Extremely Sensitive Genetically Encoded Temperature Indicator Enabling Measurement at the Organelle Level
Shun-Ichi Fukushima1, Tetsuichi Wazawa1, Kazunori Sugiura1
1SANKEN, The University of Osaka, Ibaraki, Osaka 567-0047, Japan.
ACS Sensors
|July 23, 2024
Summary
Researchers developed gMELT, a genetically encoded fluorescent temperature indicator, for precise intracellular thermogenesis monitoring. This new tool enhances sensitivity and organelle targeting for improved subcellular temperature measurements.
Area of Science:
- Cellular Biology
- Biophysics
- Biochemistry
Background:
- Intracellular temperature is crucial for biochemical reactions.
- Existing genetically encoded fluorescent temperature indicators (GETIs) require improved sensitivity and organelle targeting.
- Accurate visualization of subcellular temperature changes is needed.
Purpose of the Study:
- To develop a novel genetically encoded fluorescent temperature indicator (GETI) with enhanced sensitivity and organelle localization capabilities.
- To enable precise monitoring of intracellular thermogenesis at the subcellular level.
- To investigate the influence of macromolecular crowding on temperature indicator performance.
Main Methods:
- Development of gMELT, a Förster resonance energy transfer (FRET)-based indicator using cyan and yellow fluorescent proteins fused to temperature-sensitive domains.
- Expression of gMELT in cytoplasm and targeted delivery to endoplasmic reticulum (ER) and mitochondria using localization signals.
- Utilizing gMELT to detect chemically induced thermogenesis in cellular compartments.
- Assessing the impact of macromolecular crowding on gMELT fluorescence.
Main Results:
- gMELT demonstrated higher temperature sensitivity in cellular compartments compared to previous GETIs.
- Targeted gMELT variants successfully visualized thermogenesis in the ER and mitochondria.
- Observed temperature changes in organelles were comparable to previous findings.
- Macromolecular crowding was shown to subtly influence gMELT fluorescence readout.
Conclusions:
- gMELT provides a sensitive and versatile tool for measuring intracellular temperature changes in specific organelles.
- The development of gMELT advances the study of intracellular thermogenesis.
- Accounting for factors like macromolecular crowding is essential for accurate interpretation of thermogenesis data.
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