线粒体融合
线粒体分裂
线粒体
细胞生物学
生物
活体细胞成像
绿色荧光蛋白
细胞器
线粒体基质
荧光显微镜
融合蛋白
线粒体DNA
细胞
胞浆
荧光
生物化学
物理
基因
重组DNA
量子力学
酶
作者
Mariusz Karbowski,Megan M. Cleland,Brian A. Roelofs
出处
期刊:Methods in Enzymology
日期:2014-01-01
卷期号:: 57-73
被引量:31
标识
DOI:10.1016/b978-0-12-801415-8.00004-7
摘要
Technological improvements in microscopy and the development of mitochondria-specific imaging molecular tools have illuminated the dynamic rearrangements of these essential organelles. These rearrangements are mainly the result of two opposing processes: mitochondrial fusion and mitochondrial fission. Consistent with this, in addition to mitochondrial motility, these two processes are major factors determining the overall degree of continuity of the mitochondrial network, as well as the average size of mitochondria within the cell. In this chapter, we detail the use of advanced confocal microscopy and mitochondrial matrix-targeted photoactivatable green fluorescent protein (mito-PAGFP) for the investigation of mitochondrial dynamics. We focus on direct visualization and quantification of mitochondrial fusion and mitochondrial network complexity in living mammalian cells. These assays were instrumental in important recent discoveries within the field of mitochondrial biology, including the role of mitochondrial fusion in the activation of mitochondrial steps in apoptosis, participation of Bcl-2 family proteins in mitochondrial morphogenesis, and stress-induced mitochondrial hyperfusion. We present some basic directions that should be helpful in designing mito-PAGFP-based experiments. Furthermore, since analyses of mitochondrial fusion using mito-PAGFP-based assays rely on time-lapse imaging, critical parameters of time-lapse microscopy and cell preparation are also discussed.
科研通智能强力驱动
Strongly Powered by AbleSci AI