# New Technology for Tracking Proteins by Light and Electron Microscopy

> **NIH NIH R01** · OREGON HEALTH & SCIENCE UNIVERSITY · 2020 · $395,053

## Abstract

Project Summary
 Recent developments in fluorescence microscopy (FM), electron microscopy (EM), and correlative light
and EM (CLEM) offer unprecedented opportunities for illuminating cellular structures at the nanoscale. It is
now feasible to visualize and quantify the spatial organization of proteins and other macromolecules that
enable cells to sense and respond to their environment. However, these efforts are restricted by the shortage
of methods for attaching FM-, EM-, and CLEM-compatible reporter chemistries to target proteins.
 We will address this limitation by developing a new technology for labeling and imaging multiple cellular
proteins at once. Specifically, we propose to develop a set of heterodimeric coiled-coil tags that will
allow specific protein labeling for FM and EM. Our new versatile interacting peptide (VIP) tags will be
protein specific and cell compatible. Briefly, one coil (the “tag”) will be genetically-encoded as a fusion to a
protein of interest. After expression, the tagged protein will be subsequently labeled via heterodimer formation
with a high affinity (KD < 5 nM) “probe peptide”. We identified a set of coils that will self-sort into specific pairs,
which will enable up to four proteins to be labeled and imaged simultaneously. The reporter can be bright,
photostable fluorophores for FM or electron-dense nanoparticles for EM. In other words, VIP tags are modular,
enabling end-users to alternate between state-of-the art FM and EM imaging platforms. We will engineer VIP
tags to achieve the high labeling efficiency needed for quantitative analysis of multi-protein interactions.
 We will develop and validate our technology by investigating the iron-uptake machinery in cells. We will
use our technology to determine the differential protein trafficking and interactions of transferrin receptors 1
and 2 (TfR1 and TfR2). TfR1 is a well-studied transmembrane receptor and an ideal target for validating our
technology. Our studies of TfR2 will reveal new information on the sub-cellular distribution and trafficking of
this recently discovered iron-sensing receptor. We propose to complete two Aims. Aim 1. Develop and
validate a set of VIP tags for imaging proteins by FM. Aim 2. Use VIP tags for tracking receptor localization
and multi-protein interactions by EM.
 We believe that the VIP tags, once fully developed and validated, will be an ideal technology for
investigating the cellular organization of proteins with nanoscale precision. Furthermore, we believe this new
technology has broad utility for imaging cellular processes related to human health and disease.

## Key facts

- **NIH application ID:** 9983074
- **Project number:** 5R01GM122854-04
- **Recipient organization:** OREGON HEALTH & SCIENCE UNIVERSITY
- **Principal Investigator:** Kimberly Elizabeth Beatty
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $395,053
- **Award type:** 5
- **Project period:** 2017-08-01 → 2022-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9983074

## Citation

> US National Institutes of Health, RePORTER application 9983074, New Technology for Tracking Proteins by Light and Electron Microscopy (5R01GM122854-04). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9983074. Licensed CC0.

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