1. Mito-priming as a method to engineer Bcl-2 addiction.
- Author
-
Lopez J, Bessou M, Riley JS, Giampazolias E, Todt F, Rochegüe T, Oberst A, Green DR, Edlich F, Ichim G, and Tait SW
- Subjects
- Animals, Apoptosis physiology, Apoptosis Regulatory Proteins genetics, Apoptosis Regulatory Proteins metabolism, CRISPR-Cas Systems, Cell Line, Gene Expression Regulation, Genetic Engineering, Humans, Mice, Proto-Oncogene Proteins c-bcl-2 genetics, Mitochondrial Membranes physiology, Peptide Fragments metabolism, Proto-Oncogene Proteins metabolism, Proto-Oncogene Proteins c-bcl-2 metabolism
- Abstract
Most apoptotic stimuli require mitochondrial outer membrane permeabilization (MOMP) in order to execute cell death. As such, MOMP is subject to tight control by Bcl-2 family proteins. We have developed a powerful new technique to investigate Bcl-2-mediated regulation of MOMP. This method, called mito-priming, uses co-expression of pro- and anti-apoptotic Bcl-2 proteins to engineer Bcl-2 addiction. On addition of Bcl-2 targeting BH3 mimetics, mito-primed cells undergo apoptosis in a rapid and synchronous manner. Using this method we have comprehensively surveyed the efficacy of BH3 mimetic compounds, identifying potent and specific MCL-1 inhibitors. Furthermore, by combining different pro- and anti-apoptotic Bcl-2 pairings together with CRISPR/Cas9-based genome editing, we find that tBID and PUMA can preferentially kill in a BAK-dependent manner. In summary, mito-priming represents a facile and robust means to trigger mitochondrial apoptosis.
- Published
- 2016
- Full Text
- View/download PDF