Characterization of complement C3 as a marker of alpha-amanitin toxicity by comparative secretome profiling

Doeun Kim; Min Seo Lee; Hyunchae Sim; Sangkyu Lee; Hye Suk Lee

doi:10.1007/s43188-022-00163-z

Characterization of complement C3 as a marker of alpha-amanitin toxicity by comparative secretome profiling

Toxicol Res. 2022 Dec 22;39(2):251-262. doi: 10.1007/s43188-022-00163-z. eCollection 2023 Apr.

Authors

Doeun Kim¹, Min Seo Lee², Hyunchae Sim¹, Sangkyu Lee¹, Hye Suk Lee²

Affiliations

¹ BK21 FOUR Community-Based Intelligent Novel Drug Discovery Education Unit, College of Pharmacy and Research Institute of Pharmaceutical Sciences, Kyungpook National University, Daegu, 41566 Republic of Korea.
² BK21 Four-sponsored Advanced Program for SmartPharma Leaders, College of Pharmacy, The Catholic University of Korea, Bucheon, 14662 Republic of Korea.

Abstract

In the human body, proteins secreted into peripheral blood vessels are known as the secretome, and they represent the physiological or pathological status of cells. The unique response of cells to toxin exposure can be confirmed via secretome analysis, which can be used to discover toxic mechanisms or exposure markers. Alpha-amanitin (α-AMA) is the most widely studied amatoxin and inhibits transcription and protein synthesis by directly interacting with RNA polymerase II. However, secretory proteins released during hepatic failure caused by α-AMA have not been fully characterized. In this study, we analyzed the secretome of α-AMA-treated Huh-7 cells and mice using a comparative proteomics technique. Overall, 1440 and 208 proteins were quantified in cell media and mouse serum, respectively. Based on the bioinformatics results for the commonly downregulated proteins in cell media and mouse serum, we identified complement component 3 (C3) as a marker for α-AMA-induced hepatotoxicity. Through western blot in cell secretome and C3 ELISA assays in mouse serum, we validated α-AMA-induced downregulation of C3. In conclusion, using comparative proteomics and molecular biology techniques, we found that α-AMA-induced hepatotoxicity reduced C3 levels in the secretome. We expect that this study will aid in identifying new toxic mechanisms, therapeutic targets, and exposure markers of α-AMA-induced hepatotoxicity.

Supplementary information: The online version contains supplementary material available at 10.1007/s43188-022-00163-z.

Keywords: Alpha-amanitin; Complement component 3; Exposure marker; Secretomes; Toxic mushroom.

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