Integrating the Regulatory Components of Sphingolipid Biosynthesis in Arabidopsis

整合拟南芥中鞘脂生物合成的调节成分

• 批准号: 1158500
• 负责人: Edgar Cahoon
• 金额: $ 68.68万
• 依托单位: University of Nebraska-Lincoln
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1158500&HistoricalAwards=false
• 关键词: Integrating; Regulatory; Components; Sphingolipid; Biosynthesis

Sphingolipids contribute to protein trafficking and the organization and physical properties of membranes as major structural components of the plasma membrane, tonoplast, and endomembrane system. Sphingolipids also act as dynamic regulators of basic cellular processes, including programmed cell death (PCD), which is triggered by the accumulation of the bioactive sphingolipid metabolites long-chain bases and ceramides. The project is designed to elucidate the regulatory components and their interactions that finely mediate the requirements for sufficient amounts of sphingolipids to support cell viability and growth yet prevent the buildup of sphingolipid metabolites to levels that induce PCD. The project will build upon emerging evidence for post-translational regulation of sphingolipid synthesis through serine palmitoyltransferase (SPT), a heterodimeric enzyme that catalyzes the first step in this pathway. Arabidopsis homologs of new players in the SPT regulatory network will be characterized. These include small subunits of SPT (ssSPT) that stimulate SPT activity and ORMs that provide reversible negative-regulation of SPT activity in response to intracellular sphingolipid levels. Contributions of members of Arabidopsis ssSPT and ORM gene families to the regulation of SPT and sphingolipid synthesis will be determined. Using yeast-based tools, post-translational modifications of Arabidopsis ORMs will be assessed and interactions of SPT and regulatory proteins will be determined. Experiments will also be conducted to identify any additional components of the SPT regulatory system. Results from the yeast-based studies will provide the basis for experiments designed to uncover the in planta transduction of SPT regulation in response to perturbations in sphingolipid homeostasis. These studies will take advantage of Arabidopsis mutants with dysfunctions in sphingolipid biosynthetic regulation and fungal mycotoxins that generate enhanced levels of PCD-inducing long-chain bases. Elucidation of the mechanisms responsible for controlling the synthesis of sphingolipids will not only have implications for crop productivity but will also be of broad general interest since sphingolipids are both essential and highly bioactive in all eukaryotic cells.Broader Impacts:The project encompasses graduate and undergraduate training and high school educational outreach. The research will be conducted by graduate students at University of Nebraska-Lincoln (UNL) and the Uniformed Services University of the Health Sciences (USUHS). Opportunities for an annual two-to-four week exchange of graduate students between UNL and USUHS will broaden training experiences for student participants. To meet a growing educational need at UNL, a graduate level, team-taught course in lipid biochemistry will be established under the leadership of PI Cahoon and co-PI Stone. This course will include modules on sphingolipid function and metabolism and will broadly engage students with interests ranging from nutrition, biofuels, and lipid signaling. Undergraduate training will be incorporated into the project through the UNL Undergraduate Creative Activities and Research Experiences (UCARE) program that provides a two year opportunity for undergraduate students to develop and conduct independent research projects. Undergraduate training in line with the project goals will also be provided through established NSF REU programs at UNL and science and technology mentorship programs at USUHS, which actively recruit women and underrepresented minority students. The project will also facilitate educational outreach to high school students through lab-based workshops for the annual UNL Women in Science program and by hosting of high school students in the Nebraska EPSCoR-supported Young Nebraska Scientist Initiative, a summer program designed to strengthen the K-12 STEM student pipeline.

鞘脂有助于蛋白质运输以及膜的组织和物理特性，作为质膜，托管和内膜系统的主要结构成分。鞘脂还充当基本细胞过程的动态调节剂，包括程序性细胞死亡（PCD），这是由生物活性鞘脂代谢物长链碱和神经酰胺的积累触发的。该项目旨在阐明调节组件及其相互作用，可很好地介导足够量的鞘脂脂的需求，以支持细胞生存能力和增长，但阻止了鞘脂代谢物的堆积为诱导PCD的水平。该项目将建立在通过丝氨酸棕榈酰转移酶（SPT）（一种杂二聚体酶）促进该途径中的第一步的杂二二聚体酶（SPT）对鞘脂合成的新兴证据。 SPT监管网络中新玩家的拟南芥同源物将被表征。其中包括SPT（SSSPT）的小亚基，这些亚基刺激SPT活性和ORM，这些ORM可为响应细胞内鞘脂水平提供可逆的SPT活性负调节。将确定拟南芥SSSPT和ORM基因家族成员对SPT和鞘脂合成的调节的贡献。 使用基于酵母的工具，将评估拟南芥ORM的翻译后修饰，并确定SPT和调节蛋白的相互作用。 还将进行实验以确定SPT调节系统的任何其他组件。 基于酵母的研究的结果将为旨在发现SPT调节的planta planta转导，以响应鞘脂稳态的扰动，为实验提供基础。这些研究将利用鞘脂生物合成调节中具有功能障碍的拟南芥突变体和真菌霉菌毒素，从而产生增强的PCD诱导长链碱基水平。阐明负责控制鞘脂合成的机制不仅对农作物生产力有影响，而且还将引起广泛的兴趣，因为鞘脂型在所有真核生物中都是必不可少的，并且具有高度生物活性。 这项研究将由内布拉斯加林肯大学（UNL）和卫生科学大学（USUHS）的研究生进行。在UNL和USUHS之间进行二到四周的研究生交流的机会将扩大学生参与者的培训经验。 为了满足UNL的不断增长的教育需求，将在Pi Cahoon和Co-Pi Stone的领导下建立研究生水平的脂质生物化学课程。 本课程将包括有关鞘脂功能和代谢的模块，并将广泛吸引学生，从营养，生物燃料和脂质信号传导等等。 本科培训将通过ULL本科创意活动和研究经验（UCARE）计划纳入项目，该计划为本科生开发和开展独立研究项目提供了两年的机会。 与项目目标的本科培训还将通过UNL的既定NSF REU计划以及USUHS的科学与技术指导计划提供，该计划积极招募妇女和代表性不足的少数民族学生。 该项目还将通过基于实验室的科学领域的UNL妇女的讲习班以及在内布拉斯加州EPSCOR支持的年轻内布拉斯加州科学家倡议中为高中生的教育宣传，并在内布拉斯加州EPSCOR支持的年轻的Nebraska Progence中招募了一项夏季计划，这是一项旨在加强K-12 STEM学生管道的夏季计划。