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Title: Redox regulation of plant S-nitrosylation
Author: Chang, Tao-Ho
ISNI:       0000 0004 7225 0316
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2017
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Nitric oxide (NO), a diffusible gas molecule, is a major signal molecule in both plants and animals and regulates a plethora of biological processes. S-nitrosylation, a post-translation modification, is conducted by NO, which covalently attaches protein cysteine thiols and forms an S-nitroso thiol. S-nitrosylation plays an important role in plant development and plant immune systems. In Arabidopsis thaliana, S-nitrosoglutathione (GSNO) is the major NO donor for S-nitrosylation, and GSNO reductase (GSNOR) indirectly controls the S-nitrosylation level by turning over the GSNO. An A. thaliana T-DNA insertion mutant gsnor1-3 shows the loss of GSNOR activity and increases the S-nitrosylation level, resulting in loss of apical dominance, reduction of SA accumulation, increased hypersensitive response (HR) cell death and reduced disease resistance against virulence, avirulence and non-host pathogens. Interestingly, loss of GSNOR in Drosophila melanogaster, an animal model system, reduces the resistance against gram-positive and fungal pathogens. Catalase is an antioxidant enzyme and regulates the redox environment through scavenging the hydrogen peroxide (H2O2) to oxygen and water. Previous work in our lab had discovered two gsnor1-3 suppressor mutants, gsnor1-3 spl7 and gsnor1-3 spl8, which restore the loss of apical dominance and partially restore disease resistance. These two suppressor mutants were then identified as the point mutation in CAT3. CAT3, one of the three CAT genes in Arabidopsis, expresses catalase specifically in vascular tissues. To further extend the suppression of cat3 in gsnor1-3, the mutations in CAT3 and its paralogs CAT2 and CAT1, as well as other redox-related genes in gsnor1-3 background, were generated. In the developmental phenotype, only the gsnor1-3 cat3 showed significant changes compared with gsnor1-3. The disease susceptibility and HR cell death in gsnor1-3 cat3 were less than gsnor1- 3 and similar to wild-type. Moreover, the redox-related genes and CAT3 paralog mutations in gsnor1-3 background showed no significant changes in disease resistance against virulence pathogen compared with gsnor1-3 plant. Meanwhile, an SA-dependent (salicylic acid) defence-related gene (PR1, pathogenesis-related gene 1) showed the early expression in gsnor1-3 cat3 plant compared with gsnor1-3 plant. Results of developmental and disease-related phenotypes suggest the suppression effects which turn-over the malfunction in gsnor1- 3 are highly specific to CAT3. The previous report demonstrates that the hydroxyl radical, a reactive oxygen species by-product from H2O2, decomposes GSNO to oxidised glutathione in vitro. The interaction of GSNO and hydroxyl radical may be the possible mechanism of how cat3 suppresses gsnor1- 3. Therefore, we speculated less amount of GSNO in gsnor1-3 cat3 plant than in gsnor1-3 plant and lower level of hydroxyl radicals in gsnor1-3 cat3 plant than in cat3 plant. To evaluate our hypothesis, the content hydroxyl and GSNO were analysed in wild-type, gsnor1-3, cat3 and gsnor1-3 cat3 plants. The total S-nitrosylated protein, which indicates the GSNO content in vivo, was less in gsnor1-3 cat3 than in gsnor1-3. Furthermore, the level of hydroxyl radical in gsnor1-3 cat3 was lower than cat3. Accordingly, the reduction of hydroxyl radical in gsnor1- 3 cat3 may occur due to the reaction with GSNO and vice versa. Similar to what has been found in Arabidopsis, D. melanogaster also reported partial restoration of the immunodeficiency phenotypes of gsnor knock-out flies with an additional mutation in CAT gene. Interestingly, the content of hydroxyl radical in gsnor-/- cat-/- line was less than cat+/-. Collectively, our results suggest an interaction of hydroxyl radical and GSNO may happen both in Arabidopsis and Drosophila. Further research is needed to clarify the interaction between hydroxyl radical and GSNO in Arabidopsis as well as in Drosophila.
Supervisor: Loake, Gary ; Campbell, Colin Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: S-nitrosylation ; GSNOR1 ; cat3