# Dissecting signaling in vivo via precise control and visualization of protein activity

> **NIH NIH R35** · UNIV OF NORTH CAROLINA CHAPEL HILL · 2022 · $810,270

## Abstract

Abstract
An ongoing revolution in microscopy has revealed previously inaccessible aspects of signaling. These include
organization of signaling proteins on dynamic cytoskeletal elements, subtle but important variations in
activation kinetics, and information encoded in the oscillation frequencies of signaling circuits. We will explore
these regulatory mechanisms by developing novel approaches to visualize and control protein activity, and by
using them together in the same cell. Novel image analysis methods and computational modeling applied to
multiplexed imaging data will reveal causal connections between signaling events, including those dependent
on nested feedback loops. We will pursue new methods to produce biosensors bright enough to study the
conformational changes of individual molecules in living cells. On podosomes and adhesion complexes, where
important dynamic structures are organized on a scale below the diffraction limit, we will use these new
methods to examine the opening of stretch-activated binding sites in real time. Proteins will be controlled with
light or small molecules using engineered domains inserted away from the active site, where they can
allosterically control activity without perturbing normal interactions. We will study three model systems that are
well suited to extract basic principles re the spatio-temporal regulation of signaling: 1) In 'frustrated
phagocytosis' macrophages attempt to engulf micropatterned circles. The phagocytic apparatus they build
around the circles has precise, reproducible geometry, which will enable us to use quantitative image analysis
and simultaneous protein control/visualization to develop mathematical models. 2) We will study how cancer
cells move on aligned collagen "superhighways" to find blood vessels. There we will ask how cells use
localized signaling to sense and respond to anisotropic mechanical stresses. 3) Finally, cell
protrusion/retraction will be used to examine the integration of chemical gradients, mechanical force, and
cytoskeletal dynamics. We hope that the behavior of these model systems will reveal ubiquitous signaling
mechanisms, and that the new tools can help others explore diverse questions in cell biology.

## Key facts

- **NIH application ID:** 10406708
- **Project number:** 2R35GM122596-06
- **Recipient organization:** UNIV OF NORTH CAROLINA CHAPEL HILL
- **Principal Investigator:** Klaus M. Hahn
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $810,270
- **Award type:** 2
- **Project period:** 2017-04-01 → 2027-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10406708

## Citation

> US National Institutes of Health, RePORTER application 10406708, Dissecting signaling in vivo via precise control and visualization of protein activity (2R35GM122596-06). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10406708. Licensed CC0.

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