Hydroxymethylation at Gene Regulatory Regions Directs Stem/Early Progenitor Cell Commitment during Erythropoiesis

Hematopoietic stem cell differentiation involves the silencing of self-renewal genes and induction of a specific transcriptional program. Identification of multiple covalent cytosine modifications raises the question of how these derivatized bases influence stem cell commitment. Using a replicative primary human hematopoietic stem/progenitor cell differentiation system, we demonstrate dynamic changes of 5-hydroxymethylcytosine (5-hmC) during stem cell commitment and differentiation to the erythroid lineage. Genomic loci that maintain or gain 5-hmC density throughout erythroid differentiation contain binding sites for erythroid transcription factors and several factors not previously recognized as erythroid-specific factors. The functional importance of 5-hmC was demonstrated by impaired erythroid differentiation, with augmentation of myeloid potential, and disrupted 5-hmC patterning in leukemia patient-derived CD34+ stem/early progenitor cells with TET methylcytosine dioxygenase 2 (TET2) mutations. Thus, chemical conjugation and affinity purification of 5-hmC-enriched sequences followed by sequencing serve as resources for deciphering functional implications for gene expression during stem cell commitment and differentiation along a particular lineage. [Display omitted] •5-hmC levels change dramatically in a human erythroid differentiation system•Regions gaining 5-hmC are enriched for transcription factor binding sites•Regions gaining 5-hmC correlate with activating histone marks•TET2 deficiency disrupts 5-hmC patterns and compromises erythroid differentiation 5-hydroxymethylcytosine (5-hmC) is a DNA modification catalyzed by the TET enzymes. The significance of this modification is not completely defined, but it is known to be dysregulated in hematological malignancies. In this study, Wickrema, Godley, and colleagues demonstrate that 5-hmC plays an important role in hematopoietic stem cell commitment to the erythroid lineage and is associated with activating histone marks at a genome-wide level. TET2 deficiency disrupts 5-hmC patterns and compromises erythroid differentiation.