Feedback in organelle size regulation

 描述(由申请人提供)：MBRS评分SC2方案将填补我们在液泡/溶酶体大小和形态如何影响细胞器执行其降解功能的能力方面的一个相当大的空白。这种细胞器结构和功能的缺陷与细胞毒蛋白清除减少有关，这是阿尔茨海默病的原因之一。溶酶体功能也影响肿瘤的发生，在高转移细胞模型中，溶酶体的丰度和直径变得异常调节。我的目标是阐明控制液泡大小和形态的调节机制，以及这些特征如何影响液泡功能。这项建议应用了新的方法--包括三维图像分析和计算建模--来确定细胞器结构如何在逐个细胞的基础上影响功能。在目标1中，Pi和他的学生将(A)确定液泡的生物发生和遗传在细胞周期中是如何协调的，以及(B)是否存在液泡大小的反馈调节，使用定量建模和实验验证。我们将从延时显微镜数据中测量液泡生长和遗传的速度。这些数据还将为开发一个定量模型提供信息，以测试细胞和液泡生长之间的反馈调节是否对调节液泡大小是必要的。这一模型将在实验中得到验证，以测试液泡是否能从遗传或环境扰动引起的大小扰动中恢复。在目标2中，我们将确定空泡的大小和形态如何影响其在受体下调中的功能。在与我的教师导师，加州大学伯克利分校的大卫·德鲁宾教授的密切磋商下，我们将开发一种荧光分析方法，用于测定细胞表面受体的内吞和降解率。我们将利用这一方法来探讨空泡形态突变对受体下调动力学的影响，以及空泡体积和表面积对下调动力学的独立影响。这一建议的发现将为神经退行性疾病和癌症的进展提供一个知识基础，并对这些疾病的诊断和治疗产生影响。为了执行这一研究计划，国际和平研究所将包括研究生和本科生研究人员，重点是招收和培训来自代表性不足背景的学生。

 DESCRIPTION (provided by applicant): This MBRS SCORE SC2 proposal will close a considerable gap in our knowledge on how vacuole/lysosome size and morphology impact the ability of the organelle to perform its degradative functions. Defects in this organelle's structur and function are implicated in reduced clearance of cytotoxic proteins, which contributes to Alzheimer's disease. Lysosome function also affects tumor genesis, and lysosome abundance and diameter become abnormally regulated in highly-metastatic cell models. My goals are to elucidate the regulatory mechanisms that govern vacuole size and morphology, and how these characteristics impact vacuole function. This proposal applies novel methods - including three-dimensional image analysis, and computational modeling - to determine how organelle structure impacts function on a cell-by-cell basis. In Aim 1, the PI and his students will (A) determine how vacuole biogenesis and inheritance are coordinated during the cell cycle and (B) if there is feedback regulation of vacuole size using quantitative modeling and experimental verification. We will measure the rates of vacuole growth and inheritance from time-lapse microscopy data. These data will also inform the development of a quantitative model to test if feedback regulation between cell and vacuole growth is necessary to regulate vacuole size. This model will be verified in experiments testing whether vacuoles recover from size perturbations due to genetic or environmental perturbations. In Aim 2, we will determine how the size and morphology of a vacuole impact its function in receptor down regulation. In close consultation with my faculty mentor, Prof. David Drubin at the University of California, Berkeley, we will develop a fluorescence assay for the rates of endocytosis and degradation of a cell-surface receptor. We will apply this assay to probe the effect of vacuole morphology mutations on the kinetics of receptor down regulation, as well as determine the independent effects of vacuole volume and surface area on down regulation kinetics. The findings of this proposal will provide a foundation of knowledge of the progression of neurodegenerative disease and cancer with implications for diagnosis and treatment of these diseases. To carry out this research plan, the PI will involve graduate and undergraduate researchers, with an emphasis on recruiting and training students from under-represented backgrounds.