Determining the function of sterol lipids in the bacterial domain

该项目的总体目标是确定细菌中类胆固醇类脂质分子的功能。胆固醇在高等生物的生物学中扮演着许多重要的角色，但类似胆固醇的类脂在细菌等更简单的生物体中的重要性还没有得到很好的了解。这里要检验的假设是，类固醇使细菌能够感觉到低水平的氧气并对其做出反应，而低水平的氧气会对它们的生长能力产生不利影响。这项工作有可能扩大我们对基本细胞途径的理解，并可以为未来具有广泛社会影响的研究提供信息-从人类生理学到对环境变化的反应。该项目还将通过为博士后学者以及来自不同背景的研究生和本科生提供科学培训，为扩大STEM领域的参与这一目标做出贡献。未被充分代表的少数族裔本科生将通过斯坦福大学的两个暑期项目参与这一项目，以开展与该提案的科学目标直接相关的研究项目。此外，一系列研讨会将把职业生涯中代表性不足的科学家带到斯坦福大学，展示他们的研究成果，并与学生和博士后学者互动，由主要研究人员开发和执行，两人本身都是代表性不足的少数民族女科学家。在这个项目中，将进行各种分子研究，以调查类胆固醇脂类物质，或称甾醇，在甲氧基球菌胶囊细菌中的功能作用。胶囊支原体是一种与环境相关的微生物，通过消耗甲烷和氧气生长，是40多年前第一个被证明能产生类固醇的细菌。然而，这些非常重要的脂类在细菌中的功能仍然是一个谜。该项目将测试这样一种假设，即这种微生物中某些甾醇结构的生物合成向细胞发出信号，表明它正在经历低氧应激(称为缺氧)，这种情况会对细菌的生理产生不利影响。为了验证这一假说，研究人员将：(1)确定不同的氧气水平如何影响胶囊微囊藻的甾醇合成；(2)识别和表征在不同氧气条件下将特定的类固醇结构运输到细菌外膜所需的蛋白质；以及(3)确定当胶囊微囊藻的固醇缺乏突变细胞在氧气充足和氧气受限的条件下生长时，哪些基因处于开启和关闭状态。总之，这些研究有可能识别独特的类固醇结合蛋白和转运体，以及细菌中新的类固醇调节机制。类固醇的这种调节作用已经在真核生物中得到了展示，但它将代表细菌领域的一种新功能。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

The overall goal of this project is to determine the functions cholesterol-like lipid molecules have in bacteria. Cholesterol plays many essential roles in the biology of higher organisms, but the importance of cholesterol-like lipids in simpler organisms like bacteria is not well understood. The hypothesis to be tested here is that sterols allow bacteria to sense and react to low levels of oxygen that adversely affect their ability to grow. The work has the potential to expand our understanding of a fundamental cellular pathway and can inform future studies that have broad societal impacts - from human physiology to response to environmental change. This project will also contribute to the goal of broadening participation in STEM fields by providing scientific training for postdoctoral scholars as well as graduate and undergraduate students from diverse backgrounds. Undergraduate underrepresented minority students will participate in this project through two summer programs at Stanford to undertake research projects that are directly relevant to the scientific goals of this proposal. Further, a seminar series that will bring early career underrepresented scientists to Stanford to present their research and interact with students and postdoctoral scholars will be developed and executed by the principle investigators, both of whom are underrepresented minority women scientists themselves.In this project, a variety of molecular studies will be undertaken to investigate the functional role of cholesterol-like lipids, or sterols, in the bacterium Methylococcus capsulatus. M. capsulatus, an environmentally relevant microbe that grows by consuming methane and oxygen, was the first bacterium shown to produce sterols over 40 years ago. However, the function of these very important lipids in bacteria remains a mystery. This project will test the hypothesis that the biosynthesis of certain sterol structures within this microbe signals to the cell that it is experiencing low oxygen stress (termed hypoxia), a condition that adversely affects the bacterium's physiology. To test this hypothesis, the researchers will: (1) determine how varying oxygen levels affect sterol synthesis in M. capsulatus; (2) identify and characterize proteins necessary for transporting specific sterol structures to the outer bacterial membrane under different oxygen conditions; and 3) determine what genes are turned on and off when M. capsulatus sterol-deficient mutant cells are grown under oxygen-replete and oxygen-limited conditions. Together, these studies have the potential to identify unique sterol-binding proteins and transporters, as well as novel sterol regulatory mechanisms in bacteria. This type of a regulatory role for sterols has been shown in eukaryotes but would represent a novel function in the bacterial domain.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.