Global Analysis of Cellular Responses to Protein Damage
Our laboratory has a long-term interest in the chemical biology of cellular damage induced by lipid oxidation products such as aldehydes and epoxides. These compounds are believed to play an important role in the linkage between oxidative stress or inflammation and multiple chronic diseases (e.g., cardiovascular disease, cancer, osteoarthritis). The major emphasis of our program has been on DNA damage and mutation but it is clear that important cellular responses also result from protein modification. So we initiated a program to characterize the protein targets of modification by lipid electrophiles and relate them to the cellular responses that result. We decided to pursue a systems approach that took advantage of some novel chemistry that we had developed to globally identify the protein targets of lipid electrophiles. We first assembled an inventory of proteins modified by treatment of colon cancer cells with 4-hydroxynonenal (HNE), major product of lipid peroxidation.
In parallel, we determined the identities and magnitudes of upregulated and downregulated genes. Bioinformatic analysis was used to connect the gene expression changes to alterations in the activities of transcription factors. This provided a pattern of signaling networks that were altered by HNE treatment. These signaling pathways were compared to the identity of the HNE-modified proteins to identify protein modifications that could trigger the changes in signal transduction observed. These hypotheses can then be tested by interrupting the signaling pathways in the intact cells by a variety of techniques (e.g., gene knockout, siRNA knockdown). In addition, one can identify the sites of modification of the target proteins to alterations in their functions. This systems approach enables us to relate the chemistry of protein modification to altered signaling in the context of the entire cellular response. It has turned out to be a very powerful approach that is paying significant dividends in terms of helping us understand how cells protect themselves against or ultimately succumb to the reaction of lipid-derived electrophiles with cellular constituents.
A.Vila, K.A. Tallman, A.T. Jacobs, D.C. Liebler, N.A. Porter, and L.J. Marnett, “Identification of Protein Targets of 4-Hydroxynonenal Using Click Chemistry for Ex Vivo Biotinylation of Azido and Alkynyl Derivatives,” Chem.Res.Toxicol., 21, 432-444 (2008)
A. Jacobs and L.J. Marnett, “Heat Shock Factor-1 Attenuates 4-Hydroxynonenal-Mediated Apoptosis: Critical Role for Hsp70 Induction and Stabilization of Bcl-XL,” J.Biol.Chem. 282, 33412-33420 (2007)
A. Druckova, R.L. Mernaugh, A,-J. L. Ham, and L.J. Marnett, “Identification of the Protein Targets of the Reactive Metabolite of Teucrin A in vivo in the Rat,” Chem.Res.Toxicol., 20, 1393-1408 (2007)