Defective erythroid differentiation in miR-451 mutant mice mediated by 14-3-3zeta.

Erythrocyte formation occurs throughout life in response to cytokine signaling. We show that microRNA-451 (miR-451) regulates erythropoiesis in vivo. Mice lacking miR-451 display a reduction in hematrocrit, an erythroid differentiation defect, and ineffective erythropoiesis in response to oxidative stress. 14-3-3zeta, an intracellular regulator of cytokine signaling that is repressed by miR-451, is up-regulated in miR-451(-/-) erythroblasts, and inhibition of 14-3-3zeta rescues their differentiation defect. These findings reveal an essential role of 14-3-3zeta as a mediator of the proerythroid differentiation actions of miR-451, and highlight the therapeutic potential of miR-451 inhibitors.

Amino acid residues 268-276 of the erythropoietin receptor contain an endocytosis motif and are required for erythropoietin-mediated proliferation.

Erythropoietin (EPO) promotes viability, proliferation and differentiation of mammalian erythroid progenitor cells via its specific cell surface receptor (EPO-R). We have previously shown that truncated EPO-Rs containing 267 amino acids or less were defective in internalization of (125)I-EPO, whereas internalization via a receptor derivative containing 276 amino acids was unaffected, thus directing focus to the nine amino acid residues FEGLFTTHK at positions 268-276 [Levin, Cohen, Supino, Yoshimura, Watowich, Neumann, FEBS Lett. 427 (1998) 164-170]. Here, a panel of EPO-R mutants was generated to determine the role of these residues in EPO endocytosis, down regulation of cell surface receptors and EPO-mediated signaling. While linking amino acid residues 268-276 to a truncated EPO-R (Delta+9 EPO-R) conferred both ligand uptake and ligand-independent down regulation of the respective receptor from the cell surface, Phe 272 was crucial for EPO endocytosis but not for ligand-independent down regulation. Additional receptor motifs probably play a role in EPO endocytosis and receptor down-regulation, as these processes were not adversely impaired in Delta268-276 EPO-R. A central role of residues 268-276, in particular Phe, was demonstrated by the inability of Delta268-276 and F268,272A EPO-Rs to support EPO-mediated signal transduction.