Visualization of RNA-Quadruplexes in Live Cells.

Area of Science:

• Molecular Biology
• Cellular Imaging
• Biochemistry

Background:

• DNA and RNA quadruplexes are crucial G-rich nucleic acid structures involved in various cellular processes.
• Current methods for visualizing quadruplexes, primarily antibody-based, require cell fixation and permeabilization, limiting dynamic studies in live cells.
• There is a need for advanced probes enabling real-time, unbiased detection of quadruplexes within intact, living cells.

Purpose of the Study:

• To develop a novel fluorescent probe for the direct visualization of quadruplexes in live human and mouse cells.
• To overcome the limitations of existing immunodetection methods by enabling dynamic, label-free imaging of quadruplex formation.
• To introduce NaphthoTASQ (N-TASQ) as a next-generation tool for studying quadruplexes in their native cellular environment.

Main Methods:

• Synthesis and characterization of NaphthoTASQ (N-TASQ), a multiphoton turn-on fluorescent ligand.
• Application of N-TASQ to live human and mouse cells, requiring only a single-step incubation.
• Microscopy techniques to visualize N-TASQ fluorescence, indicating the presence and location of RNA quadruplexes.

Main Results:

• N-TASQ successfully detected RNA quadruplexes in live human and mouse cells without fixation, permeabilization, or mounting.
• The probe functions as a multiphoton turn-on fluorescent agent, enabling sensitive and specific imaging.
• This method provides a unique, unbiased visualization of quadruplexes, preserving cellular integrity and dynamics.

Conclusions:

• NaphthoTASQ (N-TASQ) represents a significant advancement in live-cell imaging of RNA quadruplexes.
• The probe's ability to detect quadruplexes in untreated cells offers unprecedented opportunities for studying their dynamic roles in biological processes.
• N-TASQ facilitates unbiased visualization, paving the way for new discoveries in nucleic acid research and cellular function.