|Artlip, Timothy - Tim|
|Norelli, John (jay) - Jay|
Submitted to: Acta Horticulturae
Publication Type: Peer reviewed journal
Publication Acceptance Date: 9/10/2004
Publication Date: 6/1/2005
Citation: Wisniewski, M.E., Bassett, C.L., Artlip, T.S., Janisiewicz, W.J., Norelli, J.L., Droby, S. 2005. Overexpression of a peach defensin gene can enhance the activity of post harvest biocontrol agents. Acta Horticulturae 682: 1999-2006. Interpretive Summary: Developing reliable and effective alternatives to chemical fungicides to control postharvest losses of fruit due to decay is a critical objective of the USDA-ARS National Program 303 addressing Plant Disease. Previous research by USDA-ARS scientists at Kearneysville, WV has led to the development of new commercial products based on the use of antagonistic yeasts as biocontrol agents. Current research is aimed at increasing the reliability and efficacy of the biocontrol agents under a wide array of conditions. One approach to improving their efficacy is to examine the potential use of antimicrobial peptides that naturally occur in fruit trees. In this regard, we isolated a gene from peach that codes for a potent antifungal protein, belonging to the defensin family of proteins, and engineered a yeast to constitutively express this protein. We demonstrated that the transgenic yeast had an improved ability to inhibit the spore germination of two major postharvest decay organisms of apple (Penicillium and Botrytis). The results from this research identified a native antimicrobial protein present in peach trees and showed that this protein can be used to enhance biocontrol activity. Our research also demonstrated that the defensin gene was also present in apple trees. Future research will characterize the antimicrobial activity of these proteins in more detail and identify other naturally-occuring proteins in fruit trees with antimicrobial activity.
Technical Abstract: A portion of our research involves developing methods to enhance the activity of yeast biocontrol agents. In the present study, we describe the cloning and characterization of defensin genes from bark tissues of peach and apple. The open reading frame of both genes code for proteins related to the defensins, a family of highly antimicrobial proteins. Sequence comparison of the encoded proteins revealed significant homology between the peach and apple defensins and defensins from other plant species. RNA gel blot analysis and semi-quantitative-PCR indicated that the genes are seasonally-expressed in bark tissues of one-year-old shoots, and the peach defensin is also expressed in early fruit development. A recombinant version of the peach defensin gene, rDFN1 was expressed in the yeast, Pichia pastoris. It was found that rDFN1 inhibited germination of the fungal pathogens Penicillium expansum and Botrytis cinerea. The potential of using defensin genes to enhance biocontrol yeasts is discussed.