The role of Orc6 in DNA replication and cytokinesis.

这是一份对当前有效的R 01 GM 121449补助金的行政补充申请， Orc 6在DNA复制和胞质分裂中的作用父母补助金的主要目的是了解 使用果蝇和人细胞在真核细胞和组织中复制起始子Orc 6和ORC的功能 as model模型systems系统. ORC，异聚六亚基蛋白复合物，是DNA复制所必需的。最小的 Orc 6蛋白是所有真核生物DNA复制的重要组成部分。我们已经证明，在 果蝇Orc 6通过与septin复合物相互作用在胞质分裂中具有额外的作用， 保守的聚合蛋白装配，在许多物种中对胞质分裂至关重要， 细胞骨架的重要组成部分。在初步研究中，我们已经确定了Orc 6的结构特征， 直接影响蛋白质在复制和胞质分裂中的功能。这个项目的总体目标是 我们的建议是使用以下方法来确定Orc 6在DNA复制和胞质分裂中功能的不同机制： 果蝇作为一个模型系统。所有主要目标的母赠款需要不断净化野生环境 型和突变蛋白复合物（如ORC和septin复合物及其亚基）来研究蛋白质 功能和蛋白质/DNA相互作用。ChemiDoc成像仪提供一流的性能和易用性， 可见光（RGB）和远红/近红外（FR/NIR）荧光和化学发光检测， 通用凝胶文档应用。免扫描成像能够立即可视化蛋白质， 凝胶染色和蛋白质转移到印迹的即时验证。行政补充将允许我们 购买一个新的ChemiDoc MP，这将大大促进和推进我们的研究和工作流程。购买 项目所需的新设备和相关的研究费用将提供必要的 投资于研究型经济。我们相信，在新发现的帮助下， 由行政补充资助的ChemiDoc MP系统将大大增加我们的机会， 通过竞争性更新申请获得未来NIH的支持。这反过来又将确保保护 新设立的和现有的研究职位由父母补助金支持。

This is an application for the Administrative Supplement to the currently active R01 GM121449 grant entitled “The Role of Orc6 in DNA replication and cytokinesis”. The major thrust of the parent grant is to understand the functions of replication initiators Orc6 and ORC in eukaryotic cells and tissues using Drosophila and human cells as model systems. ORC, heteromeric six-subunit protein complex, is essential for DNA replication. The smallest of ORC subunits, Orc6 protein, is important for DNA replication in all eukaryotes. We have shown that in Drosophila Orc6 has an additional role in cytokinesis through interaction with the septin complex, a highly conserved polymerizing protein assembly that is critical for cytokinesis in many species and is recognized as important component of the cytoskeleton. In preliminary studies we have identified structural features of Orc6 that contribute directly to the functions of the protein in replication and cytokinesis. The overall goal of this proposal is to define the distinct mechanisms of Orc6 function in DNA replication and in cytokinesis using Drosophila as a model system. All major objectives of the parent grant require constant purifications of the wild type and mutant protein complexes (such as ORC and septin complex and their subunits) to study protein functions and protein/DNA interactions. ChemiDoc Imagers offer best-in-class performance with ease of use for visible light (RGB) and far red/near infrared (FR/NIR) fluorescence and chemiluminescence detection and all general gel documentation applications. Stain-free imaging enables immediate visualization of proteins without gel staining and instant verification of protein transfer to blots. The Administrative Supplement will allow us to purchase a new ChemiDoc MP that will greatly facilitate and advance our studies and workflow. The purchase of new equipment required for the project and associated research expenses will provide the necessary investment into the research-oriented economy. We believe that the discoveries made with the help of new ChemiDoc MP system funded by the Administrative Supplement will significantly strengthen our chances for obtaining future NIH support through competitive renewal application. This in turn will ensure preservation of the newly created and current research positions supported by the parent grant.