Open AccessQuantifying the inhibitory effect of Bcl‐xl on the action of Mff using live‐cell fluorescence imaging
Author(s) -
Ma Yunyun,
Du Mengyan,
Yang Fangfang,
Mai Zihao,
Zhang Chenshuang,
Qu Wenfeng,
Wang Bin,
Wang Xiaoping,
Chen Tongsheng
Publication year - 2019
Publication title -
febs open bio
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.718
H-Index - 31
ISSN - 2211-5463
DOI - 10.1002/2211-5463.12739
Subject(s) - cytoplasm , mitochondrion , apoptosis , förster resonance energy transfer , microbiology and biotechnology , live cell imaging , fragmentation (computing) , cell , fluorescence lifetime imaging microscopy , mitochondrial fission , fluorescence , cytosol , programmed cell death , chemistry , biophysics , biology , biochemistry , physics , quantum mechanics , enzyme , ecology
Mitochondrial fission regulates mitochondrial function and morphology, and has been linked to apoptosis. The mitochondrial fission factor (Mff), a tail‐anchored membrane protein, induces excessive mitochondrial fission, contributing to mitochondrial dysfunction and apoptosis. Here, we evaluated the inhibitory effect of Bcl‐xl, an antiapoptotic protein, on the action of Mff by using live‐cell fluorescence imaging. Microscopic imaging analysis showed that overexpression of Mff induced mitochondrial fragmentation and apoptosis, which were reversed by coexpression of Bcl‐xl. Microscopic imaging and live‐cell fluorescence resonance energy transfer analysis demonstrated that Bcl‐xl reconstructs the Mff network from punctate distribution of higher‐order oligomers to filamentous distribution of lower‐order oligomers. Live‐cell fluorescence resonance energy transfer two‐hybrid assay showed that Bcl‐xl interacted with Mff to form heterogenous oligomers with 1 : 2 stoichiometry in cytoplasm and 1 : 1 stoichiometry on mitochondria, indicating that two Bcl‐xl molecules primarily interact with four Mff molecules in cytoplasm, but with two Mff molecules on mitochondria.