Abstract
Abstract CD4+ T cells (T helper cells) are cytokine-producing adaptive immune cells that activate or regulate the responses of various immune cells. The activation and functional status of CD4+ T cells is important for adequate responses to pathogen infections but has also been associated with auto-immune disorders and survival in several cancers. In the current study, we carried out a label-free high-resolution FTMS-based proteomic profiling of resting and T cell receptor-activated (72h) primary human CD4+ T cells from peripheral blood of healthy donors as well as SUP-T1 cells. We identified 5,237 proteins, of which significant alterations in the levels of 1,119 proteins were observed between resting and activated CD4+ T cells. We confirmed several known T-cell activation-related processes such as IL-2 response, metabolic and signaling changes, cell cycle induction, differentiation into effector cells among others. Several stimulatory/inhibitory immune checkpoint markers were altered considerably between resting and activated CD4+ T cells. Network analysis identified several known regulatory hubs of CD4+ T cell activation, including IFNG, IRF1, FOXP3, AURKA, and novel hubs such as RIOK2. Comparison of primary CD4+ T cell proteomic profiles with human lymphoblastic cell lines revealed a substantial overlap, while comparison with mouse CD+ T cell data suggested interspecies proteomic differences. Competing Interest Statement The authors have declared no competing interest. * Abbreviations AA Amino Acid FDR False Discovery Rate GO Gene Ontology KEGG Kyoto Encyclopedia of Genes and Genomes MACS Magnetic-Activated Cell Sorting NCBI National Center for Biotechnology Information PBMC Peripheral Blood Mononuclear Cell PSM Peptide Spectrum Match SDS Sodium Dodecyl Sulphate TCR T cell receptor TEABC Triethyl ammonium bicarbonate Th cells T helper cells TPCK L-(tosylamido-2-phenyl) ethyl chloromethyl ketone