Mechanisms Regulating Megakaryocyte Endomitosis

多倍体，即每个细胞二倍体DMA含量的增加，发生在各种细胞中，包括 巨核细胞(MKs)，在那里它是通过内有丝分裂细胞周期实现的。MK倍性程度 影响血小板水平和质量。在寻找MK内丝分裂调控因子的过程中，我们发现细胞周期蛋白D3 是这个谱系中占主导地位的D型细胞周期蛋白，它的上调，无论是异位还是通过 促血小板生成素治疗，增加体内MK倍体水平。最近，有报道称，体内有一种敲击 在已知的细胞周期蛋白D3靶点--细胞周期蛋白E的作用下，MK倍体水平显著降低。这是非常不同的 这些小鼠的其他血统和器官的正常发育。根据已知的能力， Cyclin E将细胞从静止的GO期拯救出来，根据我们的相关发现，我们提出了这一新的 认为MKs遵循少数内丝分裂细胞周期并过渡到静止期，而Cyclin E 独一无二地允许细胞周期重新进入。多倍化的MK也被编程为跳过后期和 胞质分裂。我们假设后期的纺锤体中间区是非典型的配置，这 染色体乘客蛋白的特征和相关变化(我们最近的发现)具有功能性 意义。为了加强对相关机制的探索，我们打算开发一种新的小鼠模型， MK含有标记的染色体和微管，可以对细胞进行实时成像。通过这个 方法和表达实验，我们将研究染色体和微管动力学以及 内丝分裂周期的连续性程度。提出了研究的四个具体目标：1.考察 细胞周期蛋白E在体内促进巨噬细胞倍性的能力；2.探讨细胞周期蛋白E的分子机制 MK多倍化的要求；3.建立标记染色体的MKs体内模型 研究该家系内有丝分裂的动态；4.研究 染色体乘客蛋白在MK减数分裂过程中的重新分布及其对MK倍性水平的影响。 总而言之，追求这些研究目标应该会增强我们对分子的理解 巨核细胞多倍化的机制，这一过程影响血小板的生物生成，从而影响血液 止血。

Polyploidy, the increase of diploid DMA content per cell, occurs in a variety of cells, including megakaryocytes (MKs), where it is achieved by an endomitotic cell cycle. The degree of MK ploidy influences platelet level and quality. In the search for regulators of MK endomitosis, we found that cyclin D3 is the predominant D-type cyclin in this lineage and that its upregulation, either ectopically or by thrombopoietin treatment, increases MK ploidy level in vivo. Recently, it was reported that an in vivo knock out of cyclin E, a known target of cyclin D3, significantly diminishes MK ploidy level. This is strikingly different from the normal development of other lineages and organs in these mice. Based on the known ability of cyclin E to rescue cells from a resting GOphase and on our related findings, we propose the novel contention that MKs follow few endomitotic cell cycles with transition into a resting phase, and that cyclin E uniquely allows cell cycle re-entry. Polyploidizing MKs are also programmed to skip late anaphase and cytokinesis. We hypothesize that the spindle midzone at late anaphase is atypically configured and that this feature and associated changes in chromosome passenger proteins (our recent finding) are of functional significance. To enhance exploration of related mechanisms, we intend to develop a novel mouse model with MKs containing marked chromosomes and microtubules that will allow live imaging of cells. Via this approach and expression experiments, we will study chromosome and microtubule dynamics as well as the degree of continuity of endomitotic cycles. Four Specific Aims of research are proposed: 1. To examine the ability of elevated cyclin E to promote MK ploidy in vivo; 2. To explore the molecular mechanism of cyclin E requirement for MK polyploidization; 3. To generate an in vivo model of MKs with labeled chromosomes and microtubules and to study the dynamics of endomitosis in this lineage; 4. To study the process of redistribution of chromosome passenger proteins during MK endomitosis and its effect on MK ploidy level. Taken together, pursuing these aims of research should enhance our understanding of the molecular mechanisms of megakaryocyte polyploidization, a process that impacts platelet biogenesis and hence, blood hemostasis.