SignificanceHematopoietic stem and progenitor cells (HSPCs) require tight transcriptional regulation. In-depth knowledge on how epigenetic mechanisms regulate chromatin landscape in order to control gene expression, particularly in response to external stimuli, will be key to understand the pathogenesis of hematologic disorders and improve blood stem cell-based therapies. Prostaglandin E2 (PGE2) and its metabolically resistant analog 16,16-dimethyl-PGE2 (dmPGE2) were previously identified as a potent external stimulus to enhance HSPC engraftment. PGE2has been evaluated in various clinical trials to improve engraftment of stem cells. We identified that dmPGE2mediates chromatin flexibility at HSPC-specific enhancers through histone-variant H2A.Z acetylation to promote master transcription factor (TF) binding and reinforce expression of genes involved in stem cell fate and engraftment.