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Updated: Jun 17, 2025
14:08
Study of Protein-protein Interactions in Autophagy Research
Published on: September 9, 2017
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Studying Selective Autophagy of Protein Aggregates Using Particles Induced by Multimerization (PIMs)
Giel Korsten1, Lukas C Kapitein2,3
1Cell Biology, Neurobiology and Biophysics, Department of Biology, Faculty of Science, Utrecht University, Utrecht, The Netherlands.
Methods in Molecular Biology (Clifton, N.J.)
|August 8, 2024
Summary
This study presents a protocol for the PIM assay, enabling controlled formation and visualization of protein aggregates for studying aggrephagy and cellular homeostasis.
Area of Science:
- Cell Biology
- Molecular Biology
- Autophagy Research
Background:
- Selective autophagy, or aggrephagy, is crucial for cellular homeostasis by clearing protein aggregates.
- Studying aggrephagy is difficult due to infrequent events and lack of control over aggregate formation.
- Previous work introduced the PIM (particles induced by multimerization) assay for chemically induced aggregate formation.
Purpose of the Study:
- To provide a detailed protocol for utilizing the PIM assay.
- To enable controlled and specific induction of protein aggregates for aggrephagy studies.
- To facilitate the visualization of aggregate clearance in live and fixed cells.
Main Methods:
- Utilizing two variants of the PIM assay for aggregate induction.
- Employing chemically induced, fluorescently labeled protein aggregates.
- Using pH-sensitive fluorescent proteins (e.g., GFP, mKeima) for visualization.
- Applying the protocol to both live and fixed cell imaging.
Main Results:
- The PIM assay allows for controlled formation of protein aggregates.
- Aggregates formed via the PIM assay are recognized and degraded by the selective autophagy machinery.
- The use of pH-sensitive fluorescent proteins enables direct visualization of aggregate clearance.
- The protocol is effective in both live and fixed cell imaging.
Conclusions:
- The PIM assay provides a robust method for studying aggrephagy.
- This protocol enhances the ability to investigate cellular mechanisms of protein aggregate clearance.
- The PIM assay is a valuable tool for research in cellular homeostasis and autophagy.
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