Metabolic Dynamics of In Vitro CD8+ T Cell Activation

CD8+ T cells detect and kill virally infected or cancerous cells. When activated from their naïve state, T cells undergo a complex transition, including major metabolic reprogramming. Detailed resolution of metabolic dynamics of a wide range of metabolites is needed to advance the field of immunometabolism. Here we outline methodologies that when utilized in parallel achieve broad coverage of the metabolome. Specifically, we used a combination of 2 FIA and 3 LC methods in combination with positive and negative mode high resolution MS to study the transition from naïve to effector T cells with fine-grained time resolution. Depending on the method, between 54% and 98% of cellular metabolites change in a time-dependent manner, with the major changes in both polar metabolites and lipids occurring in the first 48 h. Statistical analysis highlighted remodeling of the polyamine biosynthesis pathway, with marked differences in the dynamics of precursors, intermediates, and cofactors. Moreover, phosphatidylcholines, the major class of membrane lipids, underwent a drastic shift in acyl chain composition with polyunsaturated species decreasing from 60% to 25% and specifically depleting species containing a 20:4 fatty acid. We hope that these MS methodologies and data set of over 11000 features and 9 time points will be a useful resource for future work.