ABSTRACT
Folate, an essential vitamin, is a one-carbon acceptor and donor in key metabolic reactions. Erythroid cells harbor a unique sensitivity to folate deprivation, as revealed by the primary pathological manifestation of nutritional folate deprivation: megaloblastic anemia. To study this metabolic sensitivity, we applied mild folate depletion to human and mouse erythroid cell lines and primary murine erythroid progenitors. We show that folate depletion induces early blockade of purine synthesis and accumulation of the purine synthesis intermediate and signaling molecule, 5'-phosphoribosyl-5-aminoimidazole-4-carboxamide (AICAR), followed by enhanced heme metabolism, hemoglobin synthesis, and erythroid differentiation. This is phenocopied by inhibition of folate metabolism using the inhibitor SHIN1, and by AICAR supplementation. Mechanistically, the metabolically driven differentiation is independent of mechanistic target of rapamycin complex 1 (mTORC1) and adenosine 5'-monophosphate-activated protein kinase (AMPK) and is instead mediated by protein kinase C. Our findings suggest that folate deprivation-induced premature differentiation of erythroid progenitor cells is a molecular etiology to folate deficiency-induced anemia.
Subject(s)
Folic Acid , Purines , Mice , Humans , Animals , Folic Acid/metabolism , Cell Differentiation , Cell Line , Mechanistic Target of Rapamycin Complex 1ABSTRACT
Early erythroid progenitors are transit-amplifying cells with high proliferative capacity committed to undergoing red cell differentiation. CD71/CD24low progenitors are less mature and have greater proliferative capacity than CD71/CD24high. We present protocols for isolation of CD71/CD24low progenitors from mouse fetal liver using both fluorescence-activated cell sorting (FACS) and immunomagnetic enrichment. CD71/CD24low progenitors isolated with both approaches show similar transcriptomes at single-cell resolution and exhibit characteristic proliferative responses to glucocorticoids. For complete details on the use and execution of this protocol, please refer to Li et al. (2019).
