Metabolic quiescence of naïve-like memory T cells precedes and maintains antigen-specific T-cell memory
Metabolic activity shapes cell fate but remains challenging to capture in vivo with high resolution. We performed longitudinal metabolic and phenotypic profiling of human antigen-specific CD8+ T cells after yellow-fever vaccination on the single-cell level. During the acute phase, T cells upregulated glycolysis to fuel anabolic needs of proliferation, but predominantly used oxidative phosphorylation for energy production, as assessed via protein translation rates. Central memory T cells were the most active subset, while effector cells underwent metabolic shut-down. In contrast, weakly differentiated naïve-like memory T cells showed minimal activity, relying solely on oxidative phosphorylation already during the acute phase. Reinforcing the link between cellular quiescence and longevity, naïve-like memory cells were preferentially maintained even 26 years post vaccination. This association between differentiation degree and metabolic activity was conserved after SARS-CoV-2 vaccination and in two murine infection models. Our study dissects the metabolic profile of antigen-specific T-cell responses ex vivo, highlighting quiescence as a key feature for long-term immunological memory formation in humans. Overall design: Yellow Fever Virus (YFV)-specific CD8+ T cells from human blood before and after YFV-vaccination were were identified by DNA-baroded pMHC multimers (Dextramers) in three independent experiments. Cells were stained with DNA-barcoded CITEseq antibodies for surface markers (among them CD45RA,CD62L,CD95), cells from 27 different donors and  11 time-points were labelled with TotalSeq-C anti-human hashtag antibodies and samples were pooled for further processing. CD8+ T cells specific for eight different  YFV-epitopes (including the HLA-A2-restricted NS4B214 epitope)  and five different control-virus epitopes, as well as cells without dextramer staining were sort-isolated by FACS. Cells were subjected to 5' scRNAseq on a 10x Genomics platform including VDJ-sequencing and  CITEseq for Dextramers, hashtag and surface marker antibodies.