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Title: Molecular and genetic analysis of signal transduction pathways underlying PR-1 expression in Arabidopsis
Author: Santamaria, Marjorie
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2002
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Antimicrobial proteins are a key feature underlying the deployment of both pre-formed and inducible defence responses. Probably the most well characterised class are the pathogenesis related (PR)-proteins, which are found in both basic and acidic isoforms. This study describes the isolation and characterisation of a gene, designated AtPRBl, encoding a basic PRl-like protein from Arabidopsis. This protein showed high amino acid sequence identity with basic and acidic PR1 proteins from other plant species for example, PRB1 from Nicotiana tabacum and PR1 from Brassica napus, at 64% and 78% identity respectively. A genomic DNA fragment containing 2345 bp upstream from the putative transcriptional start site was fused to the gene encoding the luciferase (LUC) gene, in order to test for promoter activity. The resulting construct was transformed into Arabidopsis accession Col-0 and analysis of LUC activity, using an ultra low light imaging camera system, revealed that the AtPRBl promoter established an exquisite organ-specific expression pattern. LUC activity was observed in flowers, stems and roots but not in leaf tissue. Superimposed upon this organ-specific expression pattern was responsiveness, in root tissue, to ethylene (ET) and methyl-jasmonate (Me-JA), important cues during the establishment of plant disease resistance. In contrast, AtPRBl::LUC gene expression was repressed in response to salicylic acid (SA) treatment. Analysis of a limited series of AtPRBl 5'-promoter deletion mutants, identified a number of promoter regions important for both the establishment of organspecific expression and responsiveness to ET and Me-JA. While AtPRBl gene expression was not induced in response to an avirulent isolate of Peronospora parasitica in leaf tissue, this gene may contribute to constitutive resistance in other tissues and/or to Me-JA and ET dependent defence responses engaged against necrotrophic pathogens in root tissue. In the second part of this study, a population of 5000 activation-tagged lines was generated in a PR-1\.LUC genetic background to uncover novel systemic acquired resistance (SAR) mutants. The mutant screen involved imaging for constitutive LUC expression or absence of LUC expression after induction with the SA functional analogue, BTH. A set of five mutants displaying constitutive LUC activity were selected for further characterisation. Four mutants, designated esr2, esr3, esr5 and esr6 for enhanced systemic resistance, exhibited constitutive PR-1 gene expression and increased resistance to the virulent oomycete pathogen P.parasitica Noco2. Analysis of crosses between the lesion mimic mutant esr2 and other mutants that disrupt SA and JA/ET signalling showed that PR-1 expression in esr2 was largely but not strictly dependent on SA signalling. Furthermore, the ET- and JA-insensitive etrl and coil mutants were able to block PR-1 expression when introduced into esr2. Based on these results, ESR2 could act as a regulator of a signalling pathway that requires components of the SA, and JA/ET pathways. Further characterisation and cloning of esr mutants, should provide valuable insights into the function, structure and molecular interactions of novel genes involved in disease resistance.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available