Interaction with and Effects on the Profile of Proteins of Botrytis cinerea by C6 Aldehydes

The natural volatile compounds cis-3-hexenal (c-3-H) and trans-2-hexenal (t-2-H) have significant antifungal activity with potential for use as postharvest fumigants of fruits and vegetables. However, the nature of their interaction with fungi and impact on fungal growth at the molecular level are largely unknown. The sites of interaction of these six carbon (C6) aldehydes with Botrytis cinerea, a common pathogen of many plant species, was characterized using 3H-labeled c-3-H and t-2-H. Radiolabeled C6 aldehydes were produced with lipoxygenase and hydroperoxide lyase extracts using [9,10,12,13,15,16-3H6]linolenic acid as a substrate. Following exposure of B. cinerea cultures to radiolabeled C6 aldehydes, radiolabel was recovered in protein-enriched but not lipid-enriched fractions. Radiolabel was incorporated at higher levels (6-fold per milligram of fresh weight and 4-fold per microgram of protein) into conidia than mycelia. About 95% of the radiolabeled aldehyde recovered in the protein fraction was from the surface of the fungal tissue, while 5% was from protein in internal tissue (cell wall, membrane, and cytosol). Exposure to t-2-H at both 5.4 and 85.6 μmol affected the protein profile of B. cinerea, changing the intensity of over one-third of all proteins. Both up-regulation and down-regulation of specific proteins were observed by two-dimensional gel electrophoresis, indicating a clear effect of t-2-H on changes in the protein profile of B. cinerea. This is the first evidence that fungal proteins are targets of the volatile C6 aldehydes and that sublethal levels of the aldehydes cause changes in the protein profile of a fungus. Keywords: Botrytis cinerea; radiolabeled C6 aldehyde; cis-3-hexenal; trans-2-hexenal; site of interaction; protein profile