise that RBPs have been found to play key roles in regulating many aspects of coding
and non-coding RNA biology, including RNA processing, nuclear export, cellular transport, function, localization,
and stability. These efforts are carried out by >1,500 unique RBPs that utilize a variety of RNA-binding domains
to achieve oftentimes specific and high affinity interactions with target transcripts; however, non-canonical RBPs
have also been identified. Disruption of this complex network of RNA-protein interactions (RPIs) has been
implicated in a number of human diseases. Thus, the targeting of RBPs and RPIs has arisen as a new frontier
in RNA-targeted drug discovery; however, very few interactions have been validated to support a pipeline of
targets for these efforts. While the advent of sequencing and quantitative mass spectrometry has dramatically
enhanced our ability to globally profile these interactions, experimental validation of these data sets remains a
challenge. Using chemical biology- and bioorthogonal chemistry-based strategies, we have developed an
innovative new assay for the live-cell detection of RPIs, RNA interaction with Protein-mediated Complementation
Assay, or RiPCA. Through this approach, we have detected the interaction of pre-miRNAs with RBPs, in addition
to inhibition with small molecules. Moreover, to provide evidence for the potential of our technology in validating
new RPIs, we used RiPCA to confirm the interaction of a pre-miRNA with a novel RBP discovered via proteomics.
In total, these data provide encouraging proof-of-concept for this emerging technology; yet, many key questions
and challenges still remain to ensure that RiPCA is a rigorous and unbiased approach for the detection of RPIs
in distinct cellular organelles. In Specific Aim 1, we will further develop RiPCA for organelle-specific detection
to ensure its accuracy. In Specific Aim 2, we will investigate the potential of the assay by profiling additional
RPIs from various RNA and RBP families. Finally, in Specific Aim 3, we will explore its adaptability toward high-
throughput experimentation for validation of large-scale CLIP or proteomics data sets, or screening to identify
cell-active small molecule inhibitors of RPIs. Upon completion of the proposed research, our goal is to produce
a robust and user-friendly technology for the rapid validation and study of cellular RPIs to enable biomedical
research.

## Key facts

- **NIH application ID:** 10408902
- **Project number:** 3R01GM135252-03S1
- **Recipient organization:** UNIVERSITY OF MICHIGAN AT ANN ARBOR
- **Principal Investigator:** Amanda Garner
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $78,169
- **Award type:** 3
- **Project period:** 2019-09-20 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10408902

## Citation

> US National Institutes of Health, RePORTER application 10408902, Chemical Biology Approach for Validating and Manipulating Cellular RNA-Protein Interactions (3R01GM135252-03S1). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10408902. Licensed CC0.

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