Establishing a hematopoietic genetic network through locus-specific integration of chromatin regulators

Abstract

The establishment and maintenance of cell type-specific transcriptional programs require an ensemble of broadly expressed chromatin remodeling and modifying enzymes. Many questions remain unanswered regarding the contributions of these enzymes to specialized genetic networks that control critical processes, such as lineage commitment and cellular differentiation. We have been addressing this problem in the context of erythrocyte development driven by the transcription factor GATA-1 and its coregulator Friend of GATA-1 (FOG-1). As certain GATA-1 target genes have little to no FOG-1 requirement for expression, presumably additional coregulators can mediate GATA-1 function. Using a genetic complementation assay and RNA interference in GATA-1-null cells, we demonstrate a vital link between GATA-1 and the histone H4 lysine 20 methyltransferase PR-Set7/SetD8 (SetD8). GATA-1 selectively induced H4 monomethylated lysine 20 at repressed, but not activated, loci, and endogenous SetD8 mediated GATA-1-dependent repression of a cohort of its target genes. GATA-1 used different combinations of SetD8, FOG-1, and the FOG-1-interacting nucleosome remodeling and deacetylase complex component Mi2β to repress distinct target genes. Implicating SetD8 as a context-dependent GATA-1 corepressor expands the repertoire of coregulators mediating establishment/maintenance of the erythroid cell genetic network, and provides a biological framework for dissecting the cell type-specific functions of this important coregulator. We propose a coregulator matrix model in which distinct combinations of chromatin regulators are required at different GATA-1 target genes, and the unique attributes of the target loci mandate these combinations.

Histone Methyltransferase Setd8 Represses Gata2 Expression and Regulates Erythroid Maturation

“…Cell counts and trypan blue staining demonstrated that knockdown of Setd8 did not affect ESRE proliferation or viability, consistent with a previous study in erythroid cells (20) (Fig. 2A and B).…”

“…The only previously published study examining the role of Setd8 in erythroid cells suggested that Setd8 plays an important role in Gata1-mediated transcriptional repression (20). Consistent with that study, the majority of differentially expressed genes associated with Gata2/Gata1 switch sites are upregulated upon Setd8 knockdown (93/141; 66%).…”

“…Setd8 is expressed in all three murine erythroid lineages (primitive, fetal definitive, and adult definitive) (33) and throughout the erythroid differentiation of human CD34 ϩ hematopoietic stem and progenitor cells (34); however, little is known about how Setd8 interacts with the complex network of transcription factors and chromatin modifiers that govern erythroid differentiation. To date, the only published study examining the function of Setd8 in erythroid cells was done using the G1E-GATA1ER cell line, and it suggested that Setd8 is important for Gata1-mediated gene repression (20). Taken together, these preliminary data imply that Setd8 and H4K20me1play an important role during erythropoiesis, but the function of this epigenetic pathway in erythroid cells is poorly understood.…”

“…Previous studies examining the role of Setd8 and H4K20me1 in transcriptional regulation have yielded conflicting results (7,15,(17)(18)(19)(20)(21)(22)(23)(24)(25), suggesting that the impact of Setd8 on gene expression may be dependent on genomic and/or cellular context. Initial studies, which focused on the E2F family of genes, demonstrated that H4K20me1 occupancy over promoter regions was associated with gene repression (24,25).…”

“…Monomethylation of histone H4 lysine 20 has been further implicated in a number of biologic processes central to erythropoiesis, including DNA replication (13), chromatin compaction (12), cell cycle progression (12,14), and DNA repair (15,16), but little is known about its function in erythroid cells. Setd8 and H4K20me1 are also thought to be transcriptional regulators (7,15,(17)(18)(19)(20)(21)(22)(23)(24)(25); however, in contrast to some of the better-characterized histone modifications (e.g., histone H3 lysine 4 methylation), the influence of this epigenetic pathway on gene expression has not been clearly defined. Disruption of the Setd8/H4K20me1 pathway has been associated with a number of human cancers, including leukemia (26)(27)(28)(29)(30), highlighting the relevance of this pathway to human health and disease.…”