A nanoscale, multi-parametric flow cytometry-based platform to study mitochondrial heterogeneity and mitochondrial DNA dynamics
A nanoscale, multi-parametric flow cytometry-based platform to study mitochondrial heterogeneity and mitochondrial DNA dynamics
复制标题
DOI：
10.1038/s42003-019-0513-4
复制
发表时间：
2019-07-11
影响因子：
5.9
通讯作者：
Woods, Dori C.
中科院分区：
文献类型：
--
作者：
MacDonald, Julie A.;Bothun, Alisha M.;Woods, Dori C.
线粒体在能量产生和细胞死亡调节方面的功能已得到很好的表征;然而，对线粒体群体内存在的异质性知之甚少。通常作为由数百万个单独的线粒体组成的合并样品进行分析，关于线粒体亚群之间潜在的不同功能的信息很少。在这里，我们提出了一种新的方法来分析线粒体作为一个复杂的和异质性的网络，使用纳米级和多参数流式细胞仪为基础的平台的单独组件。我们使用多个下游检测来验证该平台，包括电子显微镜、ATP生成、定量质谱蛋白质组学分析和单个细胞器水平的mtDNA分析。这些策略允许对线粒体亚群进行稳健的分析和分离，以更广泛地阐明线粒体的潜在复杂性，因为这些细胞器在细胞内共同发挥作用。
Mitochondria are well-characterized regarding their function in both energy production and regulation of cell death; however, the heterogeneity that exists within mitochondrial populations is poorly understood. Typically analyzed as pooled samples comprised of millions of individual mitochondria, there is little information regarding potentially different functionality across subpopulations of mitochondria. Herein we present a new methodology to analyze mitochondria as individual components of a complex and heterogeneous network, using a nanoscale and multi-parametric flow cytometry-based platform. We validate the platform using multiple downstream assays, including electron microscopy, ATP generation, quantitative mass-spectrometry proteomic profiling, and mtDNA analysis at the level of single organelles. These strategies allow robust analysis and isolation of mitochondrial subpopulations to more broadly elucidate the underlying complexities of mitochondria as these organelles function collectively within a cell.