Structurally differentiated cis-elements that interact with PU.1 are functionally distinguishable in acute promyelocytic leukemia

Background: Transcription factor PU.1, a member of the ETS family, is a master regulator of myeloid differentiation whose functional disruption is often associated with acute myeloid leukemia (AML). Although much has been learned about PU.1 over the past decades, relatively little is known about cis-elements that interact with this factor under physiological or pathological conditions, especially in the whole-genome scale. We aimed to define the cistrome of PU.1 in acute promyelocytic leukemia (APL) cells and characterize the cis-elements bound by PU.1. Methods: Chromatin immunoprecipitation with specific antibody coupled with deep sequencing (ChIP-seq) was used to investigate the in vivo PU.1 binding sites at the whole-genome scale in APL-derived NB4 cells. The ChIP-quantitative (q)-PCR and luciferase reporter assays were used to validate the binding events and trans-activity, respectively. Various computational analyses, including motif mining, evolutionary conservation analysis and functional enrichment analysis, were performed to characterize the cis-elements that interacted with PU.1. Results: A total of 26,907 significantly enriched binding regions of PU.1 were identified under the false discovery rate 0.1% in NB4 cells. PU.1 bound to various types of genomic regions and acted as a promoter-enhancer dual binding transcription factor. Based on the sequence length and composition, two types of representative motifs were identified in PU.1 binding sites: a long and a short motif. The long motif, characterized by high sequence specificity and binding affinity, predominantly resided in the promoter-distal regions. In contrast, the short one, with strong evolutionary constraint, represented the primary PU.1 cis-elements in the promoter-proximal regions. Interestingly, the short one showed more preference to be correlated with the binding of other factors, especially PML/RARa. Moreover, genes targeted by both PU.1 and PML/RARa were significantly involved in categories associated with oncogenesis, hematopoiesis and the pathogenesis of acute myeloid leukemia. Conclusions: Our results demonstrate that structurally differentiated cis-elements that interact with PU.1 are functionally distinguishable in APL, suggesting that the sequence diversity of cis-elements might be a critical mechanism by which cells interpret the genome, and contribute to distinct physiological and/or pathological function.