Interaction of ubiquinone & complex I control longevity

描述(申请人提供)：这项工作的长期目标是在分子水平上了解线粒体功能如何影响衰老过程。我们正在研究两种线虫株，它们的突变影响线粒体功能，并改变了寿命。这些突变菌株CLK-1和GAS-1降低了电子传递链复合体I的功能。然而，它们在寿命、对氧气的敏感性以及对线粒体蛋白质的氧化损伤积累方面存在差异。这两个菌株之间在自由基损伤方面的差异是令人好奇的，因为这两个突变都影响泛醌与络合物I的相互作用。已经证明，泛醌与络合物n的相互作用在自由基形成中发挥作用，从而影响寿命。突变的clk-1缺乏泛醌的合成，泛醌是络合物I和II的电子受体。为了生存，这种突变菌株必须从它赖以生存的细菌中获得它的泛醌。然而，CLK-1从用作食物的常见形式的大肠杆菌中获得的泛醌与普通线虫泛醌(即Q8而不是Q9)不同，并延长了菌株的寿命。我们发现，络合物I和n使用新泛醌的能力不同；络合物n功能正常，而络合物I被严重抑制。第二个突变GAS-1影响复合体I的49 kDa亚基，并形成复合体I的泛醌结合位点的一部分。当我们构建双突变体CLK-1/GAS-1时，得到的动物是不育的，但存活时间是正常的两倍(34天对15天)。因此，当GAS-1暴露在Q8中时，它的寿命会延长。我们假设泛醌与复合体I的相互作用直接影响呼吸作用，继而影响ROS的产生。本应用的具体目的是：1.测量将外源泛醌替换到N_2、Clk-1和GAS-1的线粒体中对呼吸的影响；2.确定线粒体从N_2、GAS-1和ClK-1产生ROS的位置和速率；3.展示泛醌对线粒体蛋白质氧化的影响；4.鉴定抑制GAS-1寿命缩短的两个突变体seg-1和seg-2。这些结果将首次将泛醌对呼吸的影响与对寿命的影响联系起来。

DESCRIPTION (provided by applicant): The long-term goal of this work is to understand at the molecular level how mitochondrial function affects the aging process. We are studying two strains of C. elegans with mutations affecting mitochondrial function and which have altered lifespans. These mutant strains, clk-1 and gas-1, decrease the function of complex I of the electron transport chain. However, they differ in lifespan, sensitivity to oxygen, and accumulation of oxidative damage to mitochondrial proteins. The difference in free radical damage between the two strains is curious, since the mutations both affect the interaction of ubiquinone with complex I. The role of ubiquinone interaction with complex n has already been shown to play a role hi free radical formation with resulting effects on lifespan. The mutant clk-1 is deficient in the synthesis of ubiquinone, the electron acceptor for complexes I and ll. To survive, this mutant strain must obtain its ubiquinone from the bacteria on which it feeds. However, the ubiquinone which clk-1 obtains from the common form of E. coli used as food is different than the normal nematode ubiquinone (i.e. Q8 instead of Q9) and conveys a prolonged lifespan to the strain. We found that complex I and n differ in their ability to use the new ubiquinone; complex n functions normally while complex I is severely inhibited. The second mutation, gas-1, affects the 49 kDa subunit of complex I and forms part of the ubiquinone binding site of complex I. When we constructed the double mutant clk-1/gas-1 the resulting animals are sterile but live over twice as long as normal (34 days vs. 15 days). Thus, when gas-1 is exposed to Q8, it develops a prolonged lifespan. We hypothesize that the ubiquinone interaction with complex I directly affects respiration with secondary effects on ROS production. The specific aims of this application are to: 1. Measure the effects on respiration of substituting exogenous ubiquinones into mitochondria of N2, clk-1 and gas-1; 2. Determine the sites and rates of ROS production by mitochondria from N2, gas-1, and clk-1; 3. Demonstrate the resulting effects of the ubiquinones on oxidation of mitochondrial proteins; 4. Characterize two mutants, seg-1 and seg-2, that suppress the shortened lifespan of gas-1. These results will, for the first time, allow a correlation of the effects of ubiquinone on respiration with those on lifespan.