Phospholipase D Regulation of Exosome Secretion

在化学系生命过程化学项目的支持下，丹佛大学的米歇尔·诺尔斯正在研究细胞如何控制彼此之间的交流。细胞交流的一种方式是通过分泌纳米级的“外泌体”。外泌体是一种脂质包裹的容器，携带分子穿过身体，在那里它们被其他细胞吸收。外泌体可以改变其他细胞的功能，不管是好是坏。控制细胞如何制造外泌体及其释放的因素尚不完全清楚。该奖项支持的研究将侧重于脂质修饰蛋白，磷脂酶D (PLD)，一种酶，以及磷脂酸（PA），这是一种由PLD催化产生的脂质。PA可以改变膜的形状，并作为一种信号分子。PLD和PA在外泌体形成及其释放中的作用将被研究。除了产生关于外泌体的新知识之外，该项目还将为研究生和本科生创造机会，学习如何使用最先进的技术来解决细胞通信的前沿问题。Drs。诺尔斯和丹佛大学（University of Denver）的黛娜·洛克（Dinah Loerke）将为研究生开发一个同伴指导项目。科学和更广泛的影响活动的结果将在会议上提出并发表，使其他人能够建立在获得的关于细胞调节和学生指导的基础知识之上。磷脂酶D和磷脂酶A （PLD和PA）可能参与外泌体的形成和/或释放过程本身。这两个步骤都需要高度弯曲的膜。在本研究项目中，将在空间和时间上绘制外泌体分泌过程中PLD的存在和PA的形成。定量荧光显微镜方法将与标准生化方法结合使用，以评估外泌体的数量和它们从细胞中释放的频率。外泌体生命周期的形成、运输和融合阶段将在活细胞中进行评估。将改变PLD的活性和存在，以确定何时何地需要PLD和磷脂酸。这个项目有可能产生深远的科学影响；具体来说，通过展示使用生物物理工具/化学生物学来帮助阐明细胞通讯的重要机制。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。

With the support of the Chemistry of Life Processes Program in the Division of Chemistry, Michelle Knowles of the University of Denver is studying how cells control their communication with one another. One way that cells communicate is through the secretion of nanoscopic packets called “exosomes.” Exosomes are lipid-coated containers that carry molecules and travel through the body, where they are taken up by other cells. Exosomes can alter the function of other cells, for better or worse. The factors that control how cells make exosomes and their release are not fully understood. The research supported by this award will focus on the lipid-modifying protein, phospholipase D (PLD), an enzyme, and on phosphatidic acid (PA), which is a lipid whose production is catalyzed by PLD. PA can alter the shapes of membranes and acts as a signaling molecule. The role of PLD and PA in the formation of exosomes and their release will be investigated. Beyond generating new knowledge about exosomes, the project will create opportunities for graduate and undergraduate students to learn how to use state of the art technology to address problems at the forefront of cell communication. Drs. Knowles and Dinah Loerke (University of Denver) will develop a peer mentoring program for graduate students. The results of scientific and broader impact activities will be presented at conferences and published to allow others to build upon the foundational knowledge obtained about cellular regulation and student mentorship.Phospholipase D and phospholipase A (PLD and PA) could be involved in the formation of exosomes and/or in the release process itself. Both steps require highly curved membranes. In this research project, the presence of PLD and the formation of PA will be mapped in space and time during exosome secretion. Quantitative fluorescence microscopy approaches will be used in conjunction with standard biochemical methods to assess the number of exosomes and the frequency with which they are released from cells. The formation, trafficking and fusion stages of the exosome lifecycle will be assessed in live cells. The activity and presence of PLD will be altered to determine when and where PLD and phosphatidic acid are required. This project has the potential for far-reaching scientific impact; specifically, by demonstrating the use of biophysical tools/chemical biology to help to elucidate important mechanisms of cellular communication.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.