Integrative Proteomics and Phosphoproteomics Profiling Reveals Dynamic Signaling Networks and Bioenergetics Pathways Underlying T Cell Activation

Abstract

SUMMARY The molecular circuits by which antigens activate quiescent T cells remain poorly understood. We combined temporal profiling of the whole proteome and phosphoproteome via multiplexed isobaric labeling proteomics technology, computational pipelines for integrating multi-omics datasets, and functional perturbation to systemically reconstruct regulatory networks underlying T cell activation. T cell receptors activated the T cell proteome and phosphoproteome with discrete kinetics, marked by early dynamics of phosphorylation and delayed ribosome biogenesis and mitochondrial activation. Systems biology analyses identified multiple functional modules, active kinases, transcription factors and connectivity between them, and mitochondrial pathways including mitoribosomes and complex IV. Genetic perturbation revealed physiological roles for mitochondrial enzyme COX10-mediated oxidative phosphorylation in T cell quiescence exit. Our multi-layer proteomics profiling, integrative network analysis and functional studies define landscapes of the T cell proteome and phosphoproteome and reveal signaling and bioenergetics pathways that mediate lymphocyte exit from quiescence.

Hippo/Mst signalling couples metabolic state and immune function of CD8α+ dendritic cells

“…Whole and phosphoproteome profiling was performed as recently described 34 . CD8α + and CD8α − DCs were sorted from WT spleen as described above.…”

“…The analysis was performed by our in-house JUMP search engine which has been used in the data processing of numerous publications 34 . Briefly, acquired MS/MS raw files were converted into mzXML format and searched by the JUMP algorithm against a composite target/decoy database to estimate FDR.…”

MenTORing Immunity: mTOR Signaling in the Development and Function of Tissue-Resident Immune Cells

“…This is broadly consistent with the observed importance of mTORC1 in CD8 + Teff cell activation and in the differentiation and activation of some Th cell subsets. Proteomic analysis of Raptor-deficient T cells shows that expression of key transcription factors in metabolic reprogramming (MYC, YY1, GABPA, SREBF1) and components of the translational machinery are dependent upon mTORC1 (Tan et al, 2017), highlighting the central role of mTORC1 in anabolic T cell growth. Moreover, in cytotoxic CD8 + T cells, proteomic analyses have shown that rapamycin treatment to inhibit mTORC1 causes a reduction in cell size and protein content associated with changes in abundance (up as well as down) of approximately 10% of the proteome, including reductions in glucose transporters, and enzymes in the glycolysis and cholesterol synthesis metabolic pathways, as well as of the PtdIns(3,4,5)P3 phosphatase PTEN, and key effector proteins such as IFN-γ, perforin granzyme, and TNFα (Hukelmann et al, 2016).…”

The metabolic spectrum of memory T cells

“…10,14,15 Although mitochondrial ATP production appears dispensable for effector T-cell proliferation at later time points, 16 OXPHOS is upregulated after T-cell activation and has pivotal roles in the activation and development of effector T cells. 6 A number of mitochondrial pathways are engaged or remodeled early after activation, 17 including mitochondrial ROS to augment nuclear factor of activated T cells activation 18 and one carbon metabolism to support purine and thymidine synthesis. 19,20 Costimulation can contribute to enhanced mitochondrial respiratory functions early after activation.…”