RESUMO
E2F-2 is a retinoblastoma (Rb)-regulated transcription factor induced during terminal erythroid maturation. Cyclin E-mediated Rb hyperphosphorylation induces E2F transcriptional activator functions. We previously reported that deregulated cyclin E activity causes defective terminal maturation of nucleated erythroblasts in vivo Here, we found that these defects are normalized by E2F-2 deletion; however, anemia in mice with deregulated cyclin E is not improved by E2F-2-loss, which itself causes reduced peripheral red blood cell (RBC) counts without altering relative abundances of erythroblast subpopulations. To determine how E2F-2 regulates RBC production, we comprehensively studied erythropoiesis using knockout mice and hematopoietic progenitors. We found that efficient stress erythropoiesis in vivo requires E2F-2, and we also identified an unappreciated role for E2F-2 in erythroblast enucleation. In particular, E2F-2 deletion impairs nuclear condensation, a morphological feature of maturing erythroblasts. Transcriptome profiling of E2F-2-null, mature erythroblasts demonstrated widespread changes in gene expression. Notably, we identified citron Rho-interacting kinase (CRIK), which has known functions in mitosis and cytokinesis, as induced in erythroblasts in an E2F-2-dependent manner, and we found that CRIK activity promotes efficient erythroblast enucleation and nuclear condensation. Together, our data reveal novel, lineage-specific functions for E2F-2 and suggest that some mitotic kinases have specialized roles supporting enucleation of maturing erythroblasts.
Assuntos
Núcleo Celular/metabolismo , Fator de Transcrição E2F2/genética , Fator de Transcrição E2F2/metabolismo , Eritroblastos/citologia , Eritropoese , Animais , Ciclo Celular , Diferenciação Celular , Ciclina E/metabolismo , Eritroblastos/metabolismo , Deleção de Genes , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteína do Retinoblastoma/metabolismoRESUMO
The Fbw7 ubiquitin ligase critically regulates hematopoietic stem cell (HSC) function, though the precise contribution of individual substrate ubiquitination pathways to HSC homeostasis is unknown. In the work reported here, we used a mouse model in which we introduced two knock-in mutations (T74A and T393A [changes of T to A at positions 74 and 393]) to disrupt Fbw7-dependent regulation of cyclin E, its prototypic substrate, and to examine the consequences of cyclin E dysregulation for HSC function. Serial transplantation revealed that cyclin E(T74A T393A) HSCs self-renewed normally; however, we identified defects in their multilineage reconstituting capacity. By inducing hematologic stress, we exposed an impaired self-renewal phenotype in cyclin E knock-in HSCs that was associated with defective cell cycle exit and the emergence of chromosome instability (CIN). Importantly, p53 deletion induced both defects in self-renewal and multilineage reconstitution in cyclin E knock-in HSCs with serial transplantation and CIN in hematopoietic stem and progenitor cells. Moreover, CIN was a feature of fatal T-cell malignancies that ultimately developed in recipients of cyclin E(T74A T393A); p53-null HSCs. Together, our findings demonstrate the importance of Fbw7-dependent cyclin E control to the hematopoietic system and highlight CIN as a characteristic feature of HSC dysfunction and malignancy induced by deregulated cyclin E.