Genome-scale perturb-seq in primary human CD4+ T cells reveals genes regulating T cell programs and human immune traits.
CD4+ T cells orchestrate immune responses against infection and cancer, yet a comprehensive map of their gene regulatory networks has been lacking. Perturb-seq enables systematic mapping of such networks, but large-scale application in primary human T cells has been challenging. Here, we developed a scalable, probe-based Perturb-seq platform and performed genome-scale perturbations of all expressed genes across 22 million primary CD4+ T cells from 4 donors under three stimulation conditions. We demonstrate the utility of this dataset in three ways. First, we identify previously unrecognized regulators of the two important immune cytokines, IL-10 and IL-21. Second, we show that ex vivo perturbation signatures can model T cell states observed in population-scale atlases, revealing regulators of helper T cell polarization and T cell aging. Third, we leverage regulatory relationships to mechanistically explain gene-trait associations in immune-related diseases, nominating genes for uncharacterized immune functions. Thus genome-scale perturb-seq in primary human T cells provides a foundational resource for decoding immune variation in humans and reprogramming T cell states for next-generation precision therapies.