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Updated: Jun 5, 2026
07:44
Application of Membrane and Cell Wall Selective Fluorescent Dyes for Live-Cell Imaging of Filamentous Fungi
Published on: November 28, 2019
Visual mapping of cell wall biosynthesis.
Yumiko Sakuragi1, Morten H H Nørholm, Henrik V Scheller
1The Department of Plant Biology and Biotechnology, University of Copenhagen, Copenhagen, Denmark.
Methods in Molecular Biology (Clifton, N.J.)
|January 12, 2011
Summary
Researchers visualized pectin and hemicellulose biosynthetic enzymes within the Golgi apparatus using advanced live-cell imaging. This study reveals protein interactions and membrane topology, advancing our understanding of cell wall polysaccharide synthesis.
Area of Science:
- Plant biology
- Cell biology
- Biochemistry
Background:
- Pectin and hemicellulose biosynthesis occur in the Golgi apparatus.
- This process involves complex spatial regulation and enzyme/protein complex formation.
Purpose of the Study:
- To visualize biosynthetic enzymes and proteins in subcellular compartments.
- To visualize protein-protein interactions of pectin biosynthetic enzymes in situ.
- To determine the membrane topology of pectin biosynthetic enzymes in the Golgi apparatus.
Main Methods:
- Heterologous expression of recombinant proteins tagged with fluorescent proteins.
- Live-cell imaging using confocal laser scanning microscopy (CLSM).
- Bimolecular fluorescence complementation (BiFC) assays.
Main Results:
- Efficient visualization of biosynthetic enzymes and proteins in subcellular compartments.
- In situ visualization of protein-protein interactions among pectin biosynthetic enzymes.
- Determination of membrane topology for pectin biosynthetic enzymes within the Golgi apparatus.
Conclusions:
- Heterologous expression, fluorescent tagging, and CLSM are effective for visualizing biosynthetic machinery.
- BiFC is a powerful tool for studying enzyme interactions and membrane topology in the Golgi.
- This research provides new insights into the spatial organization and regulation of cell wall polysaccharide biosynthesis.
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