Exploring subcellular mitochondrial heterogeneity in neurons: a multimodal approach

线粒体在许多基本的生物过程中发挥作用，包括细胞能量的产生，并且已知这些作用在具有特定功能要求的人体不同区域中是不同的。尽管如此，目前还不完全清楚不同细胞类型中的线粒体如何调节其遗传和转录过程，以促进细胞增殖的广泛需求。此外，线粒体功能的异质性在单个细胞的水平上可能更为重要，因为单个细胞的不同部分的能量需求会随着时间的推移而波动。神经元当然也是如此，它们具有高度复杂和极化的结构，在细胞体、沿着轴突和突触中有着非常不同的能量需求，在突触中需要能量来释放神经递质，从而与周围的神经元进行交流。本项目将在单细胞和亚细胞水平上解开线粒体异质性，以更好地了解人脑线粒体功能障碍。为了做到这一点，我们将专注于单神经元中线粒体的转录输出，以及细胞不同区域的单个线粒体。这将涉及高通量RNA测序和遗传数据的计算整合，以更高的分辨率研究核-线粒体相互作用，并将通过在微流体装置中培养人iPSC衍生的神经元来促进，使得不同的细胞隔室（轴突，树突和索马）可以物理分离，并单独分析线粒体。此外，可以使用纳米镊子技术提取单个线粒体并进行单独分析。这也将得到iPSC衍生神经元的遗传操作的支持，以测试线粒体功能输出。总之，我们的目标是确定在亚细胞尺度上影响线粒体功能的特殊转录程序，并了解这些过程与大脑高级功能的相关性。

Mitochondria play a role in many fundamental biological processes, including thegeneration of cellular energy, and these roles are known to vary across differentregions of the human body that have specific functional requirements. Despite this, itis not fully known how mitochondria in different cell types modulate their genetic andtranscriptional processes to facilitate a wide range of demands from the cellularenvironment. Furthermore, heterogeneity in mitochondrial function may be even moreimportant at the level of a single cell, where different compartments of an individualcell fluctuate in their energy demands over time. This is certainly the case withneurons, which have a highly complicated and polarised structure, and have verydifferent energy requirements in the cell body, along axons and at synapses whereenergy is required to release neurotransmitters to communicate with surroundingneurons. This project will disentangle mitochondrial heterogeneity at the single celland subcellular level to better understand mitochondrial dysfunction in the humanbrain. To do this, we will focus on the transcriptional output of mitochondria in singleneurons, as well as for single mitochondria across different regions of a cell. This willinvolve the computational integration of high-throughput RNA sequencing and geneticdata to characterise nuclear-mitochondrial interactions at ever higher resolutions, andwill be facilitated by culturing human iPSC-derived neurons in microfluidic devices,such that different cellular compartments (axons, dendrites and soma) can bephysically separated, and mitochondria within seperately analysed. Additionally singlemitochondria can be extracted using nanotweezer technology and individuallyanalysed. This will also be supported by genetic manipulation of iPSC-derived neuronsto test mitochondrial functional outputs. In all, we aim to identify specialisedtranscriptional programmes that influence mitochondrial function on the sub-cellularscale, and to understand the relevance of these processes to the higher-order functionof the brain.