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4D Microscopy of Yeast
Published on: April 28, 2019
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Protocol for 4D live fluorescence microscopy to image rapid cellular responses in yeast
Yaneris M Alvarado-Cartagena1, Christopher Huppenbauer2, Nava Segev1
1Department of Biochemistry and Molecular Genetics, College of Medicine, University of Illinois Chicago, Chicago, IL, USA.
STAR Protocols
|August 15, 2025
Summary
We developed a new live-cell imaging protocol to observe micro-endoplasmic reticulum (ER)-phagy in yeast. This method visualizes rapid protein degradation processes within seconds, offering insights into cellular stress responses.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Micro-autophagy is traditionally studied using fixed cells, limiting real-time observation of dynamic processes.
- Understanding the rapid mechanisms of cellular degradation, such as micro-endoplasmic reticulum (ER)-phagy, is crucial for cellular health.
Purpose of the Study:
- To present a novel protocol for visualizing micro-ER-phagy in live yeast cells.
- To enable the study of rapid cellular processes, including cargo engulfment and lysosomal degradation.
Main Methods:
- Induction of misfolded protein accumulation and selective transport for degradation under nutritional stress.
- Live-cell time-lapse confocal microscopy and 3D reconstruction techniques.
- Observation of cargo engulfment and lysosomal internalization in real-time.
Main Results:
- Demonstration of a protocol to track micro-ER-phagy in live yeast.
- Visualization of cargo engulfment and internalization into the lysosome occurring within seconds.
- Validation of the protocol for observing rapid cellular degradation events.
Conclusions:
- The developed protocol allows for real-time observation of micro-ER-phagy.
- This method provides a powerful tool for studying rapid cellular degradation and stress responses.
- The protocol can be adapted for investigating other dynamic cellular processes.
