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Title: Nitric oxide and hydrogen peroxide mediated defence responses in Arabidopsis thaliana.
Author: Clarke, Andrew.
Awarding Body: University of the West of England at Bristol
Current Institution: University of the West of England, Bristol
Date of Award: 2001
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Incompatible plant/pathogen interactions are often manifested as the hypersensitive response (HR), characterised by host cell death and rapid tissue collapse at the site of attempted infection. A key early response during the HR is the generation of reactive oxygen species (ROS), such as the superoxide anion ( 0; -) and hydrogen peroxide (H20 2), in an oxidative burst. The ROS produced during the oxidative burst have been implicated as cellular signalling molecules for the induction of defences responses including hypersensitive cell death. Increasing evidence exist that the free radical, nitric oxide (NO) also acts as a signalling molecule in plants during plant/pathogen interactions. The generation of NO in response to bacterial challenge, and the potential signalling pathways involved in H20 2- and NO-induced defence responses in Arabidopsis were therefore investigated Arabidopsis suspension cultures were found to generate elevated levels of NO and undergo cell death analogous to HR seen in planta, in response to challenge by avirulent bacteria. Using NO donors, elevated levels of NO were found to be sufficient to induce cell death independently of ROS, but not the expression of the defence-related genes PAL or GST. The NO-induced cell death was sensitive to inhibitors of RNA processing and protein synthesis, suggesting that NO-induced cell death is a form of programmed cell death (PCD), requiring the expression of at least one gene. However, the source of NO production by Arabidopsis remains to be elucidated, but appears to be independent of nitric oxide synthase-like activity. Pharmacological studies using specific inhibitors of mammalian mitogen activated protein kinase (MAPK) signalling cascades, and guanylate cyclase, the enzyme responsible for the production of second messenger cyclic guanosine monophosphate (cGMP), suggest that a MAPK signalling cascade acts downstream or independently of the oxidative burst to initiate H20 2-induced defence responses, while NO-induced cell death requires the production of cGMP in Arabidopsis. A number of studies have attempted to establish whether PCD induced during the HR in plants is similar to apoptotic cell death of anin1al cells. The key executioners of apoptosis in animal cells are caspases. NO was found to induce caspase-like activity in Arabidopsis cells, while a specific inhibitor of caspase-l blocked harpin-, H20 2- and NO-induced cell death. A characteristic of apoptosis is chromatin condensation and DNA fragmentation into nucleosomal fragments. Chromatin condensation was observed in Arabidopsis cells treated with the NO donor Roussin's black salt, but no DNA fragmentation was found in DNA extracted from cells treated with harpin, H20 2 or NO. In addition, random DNA degradation indicative of necrosis was found in DNA extracted from cells following avirulent bacterial challenge.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: Plant pathogen interactions; Cell death