Multiomic profiling of chronically activated CD4+ T cells identifies drivers of exhaustion and metabolic reprogramming

by Matthew L. Lawton, Melissa M. Inge, Benjamin C. Blum, Erika L. Smith-Mahoney, Dante Bolzan, Weiwei Lin, Christina McConney, Jacob Porter, Jarrod Moore, Ahmed Youssef, Yashasvi Tharani, Xaralabos Varelas, Gerald V. Denis, Wilson W. Wong, Dzmitry Padhorny, Dima Kozakov, Trevor Siggers, Stefan Wuchty, Jennifer Snyder-Cappione, Andrew Emili

Repeated antigen exposure leads to T-cell exhaustion, a transcriptionally and epigenetically distinct cellular state marked by loss of effector functions (e.g., cytotoxicity, cytokine production/release), up-regulation of inhibitory receptors (e.g., PD-1), and reduced proliferative capacity. Molecular pathways underlying T-cell exhaustion have been defined for CD8+ cytotoxic T cells, but which factors drive exhaustion in CD4+ T cells, that are also required for an effective immune response against a tumor or infection, remains unclear. Here, we utilize quantitative proteomic, phosphoproteomic, and metabolomic analyses to characterize the molecular basis of the dysfunctional cell state induced by chronic stimulation of CD4+ memory T cells. We identified a dynamic response encompassing both known and novel up-regulated cell surface receptors, as well as dozens of unexpected transcriptional regulators. Integrated causal network analysis of our combined data predicts the histone acetyltransferase p300 as a driver of aspects of this phenotype following chronic stimulation, which we confirmed via targeted small molecule inhibition. While our integrative analysis also revealed large-scale metabolic reprogramming, our independent investigation confirmed a global remodeling away from glycolysis to a dysfunctional fatty acid oxidation-based metabolism coincident with oxidative stress. Overall, these data provide both insights into the mechanistic basis of CD4+ T-cell exhaustion and serve as a valuable resource for future interventional studies aimed at modulating T-cell dysfunction.