# Mapping Transport Pathways through Nuclear Pores using 3D Super-Resolution Microscopy

> **NIH NIH R01** · TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR · 2021 · $297,000

## Abstract

PROJECT SUMMARY
 Nuclear pore complexes (NPCs) mediate the bi-directional transport of proteins, RNAs and ribonucleoprotein
complexes across the double-membrane nuclear envelope of eukaryotic cells. Consequently, NPCs are
essential for the ability of many biosynthetic, signaling and gene regulatory processes to maintain cellular health
and viability. Protein mislocalization due to recognition defects or altered NPC structure and function is linked
to diseases as diverse as primary biliary cirrhosis, amyotrophic lateral sclerosis (ALS), leukemias and cancers,
and Alzheimer's and Huntington's diseases. While the protein components of the NPC and many soluble nuclear
transport factors have been identified and extensively studied, the mechanism(s) by which bi-directional transport
occurs without clogging the pore remains unknown. The NPC is an octagonally symmetric approximately
cylindrical structure with an hourglass-shaped central pore that has an internal minimal diameter of ~50 nm in
humans. Occluding this pore and decorating its exits is a network of > 200 mobile intrinsically disordered
polypeptides. Thousands of phenylalanine-glycine (FG) repeat motifs within this disordered polypeptide network
(FG-network) are binding sites for the nuclear transport receptors (NTRs) that carry cargos through the NPC. At
steady-state, at least ~100 NTRs are asymmetrically distributed throughout the FG-network. The heterogeneous
and dynamic NTR/FG-network establishes a permeability barrier while simultaneously providing pathways for
the translocation of import and export complexes of a wide range of sizes, affinities and surface properties.
Multiple preferred paths through the ~50 nm diameter pore are predicted for typical cargo complexes of ~5-10
nm. The extent of overlap in such pathways and the possibility of dynamic regulation remains largely unexplored
due to the absence of technological tools to dissect these pathways with the necessary spatial and temporal
resolution. To address this deficiency, three-dimensional (3D) super-resolution single molecule fluorescence
microscopy and single particle tracking techniques will be used to explore various transport pathways to
determine the extent of their structural and functional intersections. The Specific Aims are: 1) to develop a fast
super-resolution 3D microscopy approach to characterize the translocation pathways through functional NPCs;
and 2) to develop 3D distribution maps for NTRs and cargos undergoing transport. Aim 1 seeks to bring together
multiple existing technologies to enable high precision 3D super-resolution microscopy on NPCs. Aim 2 seeks
to then apply this technology to establishing a map of various transport pathways through the FG-network. This
work will establish whether discrete transport pathways within the FG-network exist, and thus, enhance transport
efficiency by minimizing interactions between import and export cargos. While the primary goal of the proposed
work is to develop a compr...

## Key facts

- **NIH application ID:** 10077304
- **Project number:** 5R01GM126190-04
- **Recipient organization:** TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
- **Principal Investigator:** SIEGFRIED M MUSSER
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $297,000
- **Award type:** 5
- **Project period:** 2018-01-01 → 2022-09-21

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10077304

## Citation

> US National Institutes of Health, RePORTER application 10077304, Mapping Transport Pathways through Nuclear Pores using 3D Super-Resolution Microscopy (5R01GM126190-04). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10077304. Licensed CC0.

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