Schnöder T M, Arreba-Tutusaus P, Griehl I, Bullinger L, Buschbeck M, Lane S W, Döhner K, Plass C, Lipka D B, Heidel F H, Fischer T
Department of Hematology and Oncology, Center for Internal Medicine, Otto-von-Guericke University Medical Center, Magdeburg, Germany.
Internal Medicine III, Department of Hematology/Oncology, University Hospital Ulm, Ulm, Germany.
Cell Death Differ. 2015 Jun;22(6):974-85. doi: 10.1038/cdd.2014.186. Epub 2014 Nov 14.
Erythropoiesis is a tightly regulated process. Development of red blood cells occurs through differentiation of hematopoietic stem cells (HSCs) into more committed progenitors and finally into erythrocytes. Binding of erythropoietin (Epo) to its receptor (EpoR) is required for erythropoiesis as it promotes survival and late maturation of erythroid progenitors. In vivo and in vitro studies have highlighted the requirement of EpoR signaling through Janus kinase 2 (Jak2) tyrosine kinase and Stat5a/b as a central pathway. Here, we demonstrate that phospholipase C gamma 1 (Plcγ1) is activated downstream of EpoR-Jak2 independently of Stat5. Plcγ1-deficient pro-erythroblasts and erythroid progenitors exhibited strong impairment in differentiation and colony-forming potential. In vivo, suppression of Plcγ1 in immunophenotypically defined HSCs (Lin(-)Sca1(+)KIT(+)CD48(-)CD150(+)) severely reduced erythroid development. To identify Plcγ1 effector molecules involved in regulation of erythroid differentiation, we assessed changes occurring at the global transcriptional and DNA methylation level after inactivation of Plcγ1. The top common downstream effector was H2afy2, which encodes for the histone variant macroH2A2 (mH2A2). Inactivation of mH2A2 expression recapitulated the effects of Plcγ1 depletion on erythroid maturation. Taken together, our findings identify Plcγ1 and its downstream target mH2A2, as a 'non-canonical' Epo signaling pathway essential for erythroid differentiation.
红细胞生成是一个受到严格调控的过程。红细胞的发育通过造血干细胞（HSCs）分化为更定向的祖细胞，最终分化为红细胞来实现。促红细胞生成素（Epo）与其受体（EpoR）的结合是红细胞生成所必需的，因为它能促进红系祖细胞的存活和晚期成熟。体内和体外研究都强调了通过Janus激酶2（Jak2）酪氨酸激酶和Stat5a/b进行的EpoR信号传导作为一条核心途径的必要性。在此，我们证明磷脂酶Cγ1（Plcγ1）在EpoR - Jak2下游被激活，且不依赖于Stat5。Plcγ1缺陷的早幼红细胞和红系祖细胞在分化和集落形成潜力方面表现出严重受损。在体内，免疫表型定义的造血干细胞（Lin(-)Sca1(+)KIT(+)CD48(-)CD150(+)）中Plcγ1的抑制严重减少了红系发育。为了鉴定参与红系分化调控的Plcγ1效应分子，我们评估了Plcγ1失活后在全局转录和DNA甲基化水平上发生的变化。最常见的下游效应分子是H2afy2，它编码组蛋白变体macroH2A2（mH2A2）。mH2A2表达的失活重现了Plcγ1缺失对红系成熟的影响。综上所述，我们的研究结果确定Plcγ1及其下游靶点mH2A2是红系分化所必需的“非经典”Epo信号通路。
