Identification of new regulators of mitochondrial fusion using a novel quantitative bi-molecular complementation assay. (a) Mitochondria from two cell lines expressing either the amino-terminal part of luciferase or its carboxy-terminal part are isolated. (b) Upon mixing of both populations, mitochondrial fusion occurs, leading to the reconstitution of the luciferase into a functional protein. The emission of light is quantified with a plate reader and is proportional to the amount of mitochondrial fusion. (c) Several parameters of the assay can be modified. First, one or both of the cell lines from which mitochondria are isolated can be pre-treated - for example, with chemicals (for example, forskolin) or RNA interference (RNAi; for example, PKA). Then, cytosol from different sources can be added to the fusion mixture. At the same time, different chemicals can be included in the mixture, which gives rise to the possibility to perform high throughput screens for new modulators of mitochondrial dynamics.
Jourdain and Martinou BMC Biology 2010 8:99 doi:10.1186/1741-7007-8-99