Cell Stress | Published 08.07.2025. | doi: 10.15698/cst2025 07.307
Aman Munirpasha Halikar, Aneesh Chandrasekharan, Asha Lekshmi, Aswathy Sivasailam, Jain Tiffee P J, Shivanshu Kumar Tiwari, Aijaz Ahmad Rather and TR Santhoshkumar*
Genetically identical cells in a population show cell-to-cell variability because of fluctuation in transcription. epigenetics. post translational modifications. and stochastic or extrinsically triggered non genetic alterations. The change in the interaction state of proteins also emerges as an additional layer of cell signaling that influences cell cycle and cell death. However. the interrelation between cell cycle progression and cell death under the influence of spatio-temporal changes in protein protein interaction is difficult to demonstrate in growing cells. This requires tools for cell cycle phase-resolved visualization of macromolecular interac tions in live cells. We describe an approach to visualize the interaction of pro- and anti-death signaling partners. Bax and Bcl-xL during cell cycle progression and cell death in live cells. Cells were stably expressed with Bax and Bcl-xL with FRET pairs and real-time cell cycle indicator probes. Acceptor photobleaching and Fluorescence lifetime imaging revealed interaction dynamics between Bax and Bcl-xL in isogenic stable cells. While Bcl-xL inhibited cell cycle progression. Bax promoted the cell cycle. The study highlighted an increased Bax-Bcl-xL interaction in the G1 phase compared to the non-G1 phase. Increased interaction is seen under stressed conditions and Bax-activated cells with FLIM-FRET, highlighting the nature of Bax-Bcl-xL interaction during cellular stress. In conclusion. our study explains Bax-Bcl-xL interaction dynamics in real-time and the potential utility of the approach to study macromolecular interactions along with cell cycle and cell death.
