Infection triggers large-scale changes in the phenotype and function of T cells that are critical for immune clearance, yet the gene regulatory mechanisms that control these changes are largely unknown. Using ChIP-seq for multiple histone PTM combinations, and HiC for mapping higher order chromatin structures, we mapped the dynamics of ~26,000 putative CD8⁺ T cell transcriptional enhancers (TEs) differentially utilised within ex vivo isolated naïve, effector and memory virus-specific CD8⁺ T cell during differentiation. We show that virus-specific T cell differentiation is underpinned by a combination of de novo TE gain and loss, maturation of the chromatin state via activation of “poised” TEs already present within naïve T cells, and specific transcription factor targeting of TEs that exhibit a non-canonical (H3K4me3⁺) chromatin signature upon differentiation. Our data also demonstrate that the chromatin structures within naïve CD8+ T cells are pre-configured at both the level of histone PTMs and higher order chromatin contacts. This genomic pre-configuration enables targeted maturation of lineage-specific genomic elements upon T cell activation, thus implying that the outcome of CD8⁺ T cell differentiation is largely pre-determined. These data have implications for the molecular events, and their regulation, that occur during the generation of effective T cell responses and establishment of immunological memory.