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<p>Systemic Acquired Resistance (SAR) is a plant defense response induced by an initial infection in one part of the plant that leads to broad-spectrum resistance to normally virulent pathogens in distant naïve leaves. As part of the Cameron research team, I contributed to demonstrating that the lipid transfer protein, DIR1 is required for SAR long distance signaling in <em>Arabidopsis</em> and travels from induced to distant tissues during SAR. A highly similar<em> Arabidopsis</em> protein DIR1-like was identified and is thought to be responsible for the occasional SAR-competent phenotype observed in the <em>dir1-1</em> mutant. This work provides evidence for the idea that DIR1 and DIR1-like are paralogs created by a recent duplication event and that similar to DIR1, DIR1-like may travel to distant tissues during SAR. To better understand DIR1 and DIR1-like contribution during SAR, <em>dir1-1dir1-like</em> double mutant transgenic plants were created as well as transgenic plants expressing epitope- (HA and FLAG) and fluorescent- (iLOV and phiLOV) tagged DIR1 and DIR1-like to facilitate visualization of movement during SAR. Several putative DIR1 orthologs were identified in crop plants and cucumber CucDIR1 was shown to be functionally equivalent to AtDIR1 in <em>dir1-1</em> complementation studies providing further evidence that DIR1 plays an important role in SAR across plant species. By analyzing conservation between DIR1, DIR1-like and the putative DIR1 orthologs, several protein residues were identified that may be important for DIR1 function during SAR. DIR1 proteins were modified at these sites and the importance of these residues was supported by the reduced binding of the TNS hydrophobic probe in these DIR1 variants. Taken together, this thesis suggests that DIR1 and DIR1-like both participate in SAR in <em>Arabidopsis</em>, that DIR1 crop orthologs are also important for the SAR response and that DIR1 possesses several sites that are critical for its function in long distance SAR signaling.</p> / Doctor of Philosophy (PhD)

Plants respond to pathogens both locally at the site of infection, as well as systemically in distant leaves. Systemic Acquired Resistance (SAR) is an immune response that involves the long-distance transport of SAR signal via the phloem from the site of infection to distant, uninfected leaves to establish long-lasting resistance. The Arabidopsis thaliana Defective in Induced Resistance 1 (DIR1) protein, which is required for SAR, accesses the phloem during SAR for long-distance travel to systemic leaves, and is thought to be part of a SAR signal complex. However, many questions remain about the long-distance movement of DIR1 during SAR – including the cellular route travelled to reach the phloem and whether other proteins are required for DIR1 movement. Fluorescent fusion lines of DIR1 and the related protein DIR1-like were previously created were investigated as potential tools to trace the movement of DIR1/DIR1-like during SAR. Immunoblot analysis of leaf extracts from these DIR1/DIR1-like fluorescent fusion lines revealed no signal, indicating that no fusion protein was present in these lines and therefore, they were likely not useful as a tool for assessing the movement of DIR1/DIR1-like during SAR. Lipid Transfer Protein 2 (LTP2) is required for SAR and interacted with DIR1 in a yeast-two-hybrid assay. To investigate if LTP2 is required for DIR1 movement into the phloem and long-distance, DIR1 signal was investigated by immunoblotting of phloem exudates from SAR induced ltp2-1 mutant plants. The presence of DIR1 signal in phloem exudates of local ltp2-1 leaves but not distant ltp2-1 leaves suggested that LTP2 may be required for the long-distance movement of DIR1 during SAR, but not for DIR1 to enter the phloem in induced leaves. Gene expression changes in the systemic, uninfected leaves are associated with the establishment of SAR, however, it remains less clear if there is a core set of genes important for SAR induction upregulated at the initial site of infection. To investigate this question, SAR was induced through differing treatments that first activated the PAMP-triggered immunity (PTI) pathway or Effector-triggered immunity (ETI) pathway. Common genes upregulated between all three SAR-inducing treatments were identified, revealing genes previously and currently under investigation by the Cameron lab, as well as genes that represent candidates for possible future studies. / Thesis / Master of Science (MSc)