Defining the regulatory role of mitotic cyclins on Endoplasmic Reticulum reorgani

描述(申请人提供)：细胞的主要功能，如蛋白质合成和折叠、脂肪合成、细胞内钙存储和大分子的核运输，都依赖于内质网(ER)。内质网结构或功能的缺陷与一系列慢性疾病，如2型糖尿病和自闭症，以及神经退行性疾病，如遗传性痉挛截瘫(HSP)和阿尔茨海默病有关。在有丝分裂过程中，内质网形态发生显著变化，这是适当的膜分裂和核膜形成所必需的现象。相对较少的人知道有丝分裂内质网的变化是如何发生的，它们是如何被调控的，或者是哪些分子机制导致了这些变化。有丝分裂周期蛋白：CDK激酶复合体是细胞分裂过程中核和细胞骨架动力学的主要调节者，但它们参与有丝分裂内质网重组的作用尚不清楚。此外，由于Ras超家族成员的小GTP酶控制着有丝分裂过程中的变化，它们很可能驱动有丝分裂内质网的动态。事实上，最近发现Rab5GTPase在有丝分裂的内质网结构中具有独立于其内吞作用的作用。因此，探索这些蛋白对内质网形态的影响是一个亟待解决的关键问题。本项目的目标是明确有丝分裂周期蛋白：CDK对有丝分裂过程中ER重组的调控，并确定一条与Rab5 GTPase活性协调的分子途径。这些过程将在模式生物黑腹果蝇的胚胎中进行详细研究。早期果蝇胚胎是研究有丝分裂的理想材料，因为它的细胞周期短，每15分钟就有几轮有丝分裂。具体地说，细胞周期进程或Rab5活性将通过已建立的方法进行调节，并通过GFP标记的ER标记的时间推移共聚焦成像来评估它们对ER形态的影响。本研究的目的如下：(1)确定有丝分裂周期蛋白在果蝇早期胚胎有丝分裂内质网重组中的作用。(2)研究Rab5活性的调节如何影响有丝分裂的ER重组。 公共卫生相关性：由于内质网在细胞功能中的基本作用，这项建议与人类主要的慢性疾病，如2型糖尿病、自闭症和阿尔茨海默病具有广泛而重要的相关性。拟议中的实验的完成将产生关于内质网形态在细胞周期中是如何调节的基本分子理解。这种特殊的知识将有助于开发遗传性痉挛截瘫等疾病的治疗方法。

DESCRIPTION (provided by applicant): Major cellular functions such as protein synthesis and folding, lipid synthesis, intracellular calcium storage, and nuclear transport of macromolecules all depend on the endoplasmic reticulum (ER). Failures in ER structure or function have been linked to a host of chronic diseases such as type-2 diabetes and autism, and neurodegenerative diseases such as hereditary spastic paraplegia (HSP) and Alzheimer's disease. During mitosis, ER morphology changes dramatically, a phenomenon necessary for proper membrane partitioning and nuclear membrane formation. Relatively little is known about how mitotic ER changes occur and how they are regulated, or which molecular mechanisms are responsible for these changes. Mitotic cyclin:Cdk kinase complexes are master regulators of nuclear and cytoskeletal dynamics during cell division, yet their involvement in mitotic ER reorganization is unknown. Additionally, since members of the Ras superfamily of small GTPases control changes during mitosis, they are likely to drive mitotic ER dynamics. Indeed, the Rab5 GTPase was recently found to have a role in mitotic ER structure independent of its endocytic role. Therefore, exploring how these proteins affect ER morphology is a key issue to be addressed. The goal of this project is to define the regulation of ER reorganization during mitosis by mitotic cyclin:Cdk and to define a molecular pathway that coordinates mitotic ER reorganization with Rab5 GTPase activity. These processes will be studied in detail in the embryo of the model organism, Drosophila melanogaster. The early Drosophila embryo is ideal for studying mitosis due to its rapid, abbreviated cell cycle, with initial rounds of mitosis occurring every ~15 minutes. Specifically, cell cycle progression or Rab5 activity will be modulated through established methods, and their effects on ER morphology assessed via timelapse confocal imaging of GFP-tagged ER markers. The aims of the proposal are as follows: (1) Determine the role of mitotic cyclins on mitotic ER reorganization in the early Drosophila embryo. (2) Investigate how modulation of Rab5 activity affects mitotic ER reorganization. PUBLIC HEALTH RELEVANCE: This proposal has broad and significant relevance to major chronic human diseases like type-2 diabetes, autism, and Alzheimer's disease, due to the fundamental roles of the ER in cellular function. Completion of the proposed experiments would yield a basic molecular understanding of how ER morphology is regulated during the cell cycle. This particular knowledge would contribute to the development of therapies for diseases like hereditary spastic paraplegia.