# Designing a new class of fluorescent reporters for imaging dynamic cell signaling in live animals

> **NIH NIH R35** · UNIVERSITY OF CALIFORNIA, SAN FRANCISCO · 2024 · $591,712

## Abstract

Project Summary/Abstract
Spatial organization and dynamics are the essential features of living systems. Visualizing compartmentalized
signaling dynamics is thus critical to understand biological processes. We focus on rational design of new
classes of signaling reporters that achieve large dynamic range, high brightness, fast kinetics and high spatial
resolution. These reporters are designed based on 1) new physical principles such as multivalent interaction-
driven protein phase separation forming intensely bright GFP droplets upon activation of kinases (the reporter
is named SPARK for Separation of Phases-based Activity Reporter of Kinase); 2) FlipGFP, a fluorogenic
protease reporter by flipping a beta-strand of GFP; 3) SURF (Split UnaG-based Reversible and Fluorogenic)
protein-protein interaction (PPI) reporter by splitting and engineering a new fluorescent protein scaffold UnaG
by structure-based design and directed evolution. Our primary goal in designing the new classes of signaling
reporters is to enable us in visualizing compartmentalized signaling dynamics in living systems, including PPIs
that determine specificity of signal transduction, and effectors such as kinases and proteases that regulate all
major signaling pathways. If we understand spatial organization and temporal dynamics of signaling, we will be
able to better understand molecular mechanisms of biological processes, laying the foundation for
understanding disease mechanisms and identifying drugs for the treatment of various diseases including
infectious diseases and cancer. While we have made considerable progress, in the next five years, we will
address three major unsolved issues: imaging endogenous PPIs, multicolor reporters of kinases and
proteases. We will engineer and demonstrate these multicolor and ultrasensitive reporters for multiplex
imaging of signaling network in living systems.

## Key facts

- **NIH application ID:** 10830584
- **Project number:** 2R35GM131766-06
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
- **Principal Investigator:** Xiaokun Shu
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $591,712
- **Award type:** 2
- **Project period:** 2019-06-01 → 2029-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10830584

## Citation

> US National Institutes of Health, RePORTER application 10830584, Designing a new class of fluorescent reporters for imaging dynamic cell signaling in live animals (2R35GM131766-06). Retrieved via AI Analytics 2026-06-12 from https://api.ai-analytics.org/grant/nih/10830584. Licensed CC0.

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