This is the author accepted manuscript. It is currently under an indefinite embargo pending publication by NPG. ; Transcriptional dysregulation is associated with haematological malignancy. Although mutations of the key haematopoietic transcription factor PU.1 are rare in human acute myeloid leukemia (AML), they are common in murine models of radiation-induced AML, and PU.1downregulation and/or dysfunction has been described in human AML patients carrying the fusion oncogenes RUNX1-ETO and PML-RARA. To study the transcriptional programmes associated with compromised PU.1 activity, we adapted a Pu.1-mutated murine AML cell line with an inducible wild type PU.1. PU.1 induction caused transition from leukemia phenotype to monocytic differentiation. Global binding maps for PU.1, CEBPA and the histone mark H3K27Ac with and without PU.1 induction showed that mutant PU.1 retains DNA binding ability, but induction of wild type protein dramatically increases both the number and height of PU.1-binding peaks. Correlating ChIP Seq with gene expression data, we found that PU.1 recruitment coupled with increased histone acetylation induces gene expression and activates a monocyte/macrophage transcriptional programme. PU.1 induction also caused reorganisation of a subgroup of CEBPA binding peaks. Finally, we show that the PU.1 target gene set defined in our model allows stratification of primary human AML samples, shedding light on both known and novel AML subtypes that may be driven by PU.1 dysfunction. ; X18.1.1 cells were kindly donated by Dr Wendy Cook (LaTrobe University, Melbourne). MSCV-puro-PuER plasmid was kindly donated by Dr Peter Laslo (University of Leeds). ChIP sequencing was performed at the Genomics Core Facility, CRUK Cambridge Institute. Research in the G?ttgens laboratory is supported by Leukaemia and Lymphoma Research, the MRC, BBSRC, CRUK, Leukemia and Lymphoma Society, NIHR Cambridge Biomedical Research Centre and core infrastructure support by the Wellcome Trust to the Wellcome Trust and MRC Cambridge Stem Cell Institute and CIMR. JIS is supported by CRUK and the Raymond and Beverly Sackler Foundation.