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Tropospheric ozone (O3) is a major air pollutant that decreases yield of important crops worldwide. Despite long-lasting research of its negative effects on plants, there are many gaps in our knowledge on how plants respond to O3. In this study, we used natural variation in the model plant Arabidopsis (Arabidopsis thaliana) to characterize molecular and physiological mechanisms underlying O3 sensitivity. A key parameter in models for O3 damage is stomatal uptake. Here we show that the extent of O3 damage in the sensitive Arabidopsis accession Shahdara (Sha) does not correspond with O3 uptake, pointing toward stomata-independent mechanisms for the development of O3 damage. We compared tolerant (Col-0) versus sensitive accessions (Sha, Cvi-0) in assays related to photosynthesis, cell death, antioxidants, and transcriptional regulation. Acute O3 exposure increased cell death, development of lesions in the leaves, and decreased photosynthesis in sensitive accessions. In both Sha and Cvi-0, O3-induced lesions were associated with decreased maximal chlorophyll fluorescence and low quantum yield of electron transfer from Photosystem II to plastoquinone. However, O3-induced repression of photosynthesis in these two O3-sensitive accessions developed in different ways. We demonstrate that O3 sensitivity in Arabidopsis is influenced by genetic diversity given that Sha and Cvi-0 developed accession-specific transcriptional responses to O3. Our findings advance the understanding of plant responses to O3 and set a framework for future studies to characterize molecular and physiological mechanisms allowing plants to respond to high O3 levels in the atmosphere as a result of high air pollution and climate change.

Cell walls surround all plant cells, and their composition and structure are modified in a tightly controlled, adaptive manner to meet sometimes opposing functional requirements during growth and development. The plant cell wall integrity (CWI) maintenance mechanism controls these functional modifications, as well as responses to cell wall damage (CWD). We investigated how the CWI system mediates responses to CWD in Arabidopsis thaliana. CWD induced by cell wall-degrading enzymes or an inhibitor of cellulose biosynthesis elicited similar, turgor-sensitive stress responses. Phenotypic clustering with 27 genotypes identified a core group of receptor-like kinases (RLKs) and ion channels required for the activation of CWD responses. A genetic analysis showed that the RLK FEI2 and the plasma membrane-localized mechanosensitive Ca2+ channel MCA1 functioned downstream of the RLK THE1 in CWD perception. In contrast, pattern-triggered immunity (PTI) signaling components, including the receptors for plant elicitor peptides (AtPeps) PEPR1 and PEPR2, repressed responses to CWD. CWD induced the expression of PROPEP1 and PROPEP3, which encode the precursors of AtPep1 and AtPep3, and the release of PROPEP3 into the growth medium. Application of AtPep1 and AtPep3 repressed CWD-induced phytohormone accumulation in a concentration-dependent manner. These results suggest that AtPep-mediated signaling suppresses CWD-induced defense responses controlled by the CWI mechanism. This suppression was alleviated when PTI signaling downstream of PEPR1 and PEPR2 was impaired. Defense responses controlled by the CWI maintenance mechanism might thus compensate to some extent for the loss of PTI signaling elements.

The air pollutant ozone (O3) enters plant leaves through stomata and activates apolastic reactive oxygen species (ROS) signaling. Depending on growth conditions and genotype, this results in large transcriptional reprogramming,closure of stomatal pores and activation of cell death programs. These responses are also regulated through plant stress hormones. This thesis sheds light on how stress hormone signaling is connected with apoplastic ROS signaling in the model plant Arabidopsis thaliana, and investigates regulatory mechanisms which generate specificity among sequence-specific transcription factors (TFs), the executers of apoplastic ROS -induced transcriptional reprogramming. The essential methods of the thesis include O3 exposures of Arabidopsis wild type and mutant plants followed by quantification of cell death and characterization of transcriptional responses supplemented with several protein-level analyses of selected WRKY family TFs. The O3-induced cell death was found to be inhibited by plant hormone salicylic acid, and genes RESPIRATORY BURST OXIDASE HOMOLOG F (RBOHF) and WRKY70 were found to be required for O3-induced cell death in jasmonic acid insensitive genetic background. Even though stress hormones were verified to play important roles in the regulation of cell death, the transcriptional response to apoplastic ROS in a hormone deficient/insensitive mutant was highly similar to wild type, suggesting that much of the signaling involved is independent of the studied hormones jasmonic acid, salicylic acid, and ethylene. The potential major executers of transcriptional response to apoplastic ROS, WRKY family TFs, were studied for their transcriptional regulation, DNA-binding preferences, protein-protein interactions, subcellular localization, and effects on transcriptome. The results showed that the DNA-binding preferences of WRKYs vary substantially between phylogenetic groups, implying that the specificity in signaling between different WRKYs can be, Ilmansaaste otsoni tunkeutuu kasvin lehteen ilmaraoista ja käynnistää siellä reaktiivisiin happilajeihin perustuvan viestintäketjun. Kasvuoloista ja perimästä riippuen tämä viestintä voi johtaa ilmarakojen sulkeutumiseen, ohjelmoidun solukuoleman käynnistymiseen ja suuriin muutoksiin geenien ilmentymisessä. Reaktiivisten happilajien lisäksi näitä vasteita säädellään kasvihormonien välityksellä. Tämä väitöstyö valaisee kasvin stressihormoniviestinnän ja reaktiivisiin happilajeihin perustuvan viestinnän yhteyksiä lituruohossa (Arabidopsis thaliana) ja tutkii, mitkä mekanismit luovat spesifisyyttä niiden geenien ilmentymisen säätelytekijöiden välille, jotka suurelta osin vastaavat reaktiivisten happilajien aiheuttamista muutoksista geenien ilmentymisessä. Väitöstyön keskeisiä menetelmiä ovat lituruohon sekä lituruohomutanttien otsonikäsittelyt ja niiden aiheuttaman ohjelmoidun solukuoleman mittaaminen. Lisäksi tutkittiin otsonikäsittelyn aiheuttamia muutoksia geenien ilmentymisessä sekä proteiinitason muutoksia WRKY-perheen säätelytekijöissä. Kasvihormoni salisyylihapon havaittiin estävän otsonin käynnistämää solukuolemaa lituruohossa. Lisäksi havaittiin, että geenit RESPIRATORY BURST OXIDASE HOMOLOG F (RBOHF) ja WRKY70 vaaditaan otsonin käynnistämän ohjelmoidun solukuoleman toteuttamiseksi otsoniherkässä liturohomutantissa, joka on kykenemätön aistimaan kasvihormoni jasmiinihappoa. Vaikka stressihormonien keskeinen rooli solukuoleman säätelyssä vahvistettiinkin, muutokset geenien ilmentymisessä vasteena reaktiivisiin happilajeihin olivat yllättävän samanlaisia lituruohossa ja lituruohomutantissa, jolta puuttui kyky valmistaa/aistia keskeisiä stressihormoneja jasmiinihappoa, salisyylihappoa ja etyleeniä. Tulos antaa ymmärtää, että suuri osa geenien ilmentymisen muutoksiin johtavasta viestinnästä vasteena reaktiivisiin happilajeihin kulkee reittejä, jotka eivät vaadi kyseisiä hormoneja toimiakseen. WRKY-perheen säätelytekijät ovat keskeisessä osassa apoplastin reaktiivi

OBJECTIVE: To design a multifactorial biological modulation approach targeting ischemia reperfusion injury to augment viability of porcine liver grafts from non-heart-beating donors (NHBD). BACKGROUND DATA: Liver Transplantation (LTx) from NHBD is associated with an increased risk of primary nonfunction (PNF) and biliary complications. In porcine NHBD-LTx, we previously reported a 50% risk of PNF and toxic bile formation in grafts exposed to > or =30' warm ischemia (WI). METHODS: Porcine livers exposed to 45' WI were cold stored, transplanted and either modulated (n = 6) or not (controls, n = 9). In the modulation group, donor livers were flushed with warm Ringers (avoiding cold-induced vasoconstriction), streptokinase (eliminating stagnating thrombi), and epoprostenol (vasodilator, platelet aggregation inhibitor) prior to cold storage. In recipients, glycine (Kupffer cell stabilizer), alpha1-acid-glycoprotein (anti-inflammatory protein), MAPKinase-inhibitor (pro-inflammatory cytokine generation inhibitor), alpha-tocopherol and glutathione (anti-oxidants), and apotransferrin (iron chelator) were administrated intravenously. PNF, survival, lactate, transaminase, TNF-alpha, redox-active iron, and biliary bile salt-to-phospholipid ratio were monitored. RESULTS: No PNF was observed in modulated versus 55% in control pigs (P = 0.025). Survival was 83% in modulated versus 22% in control pigs (P = 0.02). At 180' postreperfusion, lactate was lower in modulated (5.4 +/- 1.9 mmol/L) versus control pigs (9.4 +/- 2.2 mmol/L; P = 0.011). At 60' postreperfusion, there was a trend for lower AST in modulated versus control pigs at 60' (939 +/- 578 vs. 1683 +/- 873 IU/L; P = 0.089). Postreperfusion, TNF-alpha remained stable in modulated pigs (49 +/- 27 pg/mL at 15' and 85 +/- 26 pg/mL at 180'; P = 0.399) but increased in control pigs (107 +/- 36 pg/mL at 15' and 499 +/- 216 pg/mL at 180'; P = 0.023). At 180' postreperfusion, redox-active iron was higher in control pigs versus modul