RUI: Collaborative Research: SAC9, A Novel, Plant-Specific Phosphoinositide Essential for Membrane/Cytoskeleton Dynamics in Arabidopsis

RUI：合作研究：SAC9，一种对拟南芥膜/细胞骨架动力学至关重要的新型植物特异性磷酸肌醇

• 批准号: 0717904
• 负责人: Daryll Dewald
• 金额: $ 18.74万
• 依托单位: Utah State University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2010-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0717904&HistoricalAwards=false
• 关键词: RUI; Collaborative; Research; SAC9; Novel

Understanding plant cell lipid signaling is critical for understanding the regulation of many aspects of plant physiology. Phosphoinositides (PIs) play critical roles in plant responses to stress, vascular development, guard cell dynamics and cell growth and morphology. Plants have evolved novel mechanisms of PI signaling and novel PI modifying pathways that are distinct from those in animals and yeast. Studies undertaken in this project will lead to new insights into the function of PIs in plants cells and new tools and methods that will greatly enhance study of plant PI signaling, and ultimately lead to the development of novel approaches for engineering plants with greater tolerances to abiotic and biotic stresses. Primarily through their interactions with proteins, PIs regulate functions including vesicle trafficking and membrane/cytoskeleton interactions. PI accumulation is regulated in part by the action of PI phosphatases, including the Sac-domain proteins. Sac-domain proteins are conserved across eukaryotes, but a novel distinct family member, SAC9, is found only in plants and green algae. Plant and algal SAC9 proteins have many conserved differences from the other Sac-domain proteins, including the other Sac-domain proteins found in plants. Dr. Williams's group's initial studies have demonstrated that SAC9 is important for regulation of both PtdIns(4,5)P2 levels and actin cytoskeleton distribution in plants. Further characterization of the novel SAC9 protein, and phenotypes of sac9 mutant plants, will both inform studies on Sac-domain proteins across eukaryotes, and contribute to an understanding of the interactions between PIs, the actin cytoskeleton and plant cell functions. This project will (1) characterize the activity and cellular localization of AtSAC9 through in vitro and in vivo expression; (2) characterize the roles of the plant-specific C-terminal domain and SAC9-specific WW domain in AtSAC9 function and cellular distribution; (3) examine SAC9 function in vivo through transient and inducible expression studies; and (4) test our model of SAC9 function by examining co-localization of SAC9, PtdIns(4,5)P2 and actin filaments using fluorescent proteins and immunolocalization.Broader ImpactBoth principal investigators have extensive experience and demonstrated success conducting research with undergraduates, and believe that providing exciting research opportunities to undergraduates in the classroom and the research lab is instrumental for bringing them into the science pipeline. Drs. Williams and Dewald have developed ways that ensure the undergraduates in their laboratories will be trained technically in sophisticated approaches and will participate in research design and execution. The research will be carried out principally by undergraduate students at both Harvey Mudd College (HMC) and Utah State University (USU). One graduate student at USU will collaborate with the undergraduates and participate in undergraduate teaching. This project builds on an ongoing and successful collaboration that has involved undergraduates at both HMC and USU.

了解植物细胞脂质信号对于了解植物生理学的许多方面的调节是至关重要的。磷脂酰肌醇在植物逆境应答、维管发育、保卫细胞动态、细胞生长和形态等方面起着重要作用。植物已经进化出与动物和酵母中的PI信号传导和PI修饰途径不同的新机制。在这个项目中进行的研究将导致新的见解PI在植物细胞中的功能和新的工具和方法，这将大大提高植物PI信号的研究，并最终导致开发新的方法，工程植物具有更大的耐受性，以非生物和生物胁迫。PI主要通过与蛋白质的相互作用调节囊泡运输和膜/细胞骨架相互作用等功能。PI积累部分地由PI磷酸酶的作用调节，包括Sac结构域蛋白。Sac-domain蛋白在真核生物中是保守的，但是一个新的独特的家族成员，SAC 9，仅在植物和绿色藻类中发现。植物和藻类SAC 9蛋白与其他Sac-domain蛋白（包括在植物中发现的其他Sac-domain蛋白）具有许多保守差异。威廉姆斯博士小组的初步研究表明，SAC 9对调节植物中PtdIns（4，5）P2水平和肌动蛋白细胞骨架分布都很重要。进一步表征的新的SAC 9蛋白，和表型的sac 9突变体植物，都将通知跨真核生物的Sac-domain蛋白质的研究，并有助于PI之间的相互作用的理解，肌动蛋白细胞骨架和植物细胞功能。本项目将（1）通过体外和体内表达来表征AtSAC 9的活性和细胞定位;（2）表征植物特异性C-末端结构域和SAC 9特异性WW结构域在AtSAC 9功能和细胞分布中的作用;（3）通过瞬时和诱导表达研究来检测SAC 9的体内功能;（4）研究植物特异性C-末端结构域和SAC 9特异性WW结构域在AtSAC 9功能和细胞分布中的作用。和（4）通过使用荧光蛋白和免疫定位检测SAC 9、PtdIns（4，5）P2和肌动蛋白丝的共定位来测试我们的SAC 9功能模型。更广泛的影响两位主要研究者都有丰富的经验，并成功地与本科生进行了研究，并认为，提供令人兴奋的研究机会，在课堂上和研究实验室的本科生是有助于把他们纳入科学管道。威廉姆斯博士和德瓦尔德博士已经开发了一些方法，确保他们实验室的本科生在技术上接受复杂方法的培训，并参与研究设计和执行。这项研究将主要由哈维穆德学院（HMC）和犹他州州立大学（USU）的本科生进行。USU的一名研究生将与本科生合作并参与本科教学。这个项目建立在一个正在进行的和成功的合作，涉及在HMC和USU的本科生。