Thioredoxin, a master regulator of the tricarboxylic acid cycle in plant mitochondria
Thioredoxin, a master regulator of the tricarboxylic acid cycle in plant mitochondria
复制标题
DOI：
10.1073/pnas.1424840112
复制
发表时间：
2015-03-17
影响因子：
11.1
通讯作者：
Fernie, Alisdair R.
中科院分区：
文献类型：
--
作者：
Daloso, Danilo M.;Mueller, Karolin;Fernie, Alisdair R.
植物线粒体具有完全可操作的三羧酸（TCA）循环，其在产生ATP和为异养和光合组织中的一系列生物合成过程提供碳骨架方面发挥核心作用。循环酶编码基因在转录和效应子介导的调控方面已经得到很好的表征，并且也进行了反向遗传分析。然而，尽管有如此多的关注，一个中心问题仍然没有答案：“是什么调节了体内通过这一途径的流量？“先前对拟南芥的蛋白质组学实验已经揭示了许多线粒体酶，包括TCA循环和附属途径的成员，具有硫氧还蛋白（TRX）结合位点，并且可能受到氧化还原调节。我们已经跟进了这种可能性，并发现TRX是一个氧化还原敏感的TCA循环流量的调解人。在这项研究中，我们首先在酶和代谢物水平上表征了拟南芥线粒体TRX途径的突变体：NADP-TRX还原酶a和B双突变体（ntra ntrB）和位于细胞内的硫氧还蛋白o 1（trxo 1）突变体。这些研究之后是当C-13-葡萄糖、C-13-苹果酸或C-13-丙酮酸作为底物提供给突变体或WT植物的叶时同位素再分布的比较评估。在一个互补的方法中，我们评估了一系列TCA循环和相关酶在不同氧化还原状态下的体外活性。组合数据集表明，TRX可以使线粒体琥珀酸脱氢酶和琥珀酸脱氢酶失活，并在体内激活胞质ATP-柠檬酸裂解酶，作为通过TCA循环的碳流的直接调节剂，并提供协调细胞功能的机制。
Plant mitochondria have a fully operational tricarboxylic acid (TCA) cycle that plays a central role in generating ATP and providing carbon skeletons for a range of biosynthetic processes in both heterotrophic and photosynthetic tissues. The cycle enzymeencoding genes have been well characterized in terms of transcriptional and effector-mediated regulation and have also been subjected to reverse genetic analysis. However, despite this wealth of attention, a central question remains unanswered: "What regulates flux through this pathway in vivo?" Previous proteomic experiments with Arabidopsis discussed below have revealed that a number of mitochondrial enzymes, including members of the TCA cycle and affiliated pathways, harbor thioredoxin (TRX)-binding sites and are potentially redox-regulated. We have followed up on this possibility and found TRX to be a redox-sensitive mediator of TCA cycle flux. In this investigation, we first characterized, at the enzyme and metabolite levels, mutants of the mitochondrial TRX pathway in Arabidopsis: the NADP-TRX reductase a and b double mutant (ntra ntrb) and the mitochondrially located thioredoxin o1 (trxo1) mutant. These studies were followed by a comparative evaluation of the redistribution of isotopes when C-13-glucose, C-13-malate, or C-13-pyruvate was provided as a substrate to leaves of mutant or WT plants. In a complementary approach, we evaluated the in vitro activities of a range of TCA cycle and associated enzymes under varying redox states. The combined dataset suggests that TRX may deactivate both mitochondrial succinate dehydrogenase and fumarase and activate the cytosolic ATP-citrate lyase in vivo, acting as a direct regulator of carbon flow through the TCA cycle and providing a mechanism for the coordination of cellular function.