# Quantifying Physiologic and Pathologic Viscoelastic Phases of Biomolecular Condensates by Correlative Force and Fluorescence Microscopy

> **NIH NIH R35** · STATE UNIVERSITY OF NEW YORK AT BUFFALO · 2021 · $395,329

## Abstract

SUMMARY
In recent years, it has become increasingly clear that the material properties of ribonucleoprotein (RNP) granules,
which are formed via liquid-liquid phase separation, play crucial roles in both cellular physiology and pathology.
Nevertheless, mechanistic understandings of the molecular determinants and modulators of RNP granule
viscoelastic phases remain incomplete due to the limitations of currently available techniques to probe for protein
condensate dynamics across single-molecule to mesoscale. The goal of this proposal is to address this critical
gap by the development of a multi-parametric experimental toolbox that simultaneously reports on RNP
condensate structure and dynamics across different length-scales, with high sensitivity. Our approach will feature
correlative multicolor single-molecule fluorescence microscopy, dual-trap optical tweezers, and microfluidics.
Utilizing our novel toolbox, we will decipher the mechanisms of liquid-to-liquid and liquid-to-solid phase
transitions of intracellular RNP condensates, processes that critically contribute to the onset or development of
many neurodegenerative diseases including amyotrophic lateral sclerosis (ALS) and frontotemporal dementia
(FTD). Commonly used fluorescence microscopy techniques, such as fluorescence recovery after
photobleaching (FRAP), offer only probe-specific protein/RNA diffusivity within the RNP granules. In contrast,
our proposed correlative force-fluorescence microscopy platform will provide a multiscale view of RNP
condensate dynamics by taking advantage of optical tweezer-based rheological and fluid dynamics
measurements in conjunction with quantification of protein dynamics using single-molecule fluorescence. We
hypothesize that (a) a hierarchy of protein-protein and protein-nucleic acid interactions determines both
nanoscale RNP dynamics and micron-scale material properties of the condensate, and (b) post-translational
modifications, RNA/DNA and ligand binding, and pathologic mutations modulate the material properties of RNP
condensates by manipulating the long-range and short-range inter-molecular forces. Overall, our research
program will address three Key Challenges (KCs): (a) we will develop a novel experimental toolbox based
on correlative multi-color confocal fluorescence microscopy and dual-trap optical tweezer that simultaneously
reports on molecular and mesoscale protein condensate structure and dynamics (KC 1), (b) we will apply our
toolbox to map the transition pathways of physiologic RNP granules to pathologic states in c9orf72 repeat
expansion disorder (KC 2), and (c) we will identify mechanisms of ligand-dependent transcriptional condensate
regulation at DNA enhancer sites (KC 3). Our studies will provide new insights into the determinants of functional
RNP condensate material states, dynamics, and composition, as well as identify novel pathways of these
granules’ pathologic alterations.

## Key facts

- **NIH application ID:** 10231209
- **Project number:** 5R35GM138186-02
- **Recipient organization:** STATE UNIVERSITY OF NEW YORK AT BUFFALO
- **Principal Investigator:** Priya R. Banerjee
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $395,329
- **Award type:** 5
- **Project period:** 2020-08-15 → 2025-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10231209

## Citation

> US National Institutes of Health, RePORTER application 10231209, Quantifying Physiologic and Pathologic Viscoelastic Phases of Biomolecular Condensates by Correlative Force and Fluorescence Microscopy (5R35GM138186-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10231209. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*