Abstract:  Transcriptional regulatory changes have been shown to contribute to phenotypic differences between species, but many questions remain about how gene expression evolves. In this talk, I will present the first comparative study of nascent transcription in primates. We used PRO-seq to map actively transcribing RNA polymerases in resting and activated CD4+ T-cells in multiple human, chimpanzee, and rhesus macaque individuals, with rodents as outgroups. This approach allowed us to directly measure active transcription separately from post-transcriptional processes. We observed general conservation in coding and non-coding transcription, punctuated by numerous differences between species, particularly at distal enhancers and non-coding RNAs. Transcription factor binding sites are a primary determinant of transcriptional differences between species. We found evidence for stabilizing selection on gene expression levels and adaptive substitutions associated with lineage-specific transcription. Finally, rates of evolutionary change are strongly correlated with long-range chromatin interactions. These observations clarify the role of primary transcription in regulatory evolution.