Investigating the relationship between respiratory chain organisation and mitochondrial morphology by electron cryo-tomography

线粒体在公共领域很突出，突发新闻报道经常报道线粒体功能与其主要作用之间的新发现联系-利用储存在食物中的能量以ATP的形式为细胞生存提供动力。越来越多的证据表明，线粒体功能障碍与衰老和神经退行性疾病（如帕金森病）有关。呼吸酶体是一个巨大的分子机器，在线粒体内膜中进行细胞呼吸。它由四个复合物（I-IV）组成，它们将电子从NADH和琥珀酸转移到分子氧。通过这一过程获得的能量用于泵送质子穿过线粒体内膜，导致用于产生ATP的膜电位。大约50%的所有线粒体疾病影响能量代谢可以追溯到突变的一个亚基的复合物I.在令人兴奋的新数据，我们已经发现，一个辅助蛋白的复合物I起着关键作用的稳定性的线粒体。这种蛋白质的敲除显示对呼吸、活性氧（ROS）的产生和内膜电位的降低的额外有害影响。通过尖端的成像技术，我们还观察到线粒体形态受到干扰，这可能对网络形成和内膜中蛋白质复合物的结构产生额外的影响。在这个多学科的项目中，最先进的成像方法包括荧光显微镜和高分辨率电子冷冻显微镜（cryoEM），将用于研究线粒体网络的破坏以及对我们的复合物I突变体中蛋白质结构的影响。这将与生物化学实验相结合，以探索电子转移装置的结构和功能的完整性。这些将包括分析呼吸超复合物的组装状态及其电子转移，质子泵和氧摄取活动。还将评估它们限制ROS过度（和破坏性）产生的能力。获得的知识将用于阐明，并指导进一步的研究，与复杂的I功能障碍相关的疾病。

Mitochondria are prominent in the public domain, with breaking news stories often reporting on newly discovered links between mitochondrial function and their primary role- harnessing the energy stored in food to provide the power for cell survival in the form of ATP. There is increasing evidence that relates mitochondrial dysfunction with ageing and neurodegenerative disorders, such as Parkinson's disease. The respirasome is a massive molecular machine that carries out cellular respiration in the mitochondrial inner membrane. It is comprised of four complexes (I-IV), which transfer electrons from NADH and succinate to molecular oxygen. Energy gained through this process is used to pump protons across the inner mitochondrial membrane, leading to a membrane potential that is used to generate ATP. Approximately 50% of all mitochondrial disorders affecting energy metabolism can be traced back to mutations in one of the subunits of complex I.In exciting new data, we have discovered that an accessory protein of complex I plays a key role in the stability of the respirasome. Knock-down of this protein shows additional deleterious effects on respiration, production of reactive oxygen species (ROS) and reduction of the inner membrane potential. By cutting-edge imagining techniques, we have also observed that mitochondrial morphology is perturbed, which likely has additional consequences on network formation and the structure of protein complexes in the inner membrane.In this multi-disciplinary project, state of-the-art imaging methods including fluorescence light microscopy and high-resolution electron cryomicroscopy (cryoEM), will be used to investigate the break-down of mitochondrial networks and the consequence on protein structures in our complex I mutants. This will be combined with biochemical experiments to explore the structural and functional integrity of the electron transfer apparatus. These will include analysis of the assembled state of the respiratory super-complexes alongside their electron transfer, proton-pumping and oxygen uptake activities. Their ability to restrict excessive (and damaging) production of ROS will also be evaluated. Knowledge gained will be used to shed light on, and guide the further study of, diseases associated with complex I dysfunction.