RESUMO
An estimated 6% to 7% of the earth's population carries a mutation affecting red blood cell function. The ß-thalassemias and sickle cell disease are the most common monogenic disorders caused by these mutations. Increased levels of γ-globin ameliorate the severity of these diseases because fetal hemoglobin (HbF; α2γ2) can effectively replace adult hemoglobin (HbA; α2ß2) and counteract polymerization of sickle hemoglobin (HbS; α2ß(S)2). Therefore, understanding the molecular mechanism of globin switching is of biologic and clinical importance. Here, we show that the recently identified chromatin factor Friend of Prmt1 (FOP) is a critical modulator of γ-globin gene expression. Knockdown of FOP in adult erythroid progenitors strongly induces HbF. Importantly, γ-globin expression can be elevated in cells from ß-thalassemic patients by reducing FOP levels. These observations identify FOP as a novel therapeutic target in ß-hemoglobinopathies.
Assuntos
Hemoglobina Fetal/genética , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Células Cultivadas , Embrião de Mamíferos/metabolismo , Células Eritroides/citologia , Células Eritroides/metabolismo , Hemoglobina Fetal/metabolismo , Humanos , Camundongos , Proteínas Nucleares/genética , Fatores de Transcrição/genéticaRESUMO
BACKGROUND: In erythroblasts, the CoREST repressor complex is recruited to target promoters by the transcription factor Gfi1b, leading to repression of genes mainly involved in erythroid differentiation. Hmg20b is a subunit of CoREST, but its role in erythropoiesis has not yet been established. DESIGN AND METHODS: To study the role of Hmg20b in erythropoiesis, we performed knockdown experiments in a differentiation-competent mouse fetal liver cell line, and in primary mouse fetal liver cells. The effects on globin gene expression were determined. We used microarrays to investigate global gene expression changes induced by Hmg20b knockdown. Functional analysis was carried out on Hrasls3, an Hmg20b target gene. RESULTS: We show that Hmg20b depletion induces spontaneous differentiation. To identify the target genes of Hmg20b, microarray analysis was performed on Hmg20b knockdown cells and controls. In line with its association to the CoREST complex, we found that 85% (527 out of 620) of the deregulated genes are up-regulated when Hmg20b levels are reduced. Among the few down-regulated genes was Gfi1b, a known repressor of erythroid differentiation. Among the consistently up-regulated targets were embryonic ß-like globins and the phospholipase HRAS-like suppressor 3 (Hrasls3). We show that Hrasls3 expression is induced during erythroid differentiation and that knockdown of Hrasls3 inhibits terminal differentiation of proerythroblasts. CONCLUSIONS: We conclude that Hmg20b acts as an inhibitor of erythroid differentiation, through the down-regulation of genes involved in differentiation such as Hrasls3, and activation of repressors of differentiation such as Gfi1b. In addition, Hmg20b suppresses embryonic ß-like globins.
Assuntos
Células Eritroides/citologia , Células Eritroides/metabolismo , Eritropoese/genética , Proteínas de Grupo de Alta Mobilidade/metabolismo , Proteínas Repressoras/metabolismo , Animais , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Células Cultivadas , Proteínas Correpressoras , Proteínas de Ligação a DNA , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Técnicas de Silenciamento de Genes , Células HeLa , Proteínas de Grupo de Alta Mobilidade/genética , Humanos , Camundongos , Complexos Multiproteicos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Fosfolipases A2 Independentes de Cálcio/genética , Proteínas Proto-Oncogênicas/genética , Proteínas Repressoras/genéticaRESUMO
We describe the isolation and characterization of Friend of Prmt1 (Fop), a novel chromatin target of protein arginine methyltransferases. Human Fop is encoded by C1orf77, a gene of previously unknown function. We show that Fop is tightly associated with chromatin, and that it is modified by both asymmetric and symmetric arginine methylation in vivo. Furthermore, Fop plays an important role in the ligand-dependent activation of estrogen receptor target genes, including TFF1 (pS2). Fop depletion results in an almost complete block of estradiol-induced promoter occupancy by the estrogen receptor. Our data indicate that Fop recruitment to the promoter is an early critical event in the activation of estradiol-dependent transcription.