Visualizing Newly Synthesized Proteins and Their Degradation Dynamics by Using Long-Wavelength-Emitting Fluorescent...

Area of Science:

• Cell Biology
• Biochemistry
• Molecular Biology

Background:

• Maintaining protein homeostasis is crucial for cellular function.
• Existing methods for tracking protein synthesis often require cell fixation, limiting live-cell imaging.
• Live-cell imaging of protein synthesis and degradation dynamics is essential for understanding cellular processes.

Purpose of the Study:

• To develop a novel fluorescence imaging technique for monitoring newly synthesized proteins in living cells.
• To investigate the spatiotemporal dynamics of protein synthesis and degradation.
• To assess protein degradation capacity in senescent cells.

Main Methods:

• Utilized strain-promoted azide-alkyne cycloaddition (SPAAC) for live-cell protein labeling.
• Synthesized tetramethylrhodamine (TAMRA)-DBCO and silicon rhodamine (SiR)-DBCO probes.
• Employed fluorescence imaging to visualize newly synthesized proteins and their degradation over time.

Main Results:

• TAMRA-DBCO and SiR-DBCO successfully visualized newly synthesized proteins in living cells.
• Protein degradation was observed via both lysosomal and proteasomal pathways.
• Senescent cells exhibited diminished protein degradation ability compared to nonsenescent cells.

Conclusions:

• The developed SPAAC-based method allows for real-time monitoring of protein synthesis and degradation in living cells.
• This technique provides insights into the mechanisms of protein homeostasis and its dysregulation in cellular senescence.
• The findings offer a valuable tool for studying protein turnover and cellular health.