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Targeted quantitative mass spectrometric identification of differentially expressed proteins between Bax-expressing
Peng Wang1, Andy Lo, J Bryce Young
1Department of Chemistry, University of Alberta, Edmonton, Alberta T6G 2G2, Canada.
Abstract:
Bax, a Bcl-2 interacting protein, plays a central role in several stimuli-induced apoptosis pathways through its functional and physical interactions with various biologically important proteins. Identification of the Bax-modulating protein network should be useful to further our understanding of Bax-mediated apoptosis. For the first time, we performed proteome-wide quantification and identification of differentially expressed proteins between Bax+/- and Bax-/- HCT116 clones using a newly developed quantitative mass spectrometric analysis strategy. This strategy is based on forward and reverse differential isotope labeling of the proteome digests of two comparative cells, followed by two-dimensional liquid chromatography separation and automated peptide deposition to matrix-assisted laser desorption ionization sample plates for MS quantification and MS/MS peptide sequence identification. We quantified and identified 200 differentially expressed proteins involved in various cellular processes. Through bioinformatic analysis, four groups of differentially expressed proteins were highlighted for the association with Bax: mitochondria permeability transition channel proteins, Bax regulator proteins, heat shock protein family members, and oxidative stress-triggered proteins. These results indicate the functional diversity of Bax and provide new research directions to study the biology of Bax-regulated apoptosis.
Insights
Researchers identified proteins that interact with Bax, a key apoptosis regulator. This study advances understanding of Bax-mediated cell death by revealing its protein network, including mitochondrial and oxidative stress proteins.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Bax is a crucial protein in apoptosis pathways, interacting with other proteins.
- Understanding Bax's protein network is vital for studying apoptosis.
Purpose of the Study:
- To identify proteins differentially expressed in Bax+/- and Bax-/- HCT116 cells.
- To elucidate the Bax-modulating protein network for a deeper understanding of Bax-mediated apoptosis.
Main Methods:
- Utilized a novel quantitative mass spectrometry strategy for proteome-wide analysis.
- Employed forward and reverse differential isotope labeling, 2D-LC separation, and MALDI-TOF/TOF MS.
- Quantified and identified differentially expressed proteins between Bax+/-, Bax-/- HCT116 clones.
Main Results:
- Identified and quantified 200 differentially expressed proteins.
- Bioinformatic analysis highlighted four key protein groups associated with Bax: mitochondria permeability transition channel proteins, Bax regulators, heat shock proteins, and oxidative stress-triggered proteins.
Conclusions:
- Bax exhibits functional diversity, influencing multiple cellular processes.
- These findings offer new research avenues into the biology of Bax-regulated apoptosis and its associated protein interactions.
