The histone methyltransferase Setd8 alters the chromatin landscape and regulates the expression of key transcription factors during erythroid differentiation

Abstract

Background: SETD8 is the sole methyltransferase capable of mono-methylating histone H4, lysine 20. SETD8 and H4K20me1 play a role in a number of essential biologic processes, including cell cycle progression, establishment of higher order chromatin structure, and transcriptional regulation. SETD8 is highly expressed in erythroid cells and erythroid deletion of Setd8 is embryonic lethal by embryonic day 11.5 (E11.5) due to profound anemia, suggesting that it has an erythroid-specific function. The function of SETD8 in the hemopoietic system is poorly understood. The goal of our study was to gain insights into the function of SETD8 during erythroid differentiation. Results:We performed ATAC-seq (assay for transposase-accessible chromatin) on sorted populations of E10.5 Setd8 mutant and control erythroblasts. Accessibility profiles were integrated with expression changes and a mark of heterochromatin (H3K27me3) performed in wild-type E10.5 erythroblasts to further understand the role of SETD8 in erythropoiesis. Data integration identified regions of greater chromatin accessibility in Setd8 mutant cells that colocated with H3K27me3 in wild-type E10.5 erythroblasts suggesting that these regions, and their associated genes, are repressed during normal erythropoiesis. The majority of these more accessible regions were located in promoters and they frequently co-located with the NFY complex. Pathway analysis of genes identified through data integration revealed stemness-related pathways. Among those genes were multiple transcriptional regulators active in multipotent progenitors, but repressed during erythroid differentiation including Hhex, Hlx, and Gata2. Consistent with a role for SETD8 in erythroid specification, SETD8 expression is up-regulated upon erythroid commitment, and Setd8 disruption impairs erythroid colony forming ability. Conclusion:Taken together, our results suggest that SETD8 is an important regulator of the chromatin landscape during erythroid differentiation, particularly at promoters. Our results also identify a novel role for Setd8 in the establishment of appropriate patterns of lineage-restricted gene expression during erythroid differentiation.

Epigenetic and Transcriptional Control of Erythropoiesis

“…Dynamic changes in histone methylation also occur during erythropoiesis, with both histone methyltransferases, such as NSD1 ( Tauchmann et al, 2020 ) and SETD8, ( Malik et al, 2015 ; Malik et al, 2017 ; Myers et al, 2020 ; DeVilbiss et al, 2015 ), and histone demethylases, such as LSD1, ( Kerenyi et al, 2013 ; Yu et al, 2021 ), playing critical role in erythroid gene regulation and erythroid maturation. The distribution of well-characterized histone markers of activation and repression, including H3K4me3, H3K27me3, and H3K9me3 appear to be established at the time of erythroid commitment, and are relatively stable during terminal maturation.…”

“…Although Setd8 is expressed broadly in proliferating tissues, it is highly expressed in erythroid cells compared to most other cell types. ( Wu et al, 2016 ; Myers et al, 2020 ). Loss of Setd8, results in defective chromatin condensation and disruption of the nuclear membrane.…”

“…They were able to express erythroid-specific genes, such as the globins, but were unable to repress the non-erythroid transcriptome, accompanied by an increase in chromatin accessibility at those loci. ( Myers et al, 2020 ).…”

“…In maturing erythroblasts, Setd8 plays an important role in establishing appropriate patterns of gene expression, ( DeVilbiss et al, 2013 ), and represses GATA2, and other key transcriptional regulators during erythroid differentiation. ( DeVilbiss et al, 2015 ; Malik et al, 2015 ; Myers et al, 2020 ).…”

Codanin-1 mutations engineered in human erythroid cells demonstrate role of CDAN1 in terminal erythroid maturation

“…Single guide RNA and associated CRISPR/Cas9 editing [22], cell culture [12,22,23], IFC [22,24], flow cytometry [25], and transcriptomic analysis [26] were completed as previous described. For additional details please see supplementary methods.…”