CellREADR: An ADAR-based RNA sensor-actuator device.

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.