Lei Yu

Congenital dyserythropoietic anemia type II (CDAII) is an autosomal recessive disease resulting from loss-of-function mutations in SEC23 homolog B ( SEC23B ). We have previously shown that increased expression of SEC23A, a paralogous protein for SEC23B, rescues the CDAII erythroid defect. Here, we generated a human erythroid cell line that expresses enhanced green fluorescent protein (eGFP) from the endogenous SEC23A locus and performed a small-molecule screen to identify compounds that increased SEC23A-eGFP abundance. The top compound passing all filters was an inhibitor of lysine-specific demethylase 1 (LSD1). We found that LSD1 inhibition with RN1 resulted in increased SEC23A expression in erythroid cells derived from human hematopoietic stem and progenitor cells (HSPCs) at doses that did not impair erythroid cell growth or differentiation and rescued the erythroid defect resulting from SEC23B deletion. Genetic down-regulation of LSD1 led to a marked increase in SEC23A mRNA expression in HSPC-derived erythroid cells. Deletion of Lsd1 in mouse erythroid cells resulted in increased Sec23a expression, and RN1 treatment ameliorated the erythroid defect observed in a CDAII mouse model. Mechanistically, we found that LSD1 occupied a sequence in the SEC23A promoter, repressing SEC23A transcription. Deletion of the promotor sequence occupied by LSD1 resulted in increased SEC23A expression and amelioration of CDAII. These findings highlight that LSD1 represses SEC23A transcription and that LSD1 inhibition results in de-repression of SEC23A expression and amelioration of the CDAII erythroid defect, suggesting promising therapeutic strategies for CDAII.

Nuclear receptor TR4 (NR2C2) was previously shown to bind to the –117 position of the γ-globin gene promoters in vitro, which overlaps the more recently described BCL11 transcription factor A (BCL11A) binding site. The role of TR4 in human γ-globin gene repression has not been extensively characterized in vivo, whereas any relationship between TR4 and BCL11A regulation through the γ-globin promoters is unclear at present. We show here that TR4 and BCL11A competitively bind in vitro to distinct, overlapping sequences, including positions overlapping –117 of the γ-globin promoter. We found that TR4 represses γ-globin transcription and fetal hemoglobin accumulation in vivo in a BCL11A-independent manner. Finally, examination of the chromatin occupancy of TR4 within the β-globin locus, compared with BCL11A, shows that both bind avidly to the locus control region and other sites, but only BCL11A binds to the γ-globin promoters at statistically significant frequency. These data resolve an important discrepancy in the literature and, thus, clarify possible approaches to the treatment of sickle cell disease and β-thalassaemia.