A new Drosophila-based strategy to study mitochondrial transport and neuronal ageing in vivo.

世界人口正在迅速老龄化。到2047年，60岁及以上人口的数量预计将超过16岁以下儿童和青少年的数量（经社部，《2013年世界人口老龄化》）。老龄化是痴呆症的主要风险因素，许多其他神经元疾病仅在晚年影响个体。与其他与年龄有关的疾病一样，痴呆症的治愈或治疗方法尚不存在。因此，找到改善衰老神经元健康的措施对于减轻与年龄相关疾病相关的日益增加的社会和财政负担至关重要。神经元如何衰老也是一个令人着迷但却知之甚少的智力问题。越来越多的研究表明，细胞成分的正确分布对于确保神经系统在晚年的正常功能至关重要。例如，许多研究表明，在各种神经系统疾病的发病机制中存在细胞器轴突运输缺陷。该领域目前的一个假设是，增加细胞器运输的干预措施将延迟衰老过程中神经元功能障碍的发生。然而，调节老化神经元轴突运输的机制知之甚少，部分原因是缺乏合适的模型进行纵向研究。尽管存在适当的小鼠模型，但由于涉及时间和成本，小鼠中的纵向研究非常具有挑战性。此外，需要手术来允许活小鼠中轴突运输的成像。我最近开发了一种新的方法来详细研究细胞器的细胞内运输，它使用果蝇Drosophila melanogaster。由于该测定利用了半透明翼中神经元的可接近位置，因此该过程是非侵入性的。将果蝇强大的遗传学与延时实时成像相结合，我能够跟踪不同年龄活体动物的细胞器运输。重要的是，果蝇相对较短的寿命使纵向研究成为可能。目前，世界上许多研究小组使用脊椎动物整体动物模型、离体外植体和原代培养来研究轴突运输。我相信，果蝇系统的独特优势意味着，在许多轴突运输和神经元老化的未来研究中，它可以取代脊椎动物模型。我通过操纵线粒体的转运机制来增加这个系统中线粒体的转运，并观察到对年龄依赖性神经元功能障碍的实质性抑制。在我的研究过程中，我还发现了一种进化上保守的信号通路的证据，该通路上调了衰老神经元轴突中的线粒体转运。我研究的主要目的是了解这种特定信号级联与线粒体转运和神经元衰老之间的分子机制。这将首次定义一种信号级联，可以上调衰老神经元中的线粒体转运，从而更好地为未来对抗年龄相关疾病的治疗工作提供信息。为了解决这个问题，我将采取多学科的方法，将果蝇的创新试验与哺乳动物神经元的工作相结合。最初，CRISPR基因组编辑工具（在宿主实验室中优化）将与生物化学和活体果蝇轴突运输的定量延时成像结合使用。然后，将在来自小鼠胚胎干细胞的运动神经元和小鼠体内坐骨神经中验证关键发现。通过在果蝇中进行大部分工作，只需要少量的小鼠。这些动物将被用来测试我的发现的更广泛的相关性，结果可能对人类衰老产生重大的转化影响。

The world population is ageing rapidly. By 2047, the number of people aged 60 and over is expected to exceed the number of children and adolescents aged under 16 (UNDESA, World Population Ageing 2013). Ageing is the main risk factor for dementia and many other neuronal disorders affect individuals only in later life. Contrarily to other age-related diseases, a cure or treatment for dementia is not available. Finding measures to improve the health of ageing neurons is therefore crucial to ease the increasing societal and financial burdens associated with age-related diseases. How neurons age is also a fascinating, and poorly understood, intellectual problem.A growing body of work suggests that correct distribution of cellular constituents is crucial for ensuring proper function of the nervous system in later life. For example, many studies have implicated defective axonal transport of organelles in the pathogenesis of various neurological disorders. A current hypothesis in the field is that interventions that increase transport of organelles would delay the onset of neuronal dysfunction during ageing. However, the mechanisms that regulate axonal transport in ageing neurons are poorly understood, partly because of lack of suitable models to perform longitudinal studies. Although appropriate mouse models exist, longitudinal studies in mice are very challenging because of time and costs involved. In addition, surgery is required to allow imaging of axonal transport in live mice. I recently developed a new method to study in detail the intracellular transport of organelles, which uses the fruit fly Drosophila melanogaster. As this assay exploits the accessible position of neurons in the translucent wing, the procedure is non-invasive. Combining the powerful genetics of Drosophila with time-lapse live imaging, I am able to follow the transport of organelles in live animals of different ages. Importantly, the relatively short lifespan of Drosophila makes longitudinal studies feasible. Many research groups worldwide currently use vertebrate whole animal models, ex vivo explants and primary cultures to study axonal transport. I believe that the unique advantages of the Drosophila system mean that in can replace vertebrate models in many future studies of axonal transport and neuronal ageing.I have discovered a remarkable age-related decline in the axonal transport of mitochondria in wing neurons. I increased transport of mitochondria in this system by manipulating the transport machinery an observed a substantial suppression of age-dependent neuronal dysfunction. During the course of my studies, I also found evidence of an evolutionarily conserved signalling pathway that upregulates mitochondrial transport in axons of ageing neurons. The main aim of my research will be to understand the molecular mechanisms linking this specific signalling cascade to mitochondrial transport and neuronal ageing. This would for the first time define a signaling cascade that could upregulate mitochondrial transport in ageing neurons and hence better inform future therapeutic efforts to combat age-related diseases..To address this question, I will take a multidisciplinary approach by integrating the innovative assay in Drosophila with work in mammalian neurons. Initially, CRISPR genome-editing tools (optimised in the host lab) will be used in combination with biochemistry and quantitative time-lapse imaging of axonal transport in living Drosophila. Key findings will then be validated in motor neurons derived from mouse embryonic stem cells and in sciatic nerves of mice in vivo. By performing much of the work in Drosophila, only a small number of mice will be needed. These animals will be used to test the broader relevance of my findings, with the results potentially may have a significant translational impact on human ageing.