Arabidopsis 2010: Functional Analysis of Pollen Exine Assembly

本项目将鉴定构建花粉外壁（外壁）所需的基因。外皮无与伦比的强度和耐化学性对花粉的存活至关重要。外壁在雄性和雌性细胞之间的相互作用中也起着重要作用：花粉和花的雌性器官之间发生的物种特异性粘附需要外壁。Exine是由孢子花粉制成的，孢子花粉是一种异常坚固、化学惰性和独特图案的生物聚合物，可能对材料科学工业有用。因此，它在授粉中的重要性，对聚合物化学的影响，以及作为接触粘合剂的用途，使得对其组成的理解成为当务之急。外叶的惰性和不规则性使化学分析变得混乱，但最近的拟南芥遗传调查更有希望，揭示了外叶结构和功能所需的基因和途径。普鲁斯、萨姆纳、埃德隆德和斯旺森实验室的工作将扩展这些努力。本项目解决了2010年拟南芥基因功能分析的计划目标。它将评估特定基因在胞外组装，模式和粘附中的作用，并将这些基因分类为遗传和代谢途径。拟南芥建立正常外叶形态所需的基因将通过对候选基因中约250个插入的视觉表征以及对新突变体的大规模筛选来确定。目标基因是在a)生物合成途径中选择的，这些途径先前被化学或遗传分析暗示在胞外合成中起作用；B)在胞外合成中未确定作用的途径；C)类似于那些在外部发育中具有已知功能的基因；D)基因，其表达模式表明它们在花粉发育过程中可能在花药中活跃。正在研究的基因和研究进展将在http://preuss.bsd.uchicago.edu/nsf2010.html上提供。数据一般每半年发布一次。对于每一个突变体鉴定，显微镜和生化分析将被用来表征突变基因的作用。遗传和生化网络将通过表征外叶基因表达的时间，检查花粉发育过程中的代谢物积累，并分析双突变体来澄清。这项工作将影响多个学科，提高对以下方面的理解：1)介导授粉和作物育种的基因；2)外叶多样性和植物物种形成的进化控制；3)生物聚合物自组装，使研究人员能够在体外概括外叶发育的酶学步骤，最终设计出具有所需特征的外叶壁；4)有助于花粉粘附的外叶部分；这可能会合成高度特异性的接触黏合剂，其中有250个基因可能在外阴发育中发挥作用。教育和人才培训将是这个项目的一个组成部分。两名技术人员和至少4名本科生（包括2名来自主要的本科院校-斯佩尔曼和瓦尔帕莱索）将接受培训。学生，包括那些历史上在科学领域代表性不足的学生，将有机会进行暑期研究，发展他们在计划和执行实验方面的经验，并在研究会议上展示他们的工作。他们的导师将为他们的实验室和课程提供信息。此外，博士后将通过指导技术人员和本科生进行研究和指导培训，使其能够过渡到独立的职业生涯。

This project will identify genes required to construct the outer pollen wall (exine). The unparalleled strength and chemical resistance of exine is important for pollen survival. This wall also plays an important role in interactions between male and female cells: exine is required for the species-specific adhesion that takes place between pollen and the female organs of the flowers. Exine is made of sporopollenin, an unusually strong, chemically inert and distinctively patterned biopolymer that may prove useful for the materials science industry. Thus, its importance in pollination, implications for polymer chemistry, and utility as a contact adhesive make an understanding of its composition a high priority. The inert and irregular nature of exine has confounded chemical analysis, but recent Arabidopsis genetic surveys are more promising, revealing genes and pathways required for exine structure and function. The work of the Preuss, Sumner, Edlund, and Swanson labs will extend these efforts. This project addresses the 2010 program goal of functional analysis of every Arabidopsis gene. It will assess the role of specific genes in exine assembly, patterning and adhesion, and will sort these genes into genetic and metabolic pathways. Arabidopsis genes required for establishing normal exine morphology will be defined by visually characterizing ~250 insertions in candidate genes, as well as by performing a large-scale screen for novel mutants. The targeted genes are selected among a) biosynthetic pathways previously implicated by chemical or genetic analyses as playing a role in exine synthesis; b) pathways with an unresolved role in exine synthesis; c) genes resembling those with a known function in exine development; d) genes, whose expression pattern indicates they are likely active in anthers during pollen development. The genes under study and progress of the research will be available at http://preuss.bsd.uchicago.edu/nsf2010.html. Data will be released generally on a semi-annual basis. For each of the mutants identified, microscopic and biochemical assays will be employed to characterize the roles of the mutated genes. Genetic and biochemical networks will then be clarified through characterizing the timing of exine gene expression, examining metabolite accumulation during pollen development, and analyzing double mutants. This work will impact multiple disciplines, improving the understanding of 1) genes that mediate pollination and crop breeding, 2) evolutionary control over exine diversity and plant speciation, 3) biopolymer self-assembly to allow investigators to recapitulate enzymatic steps of exine development in vitro, ultimately resulting in designing exine walls with desired characteristics, 4) exine moieties that contribute to pollen adhesion, possibly allowing the synthesis of highly specific contact adhesives, 5) 250 genes that could play a role in exine development. Education and personnel training will be an integral part of this project. Two technicians and at least 4 undergraduates (including 2 from the primarily undergraduate institutions - Spelman and Valparaiso) will be trained. Students, including those historically underrepresented in science, will have an opportunity for summer research, developing their experience in planning and performing experiments and presenting their work at research conferences. Their mentors will generate information to incorporate in their laboratories and courses. In addition, postdoctoral fellows will be trained in research and mentoring via supervising technicians and undergraduates, enabling transitions to independent careers.