Trypanosoma brucei, the causative agent of human and animal African trypanosomiasis (AKA sleeping sickness), cycles between a mammalian host and a tsetse fly vector. The parasite undergoes huge changes in morphology and metabolism during adaptation to each host environment. These changes are reflected in the different transcriptomes of parasites living in each host. We have shown that genetic and chemical inhibition of chromatin interacting bromodomain proteins in bloodstream parasites results in spontaneous differentiation to the insect stage. This compromises the immune evasion mechanisms of bloodstream parasites, allowing mice to clear parasites in models of infection. Bromodomain proteins localize to transcription start sites in bloodstream parasites, but whether the localization of bromodomain proteins changes as parasites differentiate from bloodstream to insect stages remains unknown. To address this question, we performed cleavage under target and release using nuclease (CUT&RUN) against bromodomain protein 3 (Bdf3) in parasites differentiating from bloodstream to insect forms. We found that Bdf3 occupancy at most loci increased at 3 h following onset of differentiation and decreased thereafter. These alterations in occupancy might facilitate an increase in transcript levels following the quiescent stage that precedes differentiation to the insect stage. Additionally, one site appears de novo during differentiation and lies proximal to the procyclin gene locus housing genes essential for remodeling surface proteins following transition to the insect stage. These studies indicate that occupancy of chromatin-interacting proteins is dynamic during life cycle stage transitions and provide the groundwork for future studies on how bromodomain proteins may facilitate transcriptional reprogramming during life cycle stage transitions in a highly diverged eukaryote.