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Setd2 regulates hematopoietic stem cells
were identified in MDS patients,38 but whether SETD2 is involved in MDS development is still unclear. To examine whether SETD2 is down-regulated in MDS del(5q), we checked SETD2 expression levels in an MDS del(5q) cohort with 3 different probes. However, no significant differences between del(5q) and the control groups were found (Online Supplementary Figure S5A). Besides erythroid dysplasia and BM fibrosis,39 there are increased propor- tions of erythroblasts in the BM. Moreover, as the Setd2Δ/Δ mice became older, the percentage of erythroblasts accu- mulated and could even reach up to 80% in some mice, which is somewhat similar to the BM of acute erythroid leukemia (pure erythroid type).40
It is clear that Setd2 plays a critical role in maintaining the identity and functions of HSCs. Setd2Δ/Δ mice showed reduced HSCs and capability of BM reconstitution after transplantation. Setd2Δ/Δ HSCs showed loss of quiescence, increased apoptosis, and reduced multi-potent differentia- tion potential. Unbiased GSEA and GO analysis also indi- cated the upregulation of lineage development/differenti- ation pathways and related genes. Thus, there could be two explanations for the dramatic reduction in HSC num- bers. First, some HSCs exited from quiescence and com- mitted to differentiation. The balance between self- renewal and differentiation is important for the mainte- nance of the stem cell pool. Pushing HSCs to differentiate
would come at the expense of self-renewal and lead to the exhaustion of HSCs. The other reason could be that some HSCs directly underwent cell death. However, Setd2Δ/Δ mice did not progress to pancytopenia and BM failure without a stress challenge; this is in line with the finding that, at steady stage, a limited number of HSPCs would be sufficient to maintain normal hematopoiesis.41
Setd2, a histone methyltransferse, regulates H3K36me3. The direct effect after Setd2 knockout is the impact on his- tone modifications and other closely related H3K36 methyltransferases. Thus, the expression levels of Ash1l, Nsd1/2/3, and related histone markers were assessed first. The results showed significantly up-regulated Nsd1/2/3, accompanied with increased H3K36me1 and H3K36me2. Recently, we found that downregulation of SETD2 leads to a global elevation of DOT1L-mediated H3K79me2 in MLL-AF9 leukemia.42 Consistent with this finding, we observed a dramatic increase in H3K79me2 and H3K4me3, implying the promoting of transcriptional elongation. The enhanced RNA pol II elongation was fur- ther confirmed by the up-regulated pol II (Ser2P) and pol II (Ser5P) phosphorylations. We confirmed the significant upregulation of Gata1, Gata3, Klf1, and Myc in Setd2Δ/Δ SLAM-HSCs. These subsets of genes are sensitive to enhanced pol II elongation.
The enhanced elongation after Setd2 knockout was
haematologica | 2018; 103(7)
Figure 8. The diagram of our working model. In nor- mal adult hematopoietic stem cells (HSCs), Setd2, responsible for H3K36me3, could repress Nsds, which are responsible for H3K36me1/2. Nsds inter- act with Brd4, p-TEFb, and Dot1l to stimulate tran- scriptional elongation. On the other hand, Setd2 binds to pol II (Ser2P) and pol II (Ser5P) doubly mod- ified CTD repeats. Thus, a subset of genes, such as Myc, is maintained at a proper level to keep the bal- ance between quiescence and differentiation of adult stem cells (top). In Setd2Δ/Δ HSCs, Setd2 loss leads to the upregulation of NSDs, which would further enhance the Pol II phosphorylation and elongation, resulting in the upregulation of Myc. When treated with Brd4/Dot1l/p-TEFb inhibitors, the pol II (Ser2), pol II (Ser5), and the expressions of the Myc could be down-regulated (bottom).
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