Rho GTPases in Terminal Erythroid Maturation

红细胞终末成熟中的 Rho GTP 酶

基本信息

批准号：
8417358
负责人：
Theodosia Anastasios Kalfa
金额：
$ 38.25万
依托单位：
CINCINNATI CHILDRENS HOSP MED CTR
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-26 至 2016-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8417358
关键词：
Actins Actomyosin Adhesions Anemia BFU-E Binding Biochemical Breeding CFU-E Cell Nucleus Cell Polarity Cells Chronic Colchicine Cytokinesis Cytoskeletal Proteins Cytoskeleton Data Daughter Defect Development Disease Dysmyelopoietic Syndromes Erythroblasts Erythrocyte Membrane Erythrocytes Erythroid Erythroid Cells Erythropoiesis Excision Extracellular Matrix F-Actin Family member Fetal Liver Figs - dietary GTP Binding Genetic Goals Guanosine Triphosphate Phosphohydrolases Hematopoietic In Situ In Vitro Inflammatory Integrins Investigation Island LIMK1 gene Mammals Mediating Membrane Microdomains Microtubule Polymerization Microtubules Modeling Molecular Mus Myosin Type II Neoplasms Pathway interactions Play Poikilocytosis Polyploidy Process Production Protocols documentation Rattus Regulation Reticulocytes Role Safety Signal Transduction Speed Staging Surface Testing Thalassemia Therapeutic Intervention Time Transfusion Transplantation Tubulin Vascular blood supply Work abstracting anillin base bone cellular imaging citron rho-interacting kinase cofilin constriction daughter cell erythroid differentiation improved in utero in vivo macrophage non-muscle myosin novel polymerization profilin progenitor receptor reconstitution rho GTP-Binding Proteins

项目摘要

DESCRIPTION (provided by applicant): Terminal differentiation within the erythroid lineage in mammals concludes with the dramatic process of enucleation that results in reticulocyte formation. The interactions of erythroblasts with macrophages within the erythroblastic islands are critical for efficient enucleation. Defects in the final stages of erythropoiesis are common causes of anemia in chronic inflammatory diseases and neoplasia and in many primary hematological processes, like myelodysplasia and thalassemia. Limited understanding of the mechanisms involved in terminal erythroid maturation impedes development of novel therapies for such diseases. Using a novel analysis protocol of multiparameter high-speed cell imaging in flow we demonstrated evidence that enucleation is a multi-step process resembling asymmetric cytokinesis. It requires establishment of cell polarity through microtubule function, followed by formation of a contractile actomyosin ring, and coalescence of lipid rafts between reticulocytes and pyrenocytes. Moreover, we showed that Rac GTPases organize actin in the actomyosin ring and aggregate lipid rafts in the furrow between nascent reticulocyte and pyrenocyte during enucleation. RhoA GTPase is known to play a significant role in cytokinesis and abscission of the daughter cells. Based on the resemblances between erythroblast enucleation and cytokinesis we hypothesize that Rac and RhoA dynamically control erythroblast enucleation by intrinsic molecular pathways analogous to the ones conducting cytokinesis. Based on the critical role of erythroblast-macrophage interaction for efficient erythropoiesis and the known function of Rac and RhoA in signaling initiated by integrins we hypothesize that Rac and RhoA transduce critical macrophage signals to the erythroblasts, regulating terminal erythroid differentiation and enucleation within the erythroblastic islands. To define the mechanistic contribution of RhoA signaling and the relationship of RhoA/Rac in regulating actin and microtubule cytoskeleton in the final stages of erythropoiesis, we bred mice with erythroid specific deletion (EpoRGFPcre/+ driven) of RhoA and Rac1/Rac2/Rac3. To test our hypotheses, we propose in Aim 1 to define the molecular processes by which RhoA/Rac-regulated cytoskeleton dynamics contribute to erythroblast enucleation and in Aim 2 to determine the role of Rho GTPases in enucleation and erythroblast maturation within erythroblastic islands. The goal of this proposal is to investigate the intracellular and intercellular molecular mechanisms by which Rac and RhoA regulate erythroblast differentiation and enucleation via dynamic regulation of actin and microtubule cytoskeletons. This study has the potential to reveal targets for in vivo therapeutic intervention for anemias due to terminal erythroid maturation defects as well as for improving the efficiency of red blood cell production in vitro. (End of Abstract)

描述（由申请人提供）：哺乳动物中红细胞谱系中的末端分化以戏剧性的摘除过程结论，导致网状细胞形成。红细胞与巨噬细胞在红细胞岛内的巨噬细胞的相互作用对于有效的摘除至关重要。红细胞生成的最后阶段的缺陷是慢性炎症性疾病和肿瘤中贫血的常见原因，以及许多主要的血液学过程，如骨髓增生和丘脑症。对末端红细胞成熟涉及的机制的有限理解阻碍了此类疾病的新疗法的发展。使用流量中的多参数高速细胞成像的新分析方案，我们证明了摘除剂是类似于不对称细胞因子的多步过程。它需要通过微管功能建立细胞极性，然后形成收缩肌动蛋白环，以及网状细胞和肾上腺细胞之间的脂质筏的合并。此外，我们表明RAC GTP酶在肌动蛋白环中组织肌动蛋白，并在摘除核管期间在新生网状细胞和肾上腺细胞之间的沟中聚集脂质筏。众所周知，RhoA GTPase在细胞因子和子细胞的脱落中起着重要作用。基于红细胞摘除和细胞因子之间的相似之处，我们假设RAC和RhoA通过类似于进行细胞因子的固有分子途径动态控制染料细胞含糖。基于红细胞 - 巨噬细胞相互作用在有效的促红细胞生成方面的关键作用以及RAC和RhoA在整合素引发的信号传导中的已知功能，我们假设RAC和RhoA和RhoA将关键的巨噬细胞信号传达给红细胞，从。为了定义RhoA信号传导的机理贡献以及RhoA/RAC在红细胞生成的最后阶段调节肌动蛋白和微管细胞骨架中的关系，我们用Rhoa和Rac1/Rac2/Rac2/Rac3繁殖了红色小鼠，用红细胞特异性缺失（Eporgfppcre/+驱动）。为了检验我们的假设，我们在AIM 1中提出，以定义RhoA/RAC调节的细胞骨架动力学有助于红细胞摘除核的分子过程，并在AIM 2中确定Rho GTPases在依核细胞和红细胞成熟的rho gtpase在成核中的作用。该提案的目的是研究RAC和RhoA通过肌动蛋白和微管细胞骨架的动态调节调节红细胞分化和摘除的细胞内和细胞间分子机制。这项研究有可能揭示由于末端红细胞成熟缺陷以及提高体外红细胞产生的效率而导致的体内治疗干预靶标的贫血的靶标。（抽象的结尾）