RNA-Based Fluorescent Sensor with RhoBAST.

Area of Science:

• Molecular biology
• Biochemistry
• Cell biology

Background:

• RNA-based biosensors are used for cellular target imaging.
• Existing RNA sensors suffer from low brightness, poor photostability, slow kinetics, and green background noise.
• Current sensors often fail to detect multiple targets or image within subcellular compartments.

Purpose of the Study:

• To develop a novel RNA-based sensor with improved performance for cellular imaging.
• To enable detection and subcellular imaging of multiple targets simultaneously.
• To overcome the limitations of existing RNA biosensors.

Main Methods:

• Development of a novel RNA-based sensor utilizing the RhoBAST fluorogenic RNA aptamer.
• Application of the sensor for detecting small molecules and calcium ions (Ca2+) in living cells.
• Creation of an orthogonal sensor for simultaneous subcellular imaging of multiple small molecules.

Main Results:

• The RhoBAST-based sensors exhibit high red fluorescence intensity, superior photostability, and fast activation kinetics.
• The developed sensors outperform previous fluorescent sensors in key performance metrics.
• Simultaneous subcellular imaging of multiple small molecules was achieved using an orthogonal sensor design.

Conclusions:

• A novel, highly bright, stable, and rapidly responsive red fluorescent RNA sensor has been developed.
• This sensor technology addresses critical limitations of existing RNA-based biosensors.
• The sensor enables advanced applications in subcellular multi-target imaging within living mammalian cells.