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Updated: May 30, 2025

Real-time Imaging of Single Engineered RNA Transcripts in Living Cells Using Ratiometric Bimolecular Beacons
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CellREADR: An ADAR-based RNA sensor-actuator device.

Xiaolu Yang1, Kehali Woldemichael1, Xiao Guo2

  • 1Department of Neurobiology, Duke University School of Medicine, Durham, NC, United States.

Methods in Enzymology
|January 27, 2025
PubMed
Summary
This summary is machine-generated.

CellREADR is a novel RNA sensor technology that detects specific cellular RNAs to control protein production. This programmable system enables monitoring and manipulation of cells in diverse species and tissues.

Keywords:
ADARCell typeProgrammableRNA actuatorRNA editingRNA sensor

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genetics

Background:

  • RNAs are crucial for gene regulation and cell differentiation.
  • Sensing and responding to RNA profiles in living cells is vital for biological and medical applications.
  • Existing technologies lack the programmability and versatility needed for broad cellular monitoring.

Purpose of the Study:

  • To introduce CellREADR, a programmable RNA sensor-actuator technology.
  • To demonstrate CellREADR's ability to couple RNA detection with effector protein translation.
  • To provide a detailed protocol for implementing CellREADR in cell culture and animal models.

Main Methods:

  • CellREADR utilizes an adenosine deaminase acting on RNA (ADAR)-mediated A-to-I editing mechanism.
  • The system comprises a 5' sensor region complementary to a target RNA and a 3' protein payload.
  • Payload translation is activated by RNA binding, which removes a stop codon via ADAR editing.

Main Results:

  • CellREADR was successfully designed, cloned, and validated in mammalian cell cultures.
  • The technology demonstrated effective monitoring and manipulation of cellular activity based on RNA detection.
  • In vivo implementation using AAV-based delivery in brain tissue was established.

Conclusions:

  • CellREADR represents a new generation of programmable RNA devices for monitoring and manipulating animal cells.
  • The technology is simple, versatile, and generalizable across tissues and species.
  • Detailed protocols facilitate the implementation of CellREADR in various experimental settings.