Hyaloperonospora arabidopsidis (Hpa) is a naturally occurring oomycete pathogen on Arabidopsis thaliana. It is related to downy mildews of economically important crops such as cabbage, kale and broccoli, belonging to the Brassicaceae family. Downy mildew pathogens are obligate biotrophs that extract nutrients exclusively from living plant cells. As a part of its obligate biotrophy lifestyle, Hpa has lost the ability to assimilate inorganic nitrogen and sulfur. It thus has to acquire these nutrients from the host in an organic form; possibly amino acids. Using a reverse genetic approach, I was able to identify two host amino acid transporters that are up-regulated during Hpa infection: AAP3 and AAP6. Both of these transporters are localized in the vasculature of the plant, AAP3 mostly in the root, and AAP6 in the roots and shoots. Using transgenic lines of Arabidopsis containing transcriptional and translational reporter fusion constructs for these genes, I found that AAP3 displays increased mRNA accumulation which is attributable to an increased promoter activity in regions of shoot tissue colonized by Hpa. On the other hand, AAP6 displays a mild increase in mRNA accumulation under Hpa infection, but the induction becomes more prominent at the protein level as seen by fluorescence from GFP fused to AAP6. Surprisingly, null mutants of AAP3 did not impact Hpa growth whereas null mutants of AAP6 made the plant more susceptible to Hpa. Furthermore, aap6 mutants accumulate fewer free amino acids in the phloem compared to wild-type plants when infected with Hpa. Together, these results suggest that AAP6 acts a nutritional starvation gene for the pathogen and hence aids the plant during infection. While we now know more about AAP3's regulation during infection, its function remains to be elucidated. To successfully colonize a plant, a pathogen must be able to achieve both suppression of plant immunity and acquisition of nutrients from the plant host. While the former has been well studied, research on the latter is sparse. This work was a step in the direction to increase our understanding of potential players in nutrient acquisition by pathogens. / Doctor of Philosophy / A key aspect of achieving and maintaining food security is sustainable agricultural production. This is endangered by plant diseases that lead to large losses in crop production. All plant pathogens have to acquire food and nutrients from the plants they infect. Understanding how they acquire nutrients from the plant at a molecular level can give us insight into potential methods to prevent this and hence reduce the impact of plant diseases. One such nutrient is nitrogen. Nitrogen is essential to all of an organism’s cellular and metabolic processes. Organisms utilize nitrogen by converting it from inorganic forms such as nitrates to organic forms such as amino acids. Some plant pathogens, such as Hyaloperonospora arabidopsidis (Hpa), which causes downy mildew disease on the model plant Arabidopsis, complete their entire life cycle on a living plant. They are also unable to convert the inorganic nitrogen to organic forms and hence depend on acquiring organic forms of nitrogen from the plant. Thus, it is important to understand how they acquire amino acids from the plant. Plants use amino acid transporters that serve as a siphon or a pump in moving amino acids from one region of the plant to another. It is possible that pathogens manipulate plant’s amino acid transporters to move amino acids towards the infection site while, at the same time, plants might use another set of transporters to move amino acids away from the pathogen. This work was an attempt at understanding this potential role of plant amino acid transporters in plant-pathogen interactions using the model system of Hpa and Arabidopsis.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/102868 |
| Date | 04 October 2019 |
| Creators | Sonawala, Unnati Subhash |
| Contributors | Plant Pathology, Physiology and Weed Science, Pilot, Guillaume, Tholl, Dorothea, McDowell, John M., Westwood, James H. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Detected Language | English |
| Type | Dissertation |
| Format | ETD, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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