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

> **NIH NIH R01** · UNIVERSITY OF MICHIGAN AT ANN ARBOR · 2021 · $308,100

## 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:** 10242059
- **Project number:** 5R01GM135252-03
- **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:** $308,100
- **Award type:** 5
- **Project period:** 2019-09-20 → 2023-08-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10242059

## Citation

> US National Institutes of Health, RePORTER application 10242059, Chemical Biology Approach for Validating and Manipulating Cellular RNA-Protein Interactions (5R01GM135252-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10242059. Licensed CC0.

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