Chloroplast Starch Metabolism: A New Regulatory Junction for Redox and Protein Phosphorylation

科学研究表明，在叶绿体对二氧化碳的光合作用中形成的终极储存分子。淀粉在白天积累，晚上被降解成中间产物，然后输出到异养器官。虽然动物体内糖原的生物合成和降解是由蛋白质磷酸化通过教科书中的机制来调节的，但日循环控制暂时性叶绿体淀粉的生物合成和降解的机制长期以来一直是一个谜。在最近的一项研究中，该小组获得了双特异性蛋白磷酸酶DSP4在白天与淀粉颗粒结合，在晚上解离的证据。DSP4基因的破坏导致了突变体拟南芥植株中淀粉水平的急剧增加。与光和暗相关的两个调节因子，即pH和氧化还原状态改变了DSP4的活性和淀粉结合能力。结果进一步表明，DSP4在白天是颗粒结合磷酸酶活性的主要部分，而在晚上不是。所采用的策略表明，DSP4在调节淀粉积累的过程中，在光诱导的氧化还原变化和蛋白质磷酸化之间起到了桥梁作用。这也是第一个在蛋白质磷酸化和淀粉代谢之间建立功能联系的研究，为进一步了解植物淀粉平衡的机制提供了垫脚石。本项目将：(1)确定光介导的氧化还原系统对DSP4的调控机制；(2)鉴定潜在的靶标GSK3样激酶及其在淀粉代谢中的功能；(3)探索淀粉颗粒上的蛋白质组并确定哪些蛋白质被蛋白质磷酸化修饰并受DSP4调控；(4)确定其他与DSP4相关的蛋白磷酸酶是否在淀粉代谢中发挥作用。从这些研究中获得的信息将是理解叶绿体淀粉代谢昼夜调节背后的分子机制的基础。广泛影响除了对基础科学机构做出贡献外，这项研究还将影响社会和教育。与该项目相关的本科生将通过Co-Pi实验室正在进行的多机构项目，将他们的研究成果应用于食品改良。这项研究旨在改善高粱的营养特性，高粱是世界上最贫穷的人的主食。如果对拟南芥的研究结果可以推广到高粱上，与NSF这项研究相关的本科生的工作可能会对人类和世界农业产生重大影响。这项研究将通过私人投资机构在伯克利教授的主要课程和个人投资机构实验室的独立研究项目，对本科教育产生额外的影响。NSF项目不仅将通过直接的实验室培训，而且还将通过加州大学伯克利分校和中国科学院在植物生物学方面现有的国际合作，大大加强研究生教育，这将对研究生在国际层面的经历产生积极影响。最后，该项目将协助正在进行的以当地高中为重点的努力，以鼓励少数族裔学生获得高等教育并进入生物专业。

Scientific MeritStarch is the ultimate storage molecule formed in the photosynthetic fixation of carbon dioxide by chloroplasts. Starch accumulates during the day and is degraded at night to intermediates that are exported to heterotrophic organs. Although the biosynthesis and degradation of glycogen in animals are regulated by protein phosphorylation via mechanisms that are textbook material, the mechanism by which diurnal cycles control the biosynthesis and degradation of transitory chloroplast starch has long remained a mystery. In a recent study, this group obtained evidence that a dual specificity protein phosphatase, DSP4, binds to starch granules during the day and dissociates at night. Disruption of the DSP4 gene resulted in a dramatic increase in the level of starch in mutant Arabidopsis plants. Two regulatory factors linked to light and dark i.e., pH andredox status changed both the activity and the starch-binding capacity of DSP4. The results further revealed that DSP4 represents a major fraction of granule-bound phosphatase activity during the day but not at night. The strategy used suggests that DSP4 acts as a bridge between light-induced redox changes and protein phosphorylation in the regulation of starch accumulation. This is also the first study that establishes a functional connection between protein phosphorylation and starch metabolism, providing a stepping stone for further understanding the mechanism underlying starch balance in plants. This project will: (1) identify the mechanism of DSP4 regulation by the light-mediated redox system, (2) characterize the potential target GSK3-like kinase and its function in starch metabolism, (3) explore the proteome on the starch granule and determine which proteins are modified by protein phosphorylation and regulated by DSP4, (4) determine whether other DSP4-related protein phosphatases function in starch metabolism. Information gained in these studies will be fundamental to understanding the molecular mechanism underlying diurnal regulation of starch metabolism in the chloroplast.Broader ImpactIn addition to contributing to the body of fundamental science, this research will impact society as well as education. Undergraduate students associated with the project will apply findings made in their research to food improvement via an ongoing multi-institutional project in the Co-PI's laboratory. This research is designed to improve the nutritional properties of sorghum a grain that serves as a major food staple for the world's poorest people. If the findings made with Arabidopsis can be extended to sorghum, the work of undergraduates affiliated with this NSF research could have significant impact on humanity and world agriculture. The research will have an additional effect on undergraduate education through major courses the PIs teach at Berkeley and through independent research programs in PIs' laboratories. The NSF project will also significantly enhance graduate education not only through direct lab training but also through the existing international collaboration in plant biology between the Unversity of California, Berkeley and the Chinese Academy of Sciences, which will positively influence the experience of graduate students at the international level. Finally, the project will assist ongoing efforts focused on local high schools to encourage minority students to obtain a higher education and enter biology.