Biological systems are inherently heterogeneous. In vertebrates, the immune system protects the body with diverse cells of specialized functions. CD8+ T cells are a major component of adaptive immunity, contributing to pathogen clearance and tumor control. Naïve CD8+ T cells are a heterogeneous population with subsets of distinct kinetics and functions upon activation. It is increasingly clear that their divergent post-stimulation fates can be established at naïve stage, but gene expression programs differentiating the naïve subsets remain to be elucidated. Here, I analyzed a diverse set of RNA-seq and ATAC-seq profiles in naïve CD8+ T cell subsets, where extensive differences in gene expression and chromatin accessibility were observed. Leveraging those profiles, I used a powerful network inference algorithm, Inferelator, to construct a transcriptional regulatory network in naïve CD8+ T cells. Key TFs promoting each naïve subset were identified, which not only included known TFs, but also implicated novel roles for additional factors. Taken together, this work investigated transcriptional control on naïve CD8+ T cell subsets, revealing regulatory circuits that preprogram their distinct fates. In immune-related diseases, dysregulation of immune cells may display to different degrees across patient cohorts, introducing additional layers of heterogeneity that are important to decipher for elucidating disease mechanisms. Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a serious and poorly understood disease, characterized by symptom exacerbation following exertion. To understand immune dysregulation in ME/CFS, we used single-cell RNA-seq to examine immune cells in patients and controls at baseline and after symptom provocation. Monocyte dysregulation was identified as a prominent feature of immune dysregulation in ME/CFS, with patterns suggestive of inappropriate differentiation and migration to tissue, both at baseline and after symptom provocation. Importantly, mixed monocyte populations, including diseased and more normal cells, were identified in patients, and the fraction of diseased cells correlated with metrics of disease severity. Taken together, this work investigated immune dysregulation in ME/CFS at single cell resolution, and identified heterogeneous monocyte dysregulation in ME/CFS patients. Overall, this dissertation presents two projects that examine heterogeneity of immune cells: identifying regulatory circuits differentiating naïve CD8+ T cell subsets, and unraveling immune dysregulation in ME/CFS.