Arsenic-Perturbed Protein Function Mediated by Mitochondrial Targeting

The goal of the pilot project is to determine whether arsenic induces changes in the mitochondrial acetylome that are consistent with its documented ability to disrupt energy metabolism.  Progress in this project has focused on working closely with Dr. Tsaprailis in the proteomics core to develop expertise in separation and purification of protein targets for proteomic analysis of post-translational modifications.   In initial experiments we used acetylated bovine serum albumin to successfully develop in-gel digestion techniques and to verify that proteomic analyses at the core facility could resolve post-translational modifications with the subtle mass shift of an acetate group.  The experimental model being used in subsequent studies is the arsenic-induced accumulation of hypoxia-inducible factor 1-alpha (HIF1A) in BEAS-2B cells.  We have established that HIF-1A accumulation in BEAS-2B cells is not a result of transcriptional induction and is associated with protein stabilization, likely through post-translational modifications that reduce its affinity for the E3-ligase-like VHL protein.  We have successfully immunoprecipitated HIF1A in arsenic-exposed cells and are now awaiting the results of the proteomic analyses.  With successful analysis of HIF1A in this model, we will begin “omics” searches for acetylated and succinylated mitochondrial proteins induced by arsenic exposure in BEAS-2B.