Hematopoietic Stem and Progenitor Cell Proliferation and Differentiation Requires the Trithorax Protein Ash2l In Vivo

Gene transcription is controlled by post-translational modifications of core histones. Methylation of lysine 4 of histone H3 (H3K4), together with acetylation of H3K27, is closely associated with open chromatin and gene transcription. H3K4 methylation is catalyzed by KMT2 lysine methyltransferases that include the mixed-lineage leukemia 1-4 (MLL1-4) and SET1A and B enzymes. For efficient catalysis all six require a core complex of four proteins, WDR5, RBBP5, ASH2L, and DPY30.

In a newly-generated pIC-inducible Mx1-Cre mouse model, we report that targeted disruption of Ash2l in the hematopoietic system results in death of the mice due to a rapid loss of mature hematopoietic cells in the bone marrow and the spleen, associated with a significant reduction of the RBC, WBC and platelets. However, Lin-Sca1+Kit+ (LSK) cells, which are highly enriched in hematopoietic stem and multi-potent progenitor cells, were significantly increased in the bone marrow (18-fold increase). This increase was associated with a 15-fold loss of the myeloid progenitor (MP) population (p < 0.001). In detail, total cell number of granulocyte-macrophage progenitors (GMPs), megakaryocyte-erythrocyte progenitors (MEPs) and common-myeloid progenitors (CMPs) was significantly reduced, while the relative proportion remained essentially unchanged. In LSK cells, Ash2l loss interfered with H3K4 methylation and resulted in broadly deregulated gene expression, including down-regulation of mitosis-associated genes. As a consequence, LSK cells accumulated in the G2/M-phase of the cell cycle and were unable to proliferate and form colonies in semi-solid medium. Lentiviral complementation with human ASH2L in the lineage-negative population partly but not entirely restored colony forming capacity, suggesting that ASH2L-mediated shaping of hematopoietic differentiation is a very early process. In addition, we demonstrated that those genes that harbor H3K4me3 and H3K27ac marks at their promoters were preferentially transcriptionally down-regulated. In contrast, up-regulated genes were characterized by low H3K4me3 and H3K27ac. Thus, these findings suggest that H3K4me3 and H3K27ac are not essential for transcription of all genes and that other, so far unidentified chromatin marks, allow recruitment of the RNA polymerase complex and transcription. Together, our data identify Ash2l as a novel essential factor for balanced gene expression and for hematopoietic stem and multi-potent progenitor cell physiology. These data may have important implications for future therapies of hematopoietic malignancies, including epigenetic targeting of malignant stem cells.