CAREER: genetic approach to identifying proteins necessary for division plane orientation during plant development

职业：通过遗传方法识别植物发育过程中分割平面方向所需的蛋白质

• 批准号: 1350874
• 负责人: Kent Chapman
• 金额: $ 59.98万
• 依托单位: University of North Texas
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2020-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1350874&HistoricalAwards=false
• 关键词: CAREER; genetic; approach; identifying; proteins

Amanda Joy WrightIOS-1350874CAREER: A genetic approach to identifying proteins necessary for division plane orientation during plant development During plant development, cells divide and grow creating the tissues and organs that make up the adult plant. The process of cell division produces new cells by the cleavage of a pre-existing cell called a mother cell. Mother cells undergo cell divisions with different orientations such as longitudinal or transverse and equal or unequal. Since plant cells are surrounded by rigid cell walls, once they are generated, plant cells are unable to adjust their position relative to neighboring cells. Thus, ensuring that the mother cell undergoes a cell division in the correct orientation is critical for normal plant growth. To fully understand cell division orientation, this proposal lays out a plan to identify the proteins that regulate and execute this process. This research will be conducted in Zea mays (corn), an agriculturally important crop plant. The results of this project will allow for more effective engineering of crops for maximum growth and yield in addition to advancing understanding of a fundamental cellular process. Importantly, previous research has shown that many factors needed for plant cell division are also needed for animal cell division meaning this research is likely to advance our understanding of cell division in many organisms. Finally, during the course of this project, a Cell Biology lab module that allows every biology major at the University of North Texas to participate in a novel scientific investigation will be implemented and evaluated for effectiveness at promoting student learning. The final output of this project will be the identification and characterization of proteins needed for normal plant growth and development. The first objective of the proposal is to characterize the protein disrupted in the Zea mays dcd2 mutant. In this maize mutant, the partitioning cell wall that forms during critical cell divisions in the leaf is misoriented resulting in abnormal cell shapes. The methods that will be used to identify the causative mutation include a combination of positional cloning and RNA-seq while the localization of the DCD2 protein and further characterization of the phenotype will be accomplished by observing fluorescent fusion proteins and immunolocalization. The second objective is to isolate additional maize mutants that form abnormal division planes, roughly map five of the new mutants, and determine the causative mutation in two mutants. The methods that will be used to accomplish this objective include conducting a genetic screen, using RNA-seq to identify the position of the mutations within the genome, and characterizing the mutant phenotype by observing fluorescent fusion proteins and immunolocalization. The third objective is to integrate improvements in science education at the University of North Texas with further characterization of the proteins needed for division plane orientation in plants. To accomplish this objective, students in the Cell Biology Lab will conduct novel research in a classroom setting by conducting a yeast two-hybrid screen to identify proteins that interact with the proteins identified during first two objectives of the project.

在植物发育过程中，细胞分裂和生长创造了构成成年植物的组织和器官。细胞分裂的过程是通过分裂先前存在的称为母细胞的细胞产生新细胞。母细胞分裂的方向不同，如纵向或横向、等向或不等向。由于植物细胞被坚硬的细胞壁包围，一旦产生，植物细胞就无法调整相对于邻近细胞的位置。因此，确保母细胞在正确的方向上进行细胞分裂对植物的正常生长至关重要。为了充分了解细胞分裂方向，本提案提出了一个计划，以确定调节和执行这一过程的蛋白质。这项研究将在玉米（一种重要的农业作物）上进行。这个项目的结果将允许更有效的作物工程，以最大限度地提高生长和产量，除了推进对基本细胞过程的理解。重要的是，先前的研究表明，植物细胞分裂所需的许多因素也需要动物细胞分裂，这意味着这项研究可能会促进我们对许多生物细胞分裂的理解。最后，在这个项目的过程中，将实施一个细胞生物学实验室模块，该模块允许北德克萨斯大学的每个生物学专业的学生参与一项新的科学研究，并评估其在促进学生学习方面的有效性。该项目的最终成果将是鉴定和表征正常植物生长发育所需的蛋白质。该提案的第一个目标是表征玉米梅斯dcd2突变体中被破坏的蛋白质。在这个玉米突变体中，在叶片关键细胞分裂期间形成的分裂细胞壁定向错误，导致细胞形状异常。鉴定致病突变的方法将包括定位克隆和RNA-seq相结合，而DCD2蛋白的定位和进一步的表型表征将通过观察荧光融合蛋白和免疫定位来完成。第二个目标是分离出其他形成异常分裂面的玉米突变体，大致绘制出5个新突变体的图谱，并确定其中两个突变体的致病突变。将用于实现这一目标的方法包括进行遗传筛选，使用RNA-seq识别基因组中突变的位置，以及通过观察荧光融合蛋白和免疫定位来表征突变表型。第三个目标是将北德克萨斯大学科学教育的改进与进一步表征植物分裂面定位所需的蛋白质结合起来。为了实现这一目标，细胞生物学实验室的学生将在课堂上进行新的研究，通过酵母双杂交筛选来识别与项目前两个目标中确定的蛋白质相互作用的蛋白质。