A Cluster of Enhancer Modules Directs Differential Myc Expression Along the Normal and Leukemic Hematopoietic Stem Cell Hierarchies

Large enhancer regions, known as "super-enhancers", have been described as critical regulatory units governing cell identity, but their actual contribution to stem cell-driven hierarchies remains poorly understood. The transcription factor MYC has critical roles in the regulation of hematopoietic stem cells (HSCs) and progenitors as well as in hematopoietic malignancies.

Here we show that a region previously identified as a "super-enhancer", located 1.7 Mb telomeric from Myc plays an essential role in regulating Myc expression throughout hematopoiesis. Deletion of this "super enhancer" in mice leads to a complete loss of Myc expression in HSC/progenitors, causing an accumulation of differentiation-arrested multipotent progenitors and loss of myeloid and B cells, similarly to the conditional deletion of the Myc gene in HSCs. This "Blood ENhancer Cluster" (BENC), which is functionally conserved in mice and humans, is comprised of distinct modules with selective activity and recruitment of such transcription factors as GFI1b, RUNX1 and MYB in different hematopoietic cell types. Analysis of mice carrying deletions of individual enhancer modules revealed a modular and combinatorial organization of BENC controlling Myc expression levels throughout most of the hematopoietic hierarchy. Additionally, we show that individual enhancer modules are activated at distinct stages during B cell development.

In a malignant setting, we demonstrate that BENC is essential for the maintenance of MLL-AF9 leukemia in mice. Furthermore, in human acute myeloid leukemia, BENC modules show an increased accessibility in primary leukemic stem cells compared to blasts. The accessibility of the module directing Myc expression in normal HSCs correlates with Myc expression and a favorable patient outcome. Altogether, these data indicate a major role of BENC in regulating MYC levels across both normal hematopoietic and leukemic hierarchies. The composition of BENC may serve as a blueprint for other enhancer clusters operating across cellular hierarchies in other tissue types.