Understanding SNX-BAR protein structure and function in yeast

了解酵母中的 SNX-BAR 蛋白质结构和功能

• 批准号: 2028519
• 负责人: Richard Chi
• 金额: $ 67.6万
• 依托单位: University of North Carolina at Charlotte
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2028519&HistoricalAwards=false
• 关键词: Understanding; SNX; BAR; protein; structure

Autophagy is the cellular process of removing aged or damaged components from the cell during routine cellular maintenance or during times of environmental stress such as starvation. The recycled by-products of autophagy can also be used to sustain critical metabolism for cell survival. Autophagy is conserved from yeast to humans and must be tightly controlled to prevent biological miscues that can result in aberrant cellular growth or death. The sorting nexins are a family of proteins that have been directly implicated in many aspects of autophagy, however the mechanism of their action there is poorly understood. This Project seeks to determine the regulatory elements that govern sorting nexin function during autophagy by determining the atomic structure and analyzing the function of a newly discovered sorting nexin. The Project will also provide graduate and undergraduate students an opportunity for training in biophysical and biochemical research. The Project will have an impact outside the laboratory by improving undergraduate curriculum through the development of novel 3D printing and Virtual Reality (VR) inquiry-based learning modules. In addition, the Project will create a course based undergraduate experience (CURE) centered on macromolecular structure and function. Overall, the broader impact will give scientists-in-training, including those from backgrounds underrepresented in STEM, an opportunity to enhance their scientific education.The sorting nexins (SNXs) constitute a diverse family of molecules that play varied roles in membrane trafficking, cell signaling, membrane remodeling and organelle motility. The SNX-BAR subfamily of the sorting proteins are typically thought to function in endosome sorting, but recently several have been shown to participate in autophagosome biogenesis. However, the mechanism of their role in autophagy is unclear. Prevailing models indicate that an evolutionarily conserved Phox-Homology (PX) domain, a feature common in all sorting nexins, binds specifically to phosphatidylinositol 3- phosphate (PI3P), the major lipid moiety of the endosome, to drive SNX-BAR function. The goal of this research is to understand the structure and function of a previously uncharacterized yeast SNX-BAR protein termed Vps501 and its interaction with the SEA complex that is involved in autophagy. Preliminary data suggests that Vps501 demonstrates a unique specificity for binding lipids and requires conformational changes to uniquely interact with the vacuole membrane. This Project will use a combination of biochemical, genetic and structural biology experiments to accomplish its goal. The Project will connect patterns of lipid specificity and structural characteristics in SNX-BAR proteins, while providing a new paradigm for understanding the relationship between the endo-vacuolar system and autophagy.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.

自噬是在常规细胞维持期间或在环境应激（例如饥饿）期间从细胞中移除老化或受损组分的细胞过程。自噬的回收副产物也可用于维持细胞存活的关键代谢。自噬从酵母到人类都是保守的，必须严格控制以防止可能导致异常细胞生长或死亡的生物学错误。 分类连接蛋白是一个蛋白质家族，直接参与自噬的许多方面，但对其作用机制知之甚少。 本项目旨在通过确定一种新发现的分类连接蛋白的原子结构和分析其功能，来确定在自噬过程中控制分类连接蛋白功能的调控元件。 该项目还将为研究生和本科生提供生物物理和生物化学研究方面的培训机会。 该项目将通过开发新颖的3D打印和虚拟现实（VR）探究式学习模块来改善本科课程，从而在实验室之外产生影响。 此外，该项目将创建一个以大分子结构和功能为中心的基于本科生经验的课程（CURE）。 总的来说，更广泛的影响将给科学家在培训，包括那些来自STEM代表性不足的背景，一个机会，以加强他们的科学教育。排序连接蛋白（SNX）构成了一个多样化的家庭的分子，发挥不同的作用，在膜运输，细胞信号，膜重塑和细胞器运动。通常认为SNX-BAR亚家族的分选蛋白在内体分选中起作用，但最近已经显示出几种参与自噬体生物发生。 然而，它们在自噬中的作用机制尚不清楚。 流行的模型表明，进化上保守的Phox同源（PX）结构域，在所有分选连接蛋白中常见的特征，特异性结合磷脂酰肌醇3-磷酸（PI 3 P），内体的主要脂质部分，以驱动SNX-BAR功能。 本研究的目的是了解以前未表征的酵母SNX-BAR蛋白Vps 501的结构和功能及其与参与自噬的SEA复合物的相互作用。 初步数据表明，Vps 501表现出独特的结合脂质的特异性，并需要构象变化，独特地与液泡膜相互作用。 该项目将使用生物化学，遗传和结构生物学实验的组合来实现其目标。 该项目将SNX-BAR蛋白质中的脂质特异性和结构特征的模式联系起来，同时为理解内空泡系统和自噬之间的关系提供新的范例。该奖项反映了NSF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。