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Related Experiment Video

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An Optogenetic Method to Control and Analyze Gene Expression Patterns in Cell-to-cell Interactions
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Red Photoactivatable Genetic Optical-Indicators.

Wessal Hussein1, Shai Berlin1

  • 1Department of Neuroscience, The Ruth and Bruce Rappaport Faculty of Medicine, Technion - Israel Institute of Technology, Haifa, Israel.

Frontiers in Cellular Neuroscience
|June 18, 2020
PubMed
Summary
This summary is machine-generated.

Researchers developed novel photoactivatable red-emitting genetically-encoded calcium indicators (PA-R-GECIs) to overcome signal overlap in dense tissues. This innovation enables precise calcium signaling studies in individual neurons.

Keywords:
GECI genetically encoded Ca2+ indicatorcalciumoptical toolsphotoactivatablered fluorescent protein

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Area of Science:

  • Neuroscience
  • Biophotonics
  • Molecular Biology

Background:

  • Genetically-encoded calcium indicators (GECIs) are crucial for studying calcium signaling in neurons.
  • Red-fluorescent protein (RFP)-based GECIs (R-GECIs) offer advantages like deeper imaging and reduced photodamage.
  • Signal overlap from neighboring cells in dense tissues limits single-cell resolution.

Purpose of the Study:

  • To develop the first photoactivatable red-emitting GECIs (PA-R-GECIs).
  • To enable targeted activation of fluorescence for improved single-cell resolution in dense tissues.
  • To engineer a novel photoactivatable red fluorescent protein (PA-RFP).

Main Methods:

  • Exploited Ca2+-independent fluorescence artifacts in an emerging R-GECI.
  • Rationally engineered functional PA-R-GECIs based on observed artifacts.
  • Developed a novel PA-RFP, PA-mRuby3.

Main Results:

  • Successfully engineered several functional PA-R-GECIs.
  • Demonstrated the ability to activate fluorescence on demand in specific cells.
  • Created a new PA-RFP, PA-mRuby3, for broader applications.
  • Overcame the limitation of signal overlap in dense neural tissues.

Conclusions:

  • The developed PA-R-GECIs address a critical need for precise calcium imaging in dense tissues.
  • These tools enhance the ability to study neuronal calcium dynamics at the single-cell level.
  • The engineered PA-mRuby3 expands the toolkit for photoactivatable fluorescent proteins.