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United States Department of Agriculture

Agricultural Research Service

Research Project: MANAGEMENT OF NEMATODES AND VIRUS DISEASES AFFECTING POTATO AND GRAIN CROPS Title: The proteomics toolbox for agricultural research: precision biomarker discovery using genetics

Authors
item Cilia, Michelle
item Tamborindeguy, Cecilia -
item Thannhauser, Theodore
item Gray, Stewart

Submitted to: Journal of Agricultural and Urban Entomology
Publication Type: Proceedings
Publication Acceptance Date: March 28, 2010
Publication Date: November 7, 2010
Citation: Cilia, M., Tamborindeguy, C., Thannhauser, T.W., Gray, S.M. 2010. The proteomics toolbox for agricultural research: precision biomarker discovery using genetics. Journal of Agricultural and Urban Entomology. 27:20.

Technical Abstract: The vast array of proteomics technologies can often leave investigators wondering which set of tools to use to address their biological question. Deciding which set of tools to apply requires knowledge of the biology of the system and of the subtleties of each proteomics approach. Proteomics approaches can be broadly categorized as top-down, bottom up, or a combination of both. One application of top down proteomics separates a complex mixture of intact proteins using 2D gels to measure the molecular weight and isoelectric point of the proteins and their various isoforms in the sample. Proteins are enzymatically digested from the gel spots and identified using mass spectrometry. Bottom up methods accomplish protein identification differently. The entire complex protein sample is first digested with a specific protease (e. g. trypsin). The presence of particular proteins is inferred from the identification of a subset of its constituent peptides after multidimensional chromatography and mass spectrometry (LC-MS/MS). Our lab studies the aphid transmission of yellow dwarf viruses. By crossing a competent genotype and refractive genotype, we generated an F2 population of aphids that differ in virus transmission competency. We applied a combination of approaches to investigate the proteomes of the aphid genotypes that differ in their transmission capacity. Combining aphid genetics with the top-down quantitative intact proteomics (QIP) technique 2D fluorescence difference gel electrophoresis (DIGE) and LC-MS/MS, we identified more than 35 aphid proteins whose expression level correlates with differences in the intercellular transport of yellow dwarf viruses across aphid gut and salivary tissues. The combination of genetics with the initial top-down approach facilitated two exciting discoveries in our system that would not have been identified had we used any other approach. The first unexpected discovery was the implication of bacterial endosymbionts of aphids as playing an important role in determining virus transmission competency. Secondly, we discovered that several of the aphid and bacterial proteins have practical importance in that they serve as robust biomarkers for distinguishing transmission competent and transmission refractive populations of aphids. Not only are these the first protein biomarkers for virus transmission in any insect, they are the first protein biomarkers for any phenotypic trait discovered using a top-down proteomics approach. This work opens the door for novel virus and integrated pest management strategies.

Last Modified: 10/20/2014
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