Equipment Supplement for R35 GM144059 "Writers and Erasers of Ubiquitin Moieties in Control of Cell-to-Cell Transport in Plants"

项目总结 细胞间运输是多细胞生物体生命周期的核心，其 分子机制及其调控在不同的模型中得到了广泛的研究。 系统，包括植物和动物。然而，细胞间运输调控的一个关键方面 显然没有引起注意：泛素化/去泛素化的潜在参与(和 相应的泛素连接酶和去泛素酶)。在我们的父母项目中，我们有 获得了这种监管制度存在的证据，并正在表征其 分子组成和作用机理(S)。 这项工作在很大程度上依赖于使用共聚焦激光扫描显微镜来检测 并追随植物组织中带有荧光标记的蛋白质的命运。基于当前和 项目的规划轨迹，迫切需要改进我们的共焦 成像质量，即分辨率和信噪比，以提高采集数据的质量 以及他们的解释。为了解决该项目的这一关键技术问题，我们建议 用蔡司60A R2离线分析工作站获取蔡司Airyscan 2检测系统 它可以集成到我们现有的Zeiss LSM 900机器中，并提供重要的 其成像和图像分析能力的改进。

PROJECT SUMMARY Intercellular transport is central to the life cycle of multicellular organisms, and its molecular mechanisms and their regulation have been extensively studied in different model systems, including plants and animals. Yet, one key aspect of cell-to-cell transport regulation apparently escaped attention: the potential involvement of ubiquitylation/deubiquitylation (and the corresponding ubiquitin ligases and deubiquitinases). In our parental project, we have obtained evidence for the existence of such a regulatory system and are characterizing its molecular components and mechanism(s) of action. This work heavily depends on the use of a confocal laser scanning microscope to detect and follow the fate of fluorescently tagged proteins in plant tissues. Based on the current and projected trajectory of the project, there is an urgent need for improvement of our confocal imaging quality, i.e., resolution and signal-to-noise ratio, to enhance the quality of acquired data and their interpretation. To address this critical technical aspect of the project, we propose acquiring a Zeiss Airyscan 2 detection system with a Zeiss 60A R2 offline analysis workstation that can be incorporated into our existing Zeiss LSM 900 machine and provide significant improvement to its imaging and image analysis capabilities.