Jasmonic acid (JA) is a plant hormone that plays an important role in defense against wounding, insects and microbial pathogens. In defense responses against microbial pathogens JA acts... Show moreJasmonic acid (JA) is a plant hormone that plays an important role in defense against wounding, insects and microbial pathogens. In defense responses against microbial pathogens JA acts synergistically with the stress hormone ethylene. ORA59, belonging to the plant-specific class of AP2-domain transcription factors, is the main regulator of JA/ethylene-responsive defense gene expression in the model plant species Arabidopsis thaliana. The aim of the research described in this thesis was to study how the activity of ORA59 is regulated by JA. Studies on the promoter of the defense gene PDF1.2 showed that ORA59 regulates gene expression by interaction with GCCGCC motifs. Studies of ORA59 at the protein level indicated that JA controls ORA59 activity by promoting the nuclear localization and stabilization of the protein. Based on the findings it was postulated that there is an F-box protein that targets ORA59 for degradation, and a repressor protein that sequesters ORA59 in the cytoplasm. A protein interacting with ORA59 was identified by yeast two-hybrid screening. Further studies showed that this protein acts as a repressor of ORA59 by retaining it in the cytoplasm, which leads to fine-tuning of basal resistance against pathogens. Show less
The salicylic acid (SA) signaling pathway triggered by attack of biotrophic pathogens leads to broad spectrum resistance against a plethora of pathogenic fungi, bacteria and viruses and is known as... Show moreThe salicylic acid (SA) signaling pathway triggered by attack of biotrophic pathogens leads to broad spectrum resistance against a plethora of pathogenic fungi, bacteria and viruses and is known as systemic acquired resistance (SAR). One of the hallmarks of SAR is the accumulation of PR proteins and the induced expression of the PR-1 gene is often used as a marker for SAR. In this thesis we describe the characterization of a WRKY-class transcription factor (NtWRKY12) that specifically binds to the sequence TTTTCCAC. This sequence differs significantly from the consensus WRKY protein binding site TTGAC[C/T] (W-box), and was designated as the __WK-box__. The activation of PR-1a has always been proposed to occur mainly via TGA-factors. We show that NtWRKY12 can bind to TGA2.2 and has a more essential role in PR-1a activation. Using bioinformatic analysis various candidate genes have been prospected to have a role in regulation of the SA, jasmonate (JA) and ethylene (ET) biosynthesis or signaling pathways. The connection between AtWRKY28 and ICS1 was further studied. In protoplasts AtWRKY28 can activate ICS1 promoter::GUS reporter constructs and endogenous ICS1 mRNA levels. Using EMSA assays and ChIP experiments the bindingsite has been determined in vitro and confirmed in vivo. Show less