Regulation and Assembly of Pyruvate Dehydrogenase Complexes

呼吸作用是活细胞利用能量做功。生长和繁殖都受到呼吸作用的影响，必须加以仔细控制，以避免生长减少，而对植物来说，则是农业生产力下降。尽管进行了大量的研究，但植物细胞中呼吸是如何控制的细节仍然是一个谜。丙酮酸脱氢酶是一种多组分酶复合物，位于植物细胞的特定亚细胞区室内。它占据了一个十字路口的位置，在那里有多个呼吸成分之间的相互作用。这个复合体的理想位置是在呼吸的整体控制中起主要作用。此外，该综合体的多组件架构允许来自几种不同机制的输入。调控方案的一个要素可能是丙酮酸脱氢酶复合物的内在成分。对这一成分进行详细的生化和分子分析，将为推测的控制机制提供关键的见解。此外，还将采用一种利用整株植物的遗传策略。将使用一种小型模型植物，鼠耳芥。分子遗传学实验将允许完全消除提出的丙酮酸脱氢酶复合体的控制成分。如果控制假说是正确的，不受控制的呼吸作用将导致植物体积小、产量低。通过更换控制部件，这些工厂将得到拯救。本机控制组件和在实验室中修改过的版本都将被使用。这将有助于对控制机制的具体理解。与微生物或动物相比，植物细胞含有丙酮酸脱氢酶复合体的另一种成分的几种不同形式。初步研究表明，情况并不是在不同时间或不同地点有不同版本的功能那么简单。同样，将操纵鼠耳植物，以消除该组件的三个版本中的两个。这些实验将是迭代的和组合的。这意味着生成的植物只包含组件1、组件2、组件3、组件1和组件2、组件1和组件3、组件2和组件3。这些操作将使我们更好地理解每个组件对整个综合体的贡献。丙酮酸脱氢酶复合体的第三个组成部分一直是个谜。有相当多的证据表明它的存在，但到目前为止还没有被孤立。试图通过分子遗传学分离该成分的尝试尚未成功。将采取更经典的生化隔离策略。然而，这将是一个基于其他植物和动物实验系统结果的更具体的策略。分离这第三个成分将允许随后的基因分离。一旦完成，那么上述生化和分子策略将被应用。从这些实验中获得的信息将提高对植物生长发育的基本认识。此外，研究结果有可能使研究人员通过改变对植物细胞呼吸的控制来提高农业生产力。最后，研究结果将为通过传统育种或生物技术设计更高效的作物提供信息。

Respiration is the use of energy by living cells to do work. Both growth and reproduction are affected by respiration and it must be carefully controlled to avoid decreased growth and, in the case of plants, reduced agricultural productivity. Despite considerable research, details of how respiration is controlled in plant cells remain a puzzle. Pyruvate dehydrogenase is a multi-component enzyme complex located within a specific sub-cellular compartment of plant cells. It occupies a cross-roads position where there is interaction among multiple components of respiration. This complex is ideally situated to play a major role in the overall control of respiration. Furthermore, the multi-component architecture of the complex allows input from several different mechanisms. One element of the regulatory scheme may be an intrinsic component of the pyruvate dehydrogenase complex. Detailed biochemical and molecular analyses of this component will provide critical insight into the putative mechanism of control. In addition, a genetic strategy that uses whole plants will be employed. A small model plant, mouse-eared cress, will be used. Molecular genetic experiments will allow the complete elimination of the proposed control component of the pyruvate dehydrogenase complex. If the control hypothesis is correct, uncontrolled respiration will result in small, less-productive plants. These plants will be rescued by replacing the control component. Both the native control component and versions that have been modified in the laboratory will be used. This will allow specific understanding of the mechanism of control. In contrast to microbes or animals, plant cells contain several different versions of another component of the pyruvate dehydrogenase complex. Preliminary studies have shown that the situation is not so simple as having different versions functioning at different times or in different places. Again, the mouse-eared cress plant will be manipulated so as to eliminate two of the three versions of this component. These experiments will be iterative and combinatorial. This means that plants will be generated that contain only component 1, only 2, only 3, 1 and 2, 1 and 3, and 2 and 3. These manipulations will allow a better understanding of the contribution that each component makes to the whole complex. A third component of the pyruvate dehydrogenase complex has been an enigma. There is considerable evidence for its existence, but it has thus far not been isolated. Attempts to isolate this component by molecular genetics have not been successful. The return to a more classical biochemical isolation strategy will be undertaken. However it will be a more specific strategy based upon results from other plant and animal experimental systems. Isolation of this third component will allow subsequent isolation of the gene. Once this is accomplished, then the biochemical and molecular strategies described above will be applied. The information gained from these experiments will improve basic understanding of plant growth and development. Furthermore, there is the potential that the results will allow researchers to increase agricultural productivity by altering the control of plant cell respiration. Finally, the results will inform the design of more efficient crop plants through classical breeding or biotechnology.