Fbw7-dependent cyclin E regulation ensures terminal maturation of bone marrow erythroid cells by restraining oxidative metabolism

Abstract

The mechanisms that coordinate the final mitotic divisions of terminally differentiated bone marrow erythroid cells with components of their structural and functional maturation program remain largely undefined. We previously identified phenotypes resembling those found in early-stage myelodysplastic syndromes, including ineffective erythropoiesis, morphologic dysplasia, and hyper-cellular bone marrow, in a knock-in mouse model in which cyclin E mutations were introduced at its two Cdc4 phosphodegrons (CPDs) to ablate Fbw7-dependent ubiquitination and degradation. Here we have examined the physiologic consequences of cyclin E dysregulation in bone marrow erythroid cells during terminal maturation in vivo. We found cyclin E protein levels in bone marrow erythroid cells are dynamically regulated in a CPD-dependent manner and that disruption of Fbw7-dependent cyclin E regulation impairs terminal erythroid cell maturation at a discrete stage prior to enucleation. At this stage of erythroid cell maturation, CPD phosphorylation of cyclin E regulates both cell cycle arrest and survival. We also found normal regulation of cyclin E restrains mitochondrial reactive oxygen species accumulation and expression of genes that promote mitochondrial biogenesis and oxidative metabolism during terminal erythroid maturation. In the setting of dysregulated cyclin E expression, p53 is activated in bone marrow erythroid cells as part of a DNA damage response-type pathway, which mitigates ineffective erythropoiesis, in contrast to the role of p53 induction in other models of dyserythropoiesis. Finally, cyclin E dysregulation and ROS accumulation induce histone H3 lysine 9 hyper-methylation and disrupt components of the normal terminal erythroid maturation gene expression program. Thus, ubiquitin-proteasome pathway control of G1-to-S-phase progression is intrinsically linked to regulation of metabolism and gene expression in terminally differentiating bone marrow erythroid cells.

miR-214 protects erythroid cells against oxidative stress by targeting ATF4 and EZH2

“…2B and D, P o0.001). Analogous to previous studies [51,52], intracellular ROS generation in MEL cells was significantly increased by 50% upon erythroid differentiation induced by DMSO (Po 0.05), and concurrent 60% reduction of miR-214 expression was observed (Figs. 1C and 2E).…”

The heme exporter Flvcr1 regulates expansion and differentiation of committed erythroid progenitors by controlling intracellular heme accumulation

“…12 Interestingly, the switch from proliferation to differentiation in the erythroid lineage is obtained by restraining oxidative metabolism. 25 Concomitantly, the rate of heme synthesis increases to sustain hemoglobin production. At this time, the expression of FLVCR1a at the plasma membrane becomes dispensable, as indicated by our data from Flvcr1a -/-embryos.…”

E2F-2 Promotes Nuclear Condensation and Enucleation of Terminally Differentiated Erythroblasts

“…In this model, hyperstable cyclin E results in defective terminal erythroid maturation, with abnormal accumulation of basophilic erythroblasts, reduced numbers of orthochromatic erythroblasts and reticulocytes, and anemia (12,13). Because cyclin E knock-in erythroid cells exhibit hyperphosphorylated Rb (13) and E2F-2 is the major E2F transcriptional activator expressed in late-stage bone marrow erythroid cells, we hypothesized that E2F-2 deletion would revert the defective maturation phenotypes found in cyclin E knock-in bone marrows. To test this, we crossed cyclin E T74A T393A mice with E2F-2 knockout mice and studied erythroid maturation in resulting animals.…”

“…We have previously reported the generation of a cyclin E T74A T393A knock-in mouse model, in which the major ubiquitylation pathway (controlled by SCF Fbw7 ) for catalytically active cyclin E is disabled by mutations in its Cdc4 phosphodegron motifs (12). In this model, hyperstable cyclin E results in defective terminal erythroid maturation, with abnormal accumulation of basophilic erythroblasts, reduced numbers of orthochromatic erythroblasts and reticulocytes, and anemia (12,13). Because cyclin E knock-in erythroid cells exhibit hyperphosphorylated Rb (13) and E2F-2 is the major E2F transcriptional activator expressed in late-stage bone marrow erythroid cells, we hypothesized that E2F-2 deletion would revert the defective maturation phenotypes found in cyclin E knock-in bone marrows.…”

“…Upstream mitogen-stimulated pathway signaling converges upon the G 1 cyclin-Cdks, leading to Rb hyperphosphorylation and E2F transcription factor activation (9-11). Deregulation of cyclin E-Cdk2 activity, caused by disruption of the major pathway that controls ubiquitin-mediated destruction of cat-alytically active cyclin E, induces dyserythropoiesis associated with cell-autonomous effects on survival and gene expression (12)(13)(14).…”