Role of Survivin in Development of Megakaryocytes and Erythroid Cells

生存素在巨核细胞和红系细胞发育中的作用

基本信息

批准号：
8060642
负责人：
John D Crispino
金额：
$ 29.49万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-04-01 至 2012-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8060642
关键词：
Affect Alleles Apoptosis Apoptosis Inhibitor BFU-E Binding Biological Assay Biotinylation Blood Cells Bone Marrow Bone Marrow Cells Caspase Cell Cycle Cell Cycle Regulation Cell Death Cell Differentiation process Cell Survival Cells Cleaved cell Complementary DNA Complex Cytokinesis Data Defect Development Disease Down-Regulation Drops Erythrocytes Erythroid Erythroid Cells Erythropoiesis Excision Family Flow Cytometry GATA1 gene Gene Expression Gene Targeting Genes Goals Growth Hematopoiesis Hematopoietic Human In Vitro Kinetochores Knock-out Light Liquid substance Malignant Neoplasms Mass Spectrum Analysis Megakaryocytes Megakaryocytopoieses Messenger RNA Mitosis Mitotic Mitotic Spindle Apparatus Monitor Mouse Strains Mus Pancytopenia Pathway interactions Phenotype Physiological Play Polyploid Cells Process Proteins RNA Reporting Research Personnel Retroviridae Role Site Staging Tissues Transgenic Mice Transplantation base borealin caspase-3 cohort erythroid differentiation in vivo inner centromere protein knock-down leukemia member mouse development overexpression progenitor promoter protein complex recombinase segregation survivin transcription factor

项目摘要

DESCRIPTION (provided by applicant): Although erythroid cells and megakaryocytes derive from a common progenitor and share many essential transcription factors, their terminal maturation follows very different paths: erythroid cells undergo cell cycle exit and enucleation while megakaryocytes continue to progress through the cell cycle, but skip late stages of mitosis to become polyploid cells. In our efforts to identify genes that participate in this process, we discovered that survivin, a member of the inhibitor of apoptosis (IAP) family that also plays an essential role in cytokinesis, is differentially expressed during erythroid versus megakaryocyte development. Erythroid cells express survivin throughout their maturation, up through the terminal orthochromatic stage of differentiation. In contrast, purified murine megakaryocytes express nearly 4-fold lower levels of survivin mRNA compared to erythroid cells and no detectable protein. To investigate whether survivin is involved in the differentiation and/or survival of hematopoietic progenitors, we infected primary mouse bone marrow cells with retroviruses harboring the human survivin cDNA or control retrovirus, and then induced erythroid and megakaryocyte differentiation in both liquid culture and colony-forming assays. These studies revealed that overexpression of survivin antagonized megakaryocyte development, but did not affect the terminal differentiation of red blood cells. In contrast, a 50% reduction in survivin mRNA, caused by a heterozygous loss of survivin, blocked erythroid, but not megakaryocyte, development in vitro. Thus, our preliminary data support a physiologic role for persistent survivin expression in erythroid cells and a significantly reduced level of expression in megakaryocytes. Based on these findings, we hypothesize that persistent survivin expression is required for differentiation and/or survival of erythroid cells, while its reduction is essential for terminal maturation of megakaryocytes. In this application, we propose to investigate how survivin contributes to erythroid cell and megakaryocyte development. Specifically, we plan to 1) Determine the requirement for survivin in red cell and megakaryocyte development by conditional gene targeting in mice, 2) Characterize the mechanism by which survivin participates in erythroid cell differentiation by analyzing the phenotype of cultured erythroid cells with reduced survivin expression and by identifying survivin protein complexes in erythroid cells, and 3) Investigate why persistent survivin expression is inhibitory to megakaryocyte polyploidization and maturation by analysis of transgenic mice that ectopically express survivin in megakaryocytes and erythroid cells. A more detailed assessment of how survivin contributes to hematopoiesis will aid in our understanding of the role of cell cycle regulation and apoptosis in normal blood cell development, and may shed light into the mechanism by which erythroid cells and megakaryocytes arise from a common precursor.

描述（由申请人提供）：尽管红细胞细胞和巨核细胞源自一个共同的祖细胞并具有许多基本的转录因子，但它们的终末成熟遵循截然不同的路径：红细胞流失细胞周期的出口和吸收性，而巨核细胞却继续通过细胞周期，但会继续通过细胞循环，但跳过后期的细胞，成为多个细胞，成为多个细胞的裂痕。在我们识别参与这一过程的基因的努力中，我们发现在红细胞动力学与巨核细胞的发育过程中，差异表达了凋亡抑制剂（IAP）家族抑制剂（IAP）家族的成员。红细胞细胞在整个成熟过程中表达遗物，通过分化的末端正色阶段。相比之下，与红细胞细胞相比，纯化的鼠巨核细胞表达了近4倍的遗物mRNA水平，没有可检测的蛋白质。为了研究Survivin是否参与了造血祖细胞的分化和/或存活，我们用含有人类存活cDNA或对照逆转录病毒的逆转录病毒感染了原代小鼠骨髓细胞，然后在液态培养和液体培养和分类型培养分析中诱导红细胞和巨细胞分化。这些研究表明，Survivin的过表达拮抗了巨核细胞的发育，但并不影响红细胞的末期分化。相比之下，由于杂合丧失了Survivin，降低了50％的Survivin mRNA，但在体外发育却不会阻塞红细胞，但没有阻塞。因此，我们的初步数据支持红细胞细胞中持续的Survivin表达的生理作用，并且在巨核细胞中的表达水平显着降低。基于这些发现，我们假设持续的生存表达对于红细胞细胞的分化和/或存活需要，而其还原对于巨核细胞的终末成熟至关重要。在此应用中，我们建议研究Survivin如何促进红细胞和巨核细胞发育。 Specifically, we plan to 1) Determine the requirement for survivin in red cell and megakaryocyte development by conditional gene targeting in mice, 2) Characterize the mechanism by which survivin participates in erythroid cell differentiation by analyzing the phenotype of cultured erythroid cells with reduced survivin expression and by identifying survivin protein complexes in erythroid cells, and 3) Investigate why persistent survivin expression is通过分析转基因小鼠在巨核细胞和红细胞细胞中表达遗传的转基因小鼠，对大核细胞多倍体化和成熟。对遗产如何有助于造血的更详细评估将有助于我们理解细胞周期调节和凋亡在正常血细胞发育中的作用，并可能将丝状细胞和巨核细胞从常见的前体产生的机制浮出水面。

项目成果

期刊论文数量（3）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Survivin is not required for the endomitotic cell cycle of megakaryocytes.

巨核细胞的有丝分裂细胞周期不需要生存素。

DOI：
10.1182/blood-2008-11-190801
发表时间：
2009
期刊：
Blood
影响因子：
20.3
作者：
Wen,Qiang;Leung,Cindy;Huang,Zan;Small,Sara;Reddi,AlagarsamyLakku;Licht,JonathanD;Crispino,JohnD
通讯作者：
Crispino,JohnD

Erythroblast enucleation.