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Metabolic Studies and Bioengineering of Plant Trichomes Towards Enhancing Pest/Disease Resistance and Facilitating Molecular Farming
G. J. Wagner
Department of Plant and Soil Sciences
The work described in this project is directly related to world-wide efforts towards "molecular farming" to exploit plants for sustainable production of non-traditional, commercially-valuable products, i.e., non-food or fiber products. The success in these efforts will depend on acquiring a better fundamental understanding of the metabolism and metabolic regulation underlying plant surface secretion processes, coupled with direct efforts to apply available tools to manipulate these systems towards a useful purpose.
Our work is focused on exploitation of plant trichomes to enhance natural-product-based pest/disease resistance and develop possibilities for molecular farming of trichome secretions that might be used to protect other plants (e.g., food crops, ornamentals) against damage from pests and disease, or as commercial chemical feed-stocks.
A new and recent focus stems from our discovery of antifungal proteins we have called "phylloplanins" that are also produced on aerial surfaces of certain plants. There is a growing need for novel fungicides, particularly natural product based compounds, to augment and replace currently used, chemically-synthesized products whose marketability is under challenge because of environmental concerns.
2011 Project Description
We have continued studies of phylloplanins and have shown for a third year of field trials that T-phylloplanin is effective against brown patch and dollar spot after spraying perennial rye and bentgrass with aqueous solutions containing T-phylloplanin (as T-LWW). No gray leaf spot disease was observed in this year. Thus results of field trials in 2009, 2010, 2011 constitute 3 consecutive years of positive efficacy results for two diseases.
Studies relating to the mechanism of action of phylloplanins were continued to show that T-phylloplanin causes proton leakage from artificial phospholipid vesicles as well as tonoplast vesicles derived from tobacco roots. We developed a second assay to monitor proton leakage using the proton ion trap agent neutral red. This assay was used to show that spores leak protons and other ions when exposed to T-phylloplanins. Results with this assay confirmed earlier results obtained using the acridine orange assay. Results are being readied for publication.
A blue mold sensitive tobacco when transformed with a T-phylloplanin containing gene construct conferred blue mold resistance to Peronospora tabacina. This trait was shown to be retained in seed from fourth generation (T3)plants. Efficacy was best when the fusion gene was targeted to the extracellular space. A specific diterpene product of tobacco trichome exudate was shown to confer resistance to black shank and pithium diseases of tobacco.
Results of this period substantiate results found earlier that describe trichome-derived tobacco phylloplanins as broad spectrum fungicides that may serve as topical anti-fungals against a variety of fungal and oomycete pathogens. They also indicate that tobacco plants engineered to produce elevated levels of T-phylloplanin fusion proteins are protected against blue mold disease.
Regarding the mechanism of action of T- and S-phylloplanins, experiments extended to artificial phospholipid vesicles confirm that these antifungal agents cause pore formation in membranes and spores resulting in ion leakage and inhibition of pathogenicity. Field studies in this period substantiate results obtained in the previous year and thus we have three years of data showing efficacy in the field.
These field trials suggest that phylloplanins can be useful in IPM management of golf courses, athletic fields, residential and business lawns as well as on sod farms to reduce reliance on chemically synthesized fungicides.
Results of our collaborative work on honey bee protection from fungal disease is consistent with the potential usefulness of phylloplanins in this important agricultural industry.
King, B., Williams, D.W., Wagner, G.J. 2011. Phylloplanins reduce the severity of gray leaf spot and brown patch diseases on turfgrasses. Crop Science 51:2829-2839.