# RNA structures critical for dictating the contents, behavior, and function of droplets formed from liquid-liquid phase separation

> **NIH NIH F32** · UNIV OF NORTH CAROLINA CHAPEL HILL · 2022 · $25,259

## Abstract

Project Summary:
Background: Analogous to the separation of oil from water, a process called liquid-liquid phase separation
(LLPS) partitions cellular contents. LLPS is dictated by intrinsic biophysical/biochemical properties of
component molecules. Many phase-separated droplets form from RNA-binding proteins in complex with target
RNAs. These droplets play roles in neuromuscular/neurodegenerative diseases, and cancer. In pathological
contexts, the dynamics of these molecules are disrupted. It is now appreciated that RNA sequence/structure,
can dictate assembly and physical state of droplets. This provides a new role for RNA sequence, which is
encoding mesoscale biophysical properties of cellular bodies. There is a link between RNA sequence and
droplet behaviors but the details are still a mystery.
Objective/Hypothesis: To better understand how RNA structure regulates droplets, I will measure and
manipulate RNA structure temporally and spatially. I will then use these measurements to gain mechanistic
insights into how droplet maturation occurs and how droplets can facilitate or inhibit RNA translation. We
hypothesize that sequence directed RNA structures dictates the contents, behavior, and function of
condensed droplets and this in turn influences cell biology.
Specific Aims: (1) To determine the role of RNA/RNA interactions in droplets. (2) To map RNA structures and
RNA protein interaction changes that influence maturation of droplets. (3) To determine how RNA structures at
the core and shell of droplets differ and impact functions.
Study Design: I will use the model protein called Whi3 from genetically tractable fungi where RNA-structure is
known to be critical to droplet formation with target RNAs. I will measure RNA/RNA interaction, RNA structure,
and protein binding during the course of droplet maturation. I will then disrupt the structures and interactions
identified by mutating RNA sequences, to assess the impact of these features on droplet formation,
maturation, and biophysical properties (size, viscosity, diffusivity, etc.). I will next determine how the
biophysical properties of the shell influence cell biological phenotypes by measuring how droplets
regulate translation efficiency.
Innovation: Thus far, a detailed examination of how RNA structure relates to the material and functional state
of droplets has not been undertaken. The kinetics of RNA structure and protein binding are unknown, and
RNA/RNA interactions in droplets have not been measured. Ultimately, this work will provide insight into the
role of RNA structures in driving maturation and physical state in droplets formed from LLPS, and models
developed herein will be generalizable to all RNA protein complexes. Therefore, this work contributes to a
better understanding of how to manipulate RNA structures in the context of human diseases.
!

## Key facts

- **NIH application ID:** 10327720
- **Project number:** 5F32GM136164-03
- **Recipient organization:** UNIV OF NORTH CAROLINA CHAPEL HILL
- **Principal Investigator:** Christine Anne Roden
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $25,259
- **Award type:** 5
- **Project period:** 2020-02-01 → 2022-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10327720

## Citation

> US National Institutes of Health, RePORTER application 10327720, RNA structures critical for dictating the contents, behavior, and function of droplets formed from liquid-liquid phase separation (5F32GM136164-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10327720. Licensed CC0.

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