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Updated: Dec 28, 2025
Toxin Induction and Protein Extraction from Fusarium spp. Cultures for Proteomic Studies
Published on: February 16, 2010
The Fusarium mycotoxin, 2-Amino-14,16-dimethyloctadecan-3-ol (AOD) induces vacuolization in HepG2 cells
A Solhaug1, M L Torgersen2, J A Holme3
1Chemistry and Toxinology Research Group, Norwegian Veterinary Institute, 0454, Oslo, Norway.
Abstract:
The mycotoxin 2-Amino-14,16-dimethyloctadecan-3-ol (AOD) has been isolated from cultures of the fungus Fusarium avenaceum, one of the most prevalent Fusarium species. AOD is an analogue of sphinganine and 1-deoxysphinganine, important intermediates in the de novo biosynthesis of cellular sphingolipids. Here we studied cellular effects of AOD using the human liver cell line HepG2 as a model system. AOD (10 μM) induced a transient accumulation of vacuoles in the cells. The effect was observed at non-cytotoxic concentrations and was not linked to cell death processes. Proteomic analyses indicated that protein degradation and/or vesicular transport may be a target for AOD. Further studies revealed that AOD had only minor effects on the initiation rate of macropinocytosis and autophagy. However, the AOD-induced vacuoles were lysosomal-associated membrane protein-1 (LAMP-1) positive, suggesting that they most likely originate from lysosomes or late endosomes. Accordingly, both endosomal and autophagic protein degradation were inhibited. Further studies revealed that treatment with concanamycin A or chloroquine completely blocked the AOD-induced vacuolization, suggesting that the vacuolization is dependent of acidic lysosomes. Overall, the results strongly suggest that the increased vacuolization is due to an accumulation of AOD in lysosomes or late endosomes thereby disturbing the later stages of the endolysosomal process.
Insights
The mycotoxin 2-Amino-14,16-dimethyloctadecan-3-ol (AOD) causes vacuole accumulation in liver cells by disrupting lysosomal function. This fungal toxin interferes with cellular sphingolipid pathways at non-cytotoxic levels.
Area of Science:
- Biochemistry
- Cell Biology
- Mycotoxicology
Background:
- The mycotoxin 2-Amino-14,16-dimethyloctadecan-3-ol (AOD) is produced by Fusarium avenaceum.
- AOD is structurally similar to sphingolipid intermediates, suggesting a role in cellular lipid metabolism.
- Fusarium species are prevalent and can contaminate food and feed.
Purpose of the Study:
- To investigate the cellular effects of the mycotoxin AOD in a human liver cell model (HepG2).
- To elucidate the mechanism behind AOD-induced cellular changes, particularly vacuolization.
- To determine if AOD impacts sphingolipid biosynthesis or related cellular processes.
Main Methods:
- Human liver cell line (HepG2) exposure to AOD (10 μM).
- Proteomic analysis to identify cellular targets of AOD.
- Assessment of macropinocytosis and autophagy initiation rates.
- Immunofluorescence staining for lysosomal markers (LAMP-1).
- Treatment with lysosomal inhibitors (concanamycin A, chloroquine).
Main Results:
- AOD induced transient, non-cytotoxic vacuole accumulation in HepG2 cells.
- Proteomics suggested AOD affects protein degradation and vesicular transport.
- Vacuoles were LAMP-1 positive, indicating lysosomal or late endosomal origin.
- AOD inhibited endosomal and autophagic protein degradation.
- AOD-induced vacuolization was dependent on acidic lysosomes and blocked by inhibitors.
Conclusions:
- AOD accumulates in lysosomes or late endosomes, disrupting the endolysosomal pathway.
- The mycotoxin interferes with cellular sphingolipid metabolism and lysosomal function.
- AOD's effects on vacuolization are linked to impaired protein degradation within lysosomes.
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