Investigation of the effects of ochratoxin-a on global protein sumoylation

Abstract

Ochratoxin A (OTA), a secondary metabolite produced by especially Aspergillus and Penicillium species, is found as a contaminant in foods such as daily consumed corn flakes, coffee, meat and dairy products. OTA is a potential carcinogenic toxin whose mode of action is not fully understood, yet. It is known that OTA deregulates MAPK/Erk1-2 and PI3K/Akt signaling pathways responsible for the vital cellular activities. Therefore, identification of the mechanisms underlying OTA's carcinogenic and toxic effects is of great importance. SUMOylation is one of the post-translational modifications playing crucial roles in cellular homeostasis and the regulation of the cell's vital functions. There is no study examining the effects of OTA on the global protein SUMOylation changes and contributions of these changes on OTA-induced activations of MAPK/Erk1-2 and PI3K/Akt signaling pathways. In this Master's project, first we showed that OTA induces oxidative stress in immortalized human proximal tubule epithelial HK-2 cell line. Next, we demonstrated that OTA treatment leads to alterations in global protein SUMOylation and activation of PI3K/Akt and MAPK/Erk1-2 pathways, which might be linked to OTA-induced oxidative stress. Moreover, we showed that OTA-induced MAPK/Erk1-2 pathway activation is not affected when global SUMOylation is inhibited using ML-792. In contrast, our data portrayed that global SUMOylation is essential for phosphorylation of Akt under OTA exposure. Taken all together, our data suggests that SUMOylation may play a key role in cellular survival under OTA exposure. Finally, we constructed stable SUMO-1- and SUMO-2- overexpressing HK-2 cell lines to eliminate low efficiency for immunoprecipitation. By using these cell lines we will be able to detect possible SUMO targets under OTA exposure via Mass Spectrometry Analysis.