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Profiling Thiol Redox Proteome Using Isotope Tagging Mass Spectrometry
Published on: March 24, 2012
Identification of potential protein targets of isothiocyanates by proteomics
Lixin Mi1, Brian L Hood, Nicolas A Stewart
1Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University, 3800 Reservoir Road, Washington, D.C. 20057, United States.
Abstract:
Isothiocyanates (ITCs), such as phenethyl isothiocyanate (PEITC) and sulforaphane (SFN), are effective cancer chemopreventive compounds. It is believed that the major mechanism for the cancer preventive activity of ITCs is through the induction of cell cycle arrest and apoptosis. However, the upstream molecular targets of ITCs have been underexplored until recently. To identify proteins that are covalently modified by ITCs, human non-small cell lung cancer A549 cells were treated with (14)C-PEITC and (14)C-SFN, and the cell lysates were extracted for analysis by 2-D gel electrophoresis and mass spectrometry. After superimposing the colloidal Coomassie blue protein staining pattern with the pattern of radioactivity obtained from X-ray films, it was clear that only a small fraction of cellular proteins contained radioactivity, presumably resulting from selective binding with PEITC or SFN via thiocarbamation. More than 30 proteins with a variety of biological functions were identified with high confidence. Here, we report the identities of these potential ITC target proteins and discuss their biological relevance. The discovery of the protein targets may facilitate studies of the mechanisms by which ITCs exert their cancer preventive activity and provide the molecular basis for designing more efficacious ITC compounds.
Insights
Isothiocyanates (ITCs) show cancer chemopreventive effects by targeting specific proteins. This study identified over 30 potential protein targets of ITCs, advancing our understanding of their anti-cancer mechanisms.
Area of Science:
- Biochemistry
- Molecular Biology
- Cancer Research
Background:
- Isothiocyanates (ITCs), including phenethyl isothiocyanate (PEITC) and sulforaphane (SFN), are known for their cancer chemopreventive properties.
- The primary mechanisms are thought to involve cell cycle arrest and apoptosis induction.
- However, the specific upstream molecular targets of ITCs remain largely unidentified.
Purpose of the Study:
- To identify proteins that are covalently modified by ITCs.
- To explore the upstream molecular targets of ITCs in cancer cells.
- To provide a molecular basis for designing more effective ITC compounds.
Main Methods:
- Human non-small cell lung cancer A549 cells were treated with radiolabeled (14)C-PEITC and (14)C-SFN.
- Cell lysates were analyzed using 2-D gel electrophoresis and mass spectrometry.
- Radioactivity patterns were superimposed with protein staining to identify covalently modified proteins.
Main Results:
- A small fraction of cellular proteins were found to be covalently modified by ITCs via thiocarbamation.
- Over 30 potential ITC target proteins with diverse biological functions were identified with high confidence.
- The identified proteins represent novel targets for ITC action.
Conclusions:
- The study successfully identified numerous potential protein targets of ITCs.
- Understanding these targets can elucidate the mechanisms of ITC cancer chemoprevention.
- This discovery provides a foundation for developing improved ITC-based cancer therapies.
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