MRI: Acquisition of a Total Internal Reflection Fluorescence (TIRF) Microscope for Research and Education at Marquette University and the Greater Milwaukee Region

该奖项是为了支持自动化全内反射荧光（TIRF）显微镜系统的采购，以支持马奎特大学（MU）、威斯康星医学院（MCW）和密尔沃基地区其他研究小组的研究、教育和推广工作。TIRF显微镜是研究细胞表面大分子蛋白质复合物组装和运动的高分辨率机制和动力学的重要工具。TIRF也广泛适用于使用各种基于荧光的方法在单分子分辨率下研究酶。在大密尔沃基地区的MU和邻近机构需要TIRF显微镜的可用性，为本科生，研究生和博士后研究人员提供实践培训。PI和联合PI将为所有感兴趣的研究人员提供培训，包括从研究生到年轻本科生的专业知识。PI和合作PI在培养本科生和研究生方面有着良好的记录，而TIRF显微镜将成为马奎特大学和大密尔沃基地区周边机构教育工作的关键部分。PI， co -PI和其他用户的实验室正在进行的项目分为两大类TIRF显微镜应用。第一组将使用单分子FRET可视化荧光标记酶的运动，这些信息将被解卷积以了解它们的作用机制。第二组将利用荧光射频显微镜研究体内细胞过程，在细胞内跟踪荧光标记蛋白的动态。TIRF显微镜将是非常宝贵的：(1)进行atp依赖的运动蛋白的机制研究；(2)阐明载体结构域运动在生物素依赖性代谢酶中的作用；(3)明确朊病毒在肌动蛋白网络上形成的组装机制；(4)探讨温度在配子发育中的作用；(5)了解真核鞭毛的分子和细胞机制；(6)研究Ras GTPase在细胞膜上的相互作用(7)描述G蛋白和&amp；#946-阻滞素趋化因子受体信号传导的机制；(8)研究磷酸化在946-阻滞蛋白和E3泛素连接酶募集中的作用。这些研究的结果将发表在同行评议的科学期刊上，在科学会议上提出，并用于培训和外联活动。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

An award is made to support the acquisition of an automated Total Internal Reflection Fluorescence (TIRF) Microscope system to support the research, educational and outreach efforts at Marquette University (MU), Medical College of Wisconsin (MCW) and other research groups in the greater Milwaukee region. TIRF microscopy is an essential tool used to study, with high resolution, the mechanism and dynamics of macromolecular protein complex assembly and movement at cellular surfaces. TIRF has also been extensively adapted to study enzymes at single molecule resolution using various fluorescence-based approaches. The availability of a TIRF microscope at MU and neighboring institutions in the greater Milwaukee areas is required to provide hands-on training to undergraduate, graduate and postdoctoral researchers. The PI and co-PIs will provide training to all interested researchers, including students ranging in expertise from graduate to young undergraduate students. The PI and co-PIs have a strong record of training undergraduate and graduate students and the TIRF microscope will become a key part of the educational efforts at Marquette University and surrounding institutions in the greater Milwaukee region.The projects underway in the laboratories of the PI, Co-PIs and other users fall under two broad categories of TIRF microscopy applications. The first group will use single molecule FRET to visualize the motions of fluorescently labelled enzymes and this information will be deconvoluted to understand their mechanism of action. The second group will utilize the TIRF microscope to study in vivo cellular processes, where the dynamics of fluorescently labeled proteins are followed within the cell. The TIRF microscope will be invaluable to: (1) performing mechanistic studies of ATP-dependent motor proteins; (2) clarifying the role of carrier domain movements in biotin-dependent metabolic enzymes; (3) identifying the assembly mechanism of prion formation on actin networks; (4) exploring the role of temperature in gamete development; (5) understand the molecular and cellular mechanisms of eukaryotic flagella; (6) studying Ras GTPase interactions on cellular membranes (7) describing the mechanism of G protein and &#946-arrestin chemokine receptor signaling; and (8) investigating the role of phosphorylation in &#946-arrestin and E3 ubiquitin ligase recruitment. Results from these studies will be published in peer-reviewed scientific journals, presented at scientific meetings, and used in both training and outreach activities.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.