Probing Mitochondrial Heterogeneity and Dynamics During Cell State Switching Using Multiplexed, Environment-sensitive Fluorescent Dyes
Sufi O Raja1, Gandhi Sivaraman1, Ananya Mukherjee1, 3, Mamta Upadhyay,1 Sunny Kataria1,2, Colin Jamora1, Ravi Muddashetty1 and Akash Gulyani1
Institute for Stem Cell Biology & Regenerative Medicine1, National Centre for Biological Sciences2, TIFR Campus, GKVK Post, Bellary Road, Bengaluru-560065, Karnataka, India
Mitochondria are known as power house of the cell as they efficiently produce ATP through Oxidative Phosphorylation (OxPhos). Apart from this, mitochondria also play significant role in regulating cellular metabolism, calcium and ROS signaling as well as in programmed cell death. Despite decades of research in mitochondrial biology, precise information on spatio-temporal intracellular mitochondrial dynamics and its link to mitochondrial function is still limited. For example, recently it has been shown that cellular migration (with metastatic cancer cell lines) depends, in part, on mitochondrial motility and precise positioning within the cell. Similarly, morphology and activity of mitochondria are linked with maintenance of stemness as well as triggering of differentiation event. Therefore, unraveling the spatio-temporal localization as well as functional heterogeneity of mitochondria during various cellular states appears to be crucial for understanding the coherent behavior of cell. To better visualize the functional dynamics of mitochondrion, we have developed red-emitting, multi-functional, novel mitochondrial probes that are sensitive to local environment, specifically parameters like viscosity, pH, ROS, hypoxic condition, etc. The developed dyes have low toxicity and very high photo-stability, allowing their use in long term imaging. In this presentation, we will show these dyes have yielded new insights into differential ordering of mitochondria in Alzheimer patient's derived iPSCs compared to the normal individual. In a different example, we have also used our new dyes to probe mitochondrial heterogeneity within primary 'activated' cells during cell migration. These results would be placed in the context of our larger efforts to build new ways of probing 'cellular dynamics' with a focus on physico-chemical changes in the cell.