Loss of FoxM1 Promotes Erythroid Differentiation through Increased Proliferation of Erythroid Progenitors
Authors: Youn Minyoung , Bibikova Elena , LaVasseur Corinne , Glader Bertil , Sakamoto Kathleen , Narla Anupama 
Published: October 2019

FoxM1 belongs to the fork head/winged-helix family of transcription factors and regulates a network of proliferation-associated genes including the G2/M transition, chromosome segregation, and spindle assembly. FoxM1 expression is commonly upregulated in a number of human cancers such as liver, ovarian, breast, prostate, colon, and brain tumors. Its abnormal upregulation has been shown to be a key driver of cancer progression and an initiating factor of oncogenesis. In normal cells, FoxM1 is highly expressed in multipotent progenitor cells and inhibits differentiation of the progenitors, suggesting that FoxM1 plays in a role in the maintenance of multipotent progenitor cells. However, the exact molecular mechanism by which FoxM1 regulates stem/progenitor cells is still uncharacterized. In this study, we have examined the role of FoxM1 in normal hematopoiesis using human cord blood CD34+ cells.

To investigate the role of FoxM1 in normal hematopoiesis, we infected human cord blood CD34+ cells with FoxM1 shRNA lentivirus and observed blood cell differentiation using FACS analysis with a range of cell surface markers. We found that knockdown of FoxM1 resulted in an increase of the erythroid population (CD71+/GlyA+), a decrease of the myeloid population (CD11b+), and an unchanged megakaryocyte population (CD41a+) in two phase liquid culture system. Overall, we found a 2-fold increase in the erythroid population compared to the myeloid population. Importantly, methylcellulose colony assays also demonstrated increased numbers of CFU-E colonies (2-2.5 fold increase compared to control) and decreased numbers of CFU-GM colonies in FoxM1 knockdown cells. Taken together, these findings imply a role for FoxM1 in normal erythropoiesis.

To better define the function of FoxM1 in hematopoietic cells, we sorted distinct populations of cells based on their cell surface marker expression and quantitated FoxM1 expression level by RT-qPCR. FoxM1 had a 3-fold increased expression in CD71+ (erythroid) cells compared to CD11b+ (myeloid) cells. Additionally, we found FoxM1 expression was particularly elevated in the BFU-E and CFU-E stages of erythropoiesis, suggesting a functional role for FoxM1 in erythroid progenitor proliferation.

Finally, to study the potential molecular mechanism of FoxM1 in normal hematopoiesis, we analyzed cell cycle progress in FoxM1 knockdown cells with DAPI staining. We found increased S and G2/M phases in FoxM1 knockdown cells, which was significant only in the CD71+ (erythroid) population and not in the unsorted cell populations. We also detected an increase of BrdU+ cells in FoxM1 knockdown CD71+ population by BrdU incorporation assay, indicating faster proliferation of erythroid progenitors with FoxM1 knockdown.

These findings suggest a novel function of FoxM1 in normal human hematopoiesis, in which FoxM1 deficiency leads to increased proliferation of erythroid progenitors resulting in increased erythroid differentiation. Our data indicate that FoxM1 inhibitors, such as Thiostrepton or FDI-6, may be beneficial in treating patients with anemia.

Disclosures

No relevant conflicts of interest to declare.