All living organisms are composed of proteins plus other substances made by proteins including fat, sugar, and DNA. Proteins do the work of life; they provide shape, transmit molecular "news", make all of the parts of a cell, capture and utilize energy, provide weapons and defenses, and copy genes required for transmittal to the next generation. Despite their importance, proteins are rarely the first things measured by scientists because existing technologies reveal only a small fraction of the total. This proposal has two principal aims. The first is to increase the sensitivity and precision of protein identification by mass spectrometry. The goal is to identify 90% of the proteins in a sample, or about 10,000 proteins. Current state-of-the-art technology identifies only about 10% of the proteins. Quantitative aspects of the method will be developed to detect changes of two-fold or greater in the level of each protein, compared with existing methods that are qualitative (proteins are designated as either present or absent). Such improvements could be used by all scientists and should have broad application. The second aim is to use this new technology to answer an important scientific question with agricultural significance: how does the root knot nematode infect plants? Nematodes cause many of the most serious diseases of crops. They burrow into the roots and activate abnormal growth patterns which eventually disrupt root functions, leading to water and nutrient stress. Plant proteins will be identified whose levels are affected by infection. These changes will be compared between different plant species infected by the same nematode to learn which changes are in common. These common or conserved changes may be the most important because they have persisted over millions of years. Genetic alterations will be made in the plant that either mimic or prevent the changes caused by infection, as a way to test whether the changes caused by nematodes are necessary for the infection to take place. This will aid in the basic understanding of how to make crops that resist disease. This project will provide training opportunities in state-of-the-art proteomics for postdoctoral researchers and graduate students.
