Dynamic Mitochondrial Migratory Features Associated with Calcium Responses during T Cell Antigen Recognition

• Published in: The Journal of immunology (1950) Vol. 203; no. 3; pp. 760 - 768
• Main Authors: He, Luye, Raddatz, Andrew D, Zhou, Fangyuan, Hwang, Hyundoo, Kemp, Melissa L, Lu, Hang
• Format: Journal Article
• Language: English
• Published: United States 01-08-2019
• Subjects: Animals; Antigens, Differentiation, T-Lymphocyte - immunology; Calcium - metabolism; CD4-Positive T-Lymphocytes - immunology; Cell Line, Tumor; Encephalomyelitis, Autoimmune, Experimental - immunology; Humans; Hybridomas; Mice; Microfluidic Analytical Techniques - methods; Mitochondria - metabolism; Signal Transduction - immunology
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.1800299

A T cell clone is able to distinguish Ags in the form of peptide-MHC complexes with high specificity and sensitivity; however, how subtle differences in peptide-MHC structures translate to distinct T cell effector functions remains unknown. We hypothesized that mitochondrial positioning and associated calcium responses play an important role in T cell Ag recognition. We engineered a microfluidic system to precisely manipulate and synchronize a large number of cell-cell pairing events, which provided simultaneous real-time signaling imaging and organelle tracking with temporal precision. In addition, we developed image-derived metrics to quantify calcium response and mitochondria movement. Using myelin proteolipid altered peptide ligands and a hybridoma T cell line derived from a mouse model of experimental autoimmune encephalomyelitis, we observed that Ag potency modulates calcium response at the single-cell level. We further developed a partial least squares regression model, which highlighted mitochondrial positioning as a strong predictor of calcium response. The model revealed T cell mitochondria sharply alter direction within minutes following exposure to agonist peptide Ag, changing from accumulation at the immunological synapse to retrograde movement toward the distal end of the T cell body. By quantifying mitochondria movement as a highly dynamic process with rapidly changing phases, our result reconciles conflicting prior reports of mitochondria positioning during T cell Ag recognition. We envision applying this pipeline of methodology to study cell interactions between other immune cell types to reveal important signaling phenomenon that is inaccessible because of data-limited experimental design.