A Systems Approach Identifies Essential FOXO3 Functions at Key Steps of Terminal Erythropoiesis

Abstract

Circulating red blood cells (RBCs) are essential for tissue oxygenation and homeostasis. Defective terminal erythropoiesis contributes to decreased generation of RBCs in many disorders. Specifically, ineffective nuclear expulsion (enucleation) during terminal maturation is an obstacle to therapeutic RBC production in vitro. To obtain mechanistic insights into terminal erythropoiesis we focused on FOXO3, a transcription factor implicated in erythroid disorders. Using an integrated computational and experimental systems biology approach, we show that FOXO3 is essential for the correct temporal gene expression during terminal erythropoiesis. We demonstrate that the FOXO3-dependent genetic network has critical physiological functions at key steps of terminal erythropoiesis including enucleation and mitochondrial clearance processes. FOXO3 loss deregulated transcription of genes implicated in cell polarity, nucleosome assembly and DNA packaging-related processes and compromised erythroid enucleation. Using high-resolution confocal microscopy and imaging flow cytometry we show that cell polarization is impaired leading to multilobulated Foxo3 -/- erythroblasts defective in nuclear expulsion. Ectopic FOXO3 expression rescued Foxo3 -/- erythroblast enucleation-related gene transcription, enucleation defects and terminal maturation. Remarkably, FOXO3 ectopic expression increased wild type erythroblast maturation and enucleation suggesting that enhancing FOXO3 activity may improve RBCs production. Altogether these studies uncover FOXO3 as a novel regulator of erythroblast enucleation and terminal maturation suggesting FOXO3 modulation might be therapeutic in disorders with defective erythroid maturation.

FOXO3 Transcription Factor Is Essential for Protecting Hematopoietic Stem and Progenitor Cells from Oxidative DNA Damage

“…6). As we had noted previously, the transcript expression in primary Foxo3 Ϫ/Ϫ hematopoietic cells may not always fully correlate with the protein expression (41,42). These results together implicate FOXO3 in the regulation of BER in hematopoietic stem and progenitor cells.…”

“…XRCC1 is an essential element of base excision repair (40). Loss of FOXO3 may lead to discrepancies between mRNA and protein expressions (41,42). Given the critical OGG1-XRCC1 interaction for BER, it is likely that reduced expression of XRRC1 mediates the defective OGG1 activity (43)(44)(45) Ϫ/Ϫ mice and kept in vitro displayed similar numbers of ␥H2AX nuclear foci 2 h after 10 Gy IR (Fig.…”

Neomorphic effects of the neonatal anemia (Nan-Eklf) mutation contribute to deficits throughout development

“…In the present case, the levels of hepcidin in the Nan/+ mouse are high enough to exert a physiological effect (Nemeth et al, 2004b). (Kingsley et al, 2013;Liang et al, 2015), its levels are nonetheless significantly increased in the adult Nan/+ bone marrow. (C) Sera from wild-type (+/+) or Nan/+ adult littermates ('Nan') were monitored for expression of bilirubin (n=10-12, P<0.0001) or iron (n=5-7, P=0.0025).…”

What can we learn from ineffective erythropoiesis in thalassemia?

“…AKT kinase, the downstream target of PI3K, phosphorylates several transcription factors, including FOXOs, and regulates their activity 19, 20 . FOXOs play an important role in regulating cell growth, survival and differentiation 21, 22 .…”