Browsing by Subject "Arabidopsis thaliana"

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  • Wang, Shuyuan; Alenius, Harri; El-Nezami, Hani; Karisola, Piia (2022)
    Titanium dioxide (TiO2) and zinc oxide (ZnO) nanoparticles (NPs) have attracted a great deal of attention due to their excellent electrical, optical, whitening, UV-adsorbing and bactericidal properties. The extensive production and utilization of these NPs increases their chances of being released into the environment and conferring unintended biological effects upon exposure. With the increasingly prevalent use of the omics technique, new data are burgeoning which provide a global view on the overall changes induced by exposures to NPs. In this review, we provide an account of the biological effects of ZnO and TiO2 NPs arising from transcriptomics in in vivo and in vitro studies. In addition to studies on humans and mice, we also describe findings on ecotoxicology-related species, such as Danio rerio (zebrafish), Caenorhabditis elegans (nematode) or Arabidopsis thaliana (thale cress). Based on evidence from transcriptomics studies, we discuss particle-induced biological effects, including cytotoxicity, developmental alterations and immune responses, that are dependent on both material-intrinsic and acquired/transformed properties. This review seeks to provide a holistic insight into the global changes induced by ZnO and TiO2 NPs pertinent to human and ecotoxicology.
  • Sageman-Furnas, Katelyn; Nurmi, Markus; Contag, Meike; Ploetner, Bjoern; Alseekh, Saleh; Wiszniewski, Andrew; Fernie, Alisdair R.; Smith, Lisa M.; Laitinen, Roosa A. E. (2022)
    Hybrids between Arabidopsis thaliana accessions are important in revealing the consequences of epistatic interactions in plants. F-1 hybrids between the A. thaliana accessions displaying either defense or developmental phenotypes have been revealing the roles of the underlying epistatic genes. The interaction of two naturally occurring alleles of the OUTGROWTH-ASSOCIATED KINASE (OAK) gene in Sha and Lag2-2, previously shown to cause a similar phenotype in a different allelic combination in A. thaliana, was required for the hybrid phenotype. Outgrowth formation in the hybrids was associated with reduced levels of salicylic acid, jasmonic acid and abscisic acid in petioles and the application of these hormones mitigated the formation of the outgrowths. Moreover, different abiotic stresses were found to mitigate the outgrowth phenotype. The involvement of stress and hormone signaling in outgrowth formation was supported by a global transcriptome analysis, which additionally revealed that TCP1, a transcription factor known to regulate leaf growth and symmetry, was downregulated in the outgrowth tissue. These results demonstrate that a combination of natural alleles of OAK regulates growth and development through the integration of hormone and stress signals and highlight the importance of natural variation as a resource to discover the function of gene variants that are not present in the most studied accessions of A. thaliana.
  • Tran, Cuong (Helsingin yliopisto, 2018)
    Salicylic acid (SA) is a well-known phytohormone involved in pathogen defense, development and controlling the cellular redox balance. In response to stresses, Arabidopsis thaliana synthesizes SA in the chloroplasts mainly via the isochorismate (IC) pathway: IC synthase (ICS) uses chorismate to produce IC, which is in turn converted to SA. ICS1 is the rate limiting enzyme in SA biosynthesis. The ICS1 protein localizes in the chloroplasts and contains a chloroplast transit peptide sequence, which is the first 45 amino acids in the N-terminus. Under different light conditions, plants employ different stress defense strategies. In a previous study, wild type (WT) Arabidopsis thaliana ecotype Columbia-0 grown under different light conditions [short day (SD; 8h light/16h dark), long day (LD; 16h light/8h dark) and 12h light/12h dark] was exposed to ozone for 1 hour (350 ppb). In a phosphoproteomic approach to study signaling mechanisms, ICS1 has been found as an in vivo phosphoprotein in ozone-treated plants grown under SD condition by mass spectrometry (MS). To analyze the phosphorylation of ICS1 in vitro, WT and phospho-negative ICS1 proteins were produced recombinantly as GST-fusion proteins in E. coli and used as the substrates for targeted kinase assays. Phospho-negative ICS1 protein was generated by mutating the phospho-sites to alanines by site-directed mutagenesis. ICS1 could be phosphorylated either in the cytosol or in the chloroplasts. Therefore, different cytosolic kinases, which are involved in various stress signaling events, and a chloroplast protein kinase that functions as a dominant regulator of chloroplast processes, were selected as the kinases to be tested. In this study, GST-ICS1 could be successfully phosphorylated in vitro. To a similar extent, GST-phospho-negative ICS1 was phosphorylated, indicating that ICS1 was phosphorylated in vitro at sites that were different from the ones found by MS from plant material. In order to study the identified phospho-sites in vivo, transgenic Arabidopsis carrying WT, phospho-positive, and phospho-negative ICS1 proteins were generated. Transgenic plants were confirmed by genotyping. Western blotting was carried out to evaluate ECFP-WT ICS1 protein expression. However, none of the transgenic plants could be confirmed to carry ECFP-WT ICS1. Additionally, an antibody against AtICS1 produced by the company Agrisera was tested for its specificity against the endogenous ICS1 protein. While the antibody could detect recombinant GST-ICS1 protein, ICS1 could not be detected from a total protein extract.
  • Eusuf, Saad Bin (Helsingin yliopisto, 2020)
    Stress response in plants is influenced by several external and internal factors and is executed in a modular way. Environmental stimuli or stress is sensed by cellular receptors and the signal is transduced inside cell via the phospho-activation of highly conserved intracellular signaling cascades like mitogen activated protein kinase (MAPK) cascades. The signal then activates biosynthesis pathways of major stress response hormones like Salicylic acid (SA). In Arabidopsis about 90% SA is synthesized via isochorismate pathway and Isochorismate synthase 1 (ICS1) is a rate limiting enzyme in this pathway. In this study, goal was to select transgenic ICS1 (homozygous) candidate lines from parent ICS1-CFP by selective regeneration. Then, by molecular and physiological characterization of transgenic ICS1-CFP plants, the function of ICS1 phosphorylation, more specifically, impact of different photoperiods (Long day; LD and Short day; SD) and stress conditions on ICS1 activity would have resolved. However, there were no homozygous candidate line from any parent ICS1-CFP plants after several screening. Nevertheless, ozone treated stress sensitivity test was performed with heterozygous ICS1-CFP candidate plants (T2 generation). Ozone treated stress depends on stomata factor because ozone enters into plants through stomata. Therefore, stomata index analysis was performed with sid2 and WT (Col-0) phenotypes and grown in LD and SD conditions. Since, stomata number was different between LD and SD plants of both sid2 and WT phenotypes, a different method named Xanthine-Xanthine oxidase (X/XO) treatment was applied that induce oxidative stress regardless of stomata. Although, WT and sid2 had shown sensitivity to the treatment, the overall cell death percentage was very low. Lastly, our aim was to observe the impact of different photoperiods on the activation of two particular MAPKs i.e MPK3 and MPK6 under stress conditions. The phosphorylated (P-MPK3 and P-MPK6) are found abundantly in ozone treated plants as an early response. In this experiment, plants were grown in both LD and SD, stressed with both ozone and X/XO treatments, the activation of P-MPK3 and P-MPK6 was observed by protein level analysis (western blotting) in detailed time course. Although, the activation was visualized in both LD and SD plants, qualitatively the pattern was similar between day type samples and activation signal was very weak in both stress methods. In addition, anti-ICS1 antibody provided by Agrsera TM was tested for its efficiency to detect endogenous ICS1 protein in plants in two experimental set-up. Although the antibody could detect overexpressed ICS1-CFP protein in samples, it was not that efficient to detect endogenous ICS1 in any of the experiments.
  • Wirthmueller, Lennart; Asai, Shuta; Rallapalli, Ghanasyam; Sklenar, Jan; Fabro, Georgina; Kim, Dae Sung; Lintermann, Ruth; Jaspers, Pinja; Wrzaczek, Michael; Kangasjärvi, Jaakko; MacLean, Daniel; Menke, Frank L. H.; Banfield, Mark J.; Jones, Jonathan D. G. (2018)
    The oomycete pathogen Hyaloperonospora arabidopsidis (Hpa) causes downy mildew disease on Arabidopsis. To colonize its host, Hpa translocates effector proteins that suppress plant immunity into infected host cells. Here, we investigate the relevance of the interaction between one of these effectors, HaRxL106, and Arabidopsis RADICAL-INDUCED CELL DEATH1 (RCD1). We use pathogen infection assays as well as molecular and biochemical analyses to test the hypothesis that HaRxL106 manipulates RCD1 to attenuate transcriptional activation of defense genes. We report that HaRxL106 suppresses transcriptional activation of salicylic acid (SA)-induced defense genes and alters plant growth responses to light. HaRxL106-mediated suppression of immunity is abolished in RCD1 loss-of-function mutants. We report that RCD1-type proteins are phosphorylated, and we identified Mut9-like kinases (MLKs), which function as phosphoregulatory nodes at the level of photoreceptors, as RCD1-interacting proteins. An mlk1,3,4 triple mutant exhibits stronger SA-induced defense marker gene expression compared with wild-type plants, suggesting that MLKs also affect transcriptional regulation of SA signaling. Based on the combined evidence, we hypothesize that nuclear RCD1/MLK complexes act as signaling nodes that integrate information from environmental cues and pathogen sensors, and that the Arabidopsis downy mildew pathogen targets RCD1 to prevent activation of plant immunity.
  • Kemppinen, Jasmin (Helsingin yliopisto, 2020)
    Reactive oxygen species (ROS) are one of the prominent groups of signal compounds that are produced in stress conditions such as excess light. Nuclear protein RADICAL-INDUCED CELL DEAT (RCD1) is sensitive to ROS and controls the expression of organelle components, e.g. mitochondrial alternative oxidases (AOX), thus balancing the redox-status of a plant cell. Plants have fast responses to fluctuating light conditions that happen even before gene expression: i.e. readjusting the capability to receive light energy between the two photosystems by state transitions and increasing the capacity to remove excess energy by non-photochemical quenching (NPQ). Various small auxiliary proteins function in these fast acclimation events. However, many of them are identified on gene level only. The goal of this master’s thesis is to describe the role of a hypothetical protein, PPD8 in Arabidopsis thaliana. We evaluate how PPD8 is associated with RCD1 and a chloroplast thiol-regulator enzyme NTRC. We created double (rcd1 ppd8) and triple mutant plant lines (rcd1 ppd8 ntrc) by crossing single knockout lines ppd8, rcd1 and ntrc. Photosynthetic performance, NPQ and sensitivity to ROS were observed in each line by using two different chlorophyll fluorescence measurement methods: pulse-amplitude-modulation (PAM) and novel OJIP imaging fluorometry. The leaves were exposed to methyl viologen (MV), which accelerates the chloroplastic ROS production in light, and also to hypoxic conditions in order to study how the effect of MV is altered in low concentrations of oxygen. Additionally, we examined the amount of photosynthetic proteins and stoichiometry of photosystems in ppd8, rcd1 and rcd1 ppd8 by immunological methods. Finally, PPD8 gene with attached hemagglutinin encoding tags was generated by cloning and reintroduced back to the ppd8 knockout lines. Plants lacking RCD1 are very tolerant against MV and ROS, but when rcd1 was crossed with ppd8 the resistance was suppressed. Both rcd1 ppd8 and ppd8 exhibited elevated chlorophyll fluorescence and NPQ values. The removal of PPD8 gene had an impact on the abundance and the stoichiometry of photosynthetic proteins reducing the plants’ performance. When RCD1, PPD8 and NTRC were simultaneously absent the plants had major defects: their NPQ and fluorescence values were drastically increased. Furthermore, several results hinted towards possible issues in the function of ATP synthase in ppd8 background plants. It is also known that NTRC regulates ATP synthase: taken together, the results suggest that PPD8 is necessary for a fully operative ATP synthase and photosynthetic machinery. By reintroducing PPD8 to knockout line ppd8, the phenotype could be reverted back to wild type -like, thus confirming the significance of the PPD8 gene product in plant.
  • Gawronski, Piotr; Burdiak, Pawel; Scharff, Lars B.; Mielecki, Jakub; Gorecka, Magdalena; Zaborowska, Magdalena; Leister, Dario; Waszczak, Cezary; Karpinski, Stanislaw (2021)
    Chloroplast-to-nucleus retrograde signaling is essential for cell function, acclimation to fluctuating environmental conditions, plant growth and development. The vast majority of chloroplast proteins are nuclear-encoded, and must be imported into the organelle after synthesis in the cytoplasm. This import is essential for the development of fully functional chloroplasts. On the other hand, functional chloroplasts act as sensors of environmental changes and can trigger acclimatory responses that influence nuclear gene expression. Signaling via mobile transcription factors (TFs) has been recently recognized as a way of communication between organelles and the nucleus. In this study, we performed a targeted reverse genetic screen to identify dual-localized TFs involved in chloroplast retrograde signaling during stress responses. We found that CHLOROPLAST IMPORT APPARATUS 2 (CIA2) has a functional plastid transit peptide, and can be located both in chloroplasts and the nucleus. Further, we found that CIA2, along with its homolog CIA2-like (CIL) are involved in the regulation of Arabidopsis responses to UV-AB, high light and heat shock. Finally, our results suggest that both CIA2 and CIL are crucial for chloroplast translation. Our results contribute to a deeper understanding of signaling events in the chloroplast-nucleus cross-talk.
  • Wang, Fang; Robson, T Matthew; Casal, Jorge J; Aphalo, Pedro J. (2020)
    The UV-A/blue photoreceptors phototropins and cryptochromes are both known to contribute to stomatal opening (∆gs) in blue light. However, their relative contributions to maintenance of gs in blue light through the whole photoperiod remains unknown. To elucidate this question, Arabidopsis phot1 phot2 and cry1 cry2 mutants (MTs) and their respective wild types (WTs) were irradiated with 200 μmol m-2 s-1 of blue-, green- or red-light (BL, GL or RL) throughout a 11-hour photoperiod. Stomatal conductance (gs) was higher under BL, than under RL or GL. Under RL, gs was not affected by either of the photoreceptor mutations, but under GL gs was slightly lower in cry1 cry2 than its WT. Under BL, the presence of phototropins was essential for rapid stomatal opening at the beginning of the photoperiod, while maximal stomatal opening beyond 3 h of irradiation required both phototropins and cryptochromes. Time courses of whole-plant net carbon assimilation rate (Anet) and the effective quantum yield of photosystem II photochemistry (ΦPSII) were consistent with an Anet-independent contribution of BL on gs both in phot1 phot2 and cry1 cry2 mutants. The changing roles of phototropins and cryptochromes through the day may allow more flexible coordination between gs and Anet.
  • Nuorti, Ninni (Helsingfors universitet, 2017)
    DRama Queen (DRQ) is an unknown conserved gene that might be involved with sensing cytokinin signalling in the roots of Arabidopsis thaliana. It was originally found in a mutant screen when a mutated version of it – mDRQ – was found to improve the cytokinin sensitivity of plants in a desensitized genetic background. This thesis was done to define the protoxylem phenotype of mDRQ single mutant in Columbia (Col-0) background and to test the functionality of the gene by analyzing the complementation lines. The protoxylem phenotypes were scored from fuchsin stained samples by DIC microscopy. Protoxylem differentiation in the root of A. thaliana is an outcome of a mutually inhibitory signalling mechanism of auxins and cytokinins. Analysis of protoxylem status is a good tool for studying the cytokinin signalling because the differentiation is affected both by increased and reduced signalling levels. High signalling causes loss of protoxylem phenotypes whereas whereas low cytokinin signalling leads to ectopic protoxylem formation. The data of this thesis was analysed by SPSS version 22 software using cumulative logit modelling for the analysis. It was considered to be the most suitable alternative for analysis since the protoxylem phenotype data is ordinal by its nature. A protoxylem phenotype distribution was defined for mDRQ line and it was statistically different from Col-0. Most of the complementation lines were functional at a statistically significant level though the phenotype distributions of the complementation lines were not identical with original background. The difference could possibly be explained by the fact that the mDRQ gene is partially functional and may influence the phenotype distributions.
  • Shapiguzov, Alexey; Nikkanen, Lauri; Fitzpatrick, Duncan; Vainonen, Julia P.; Gossens, Richard; Alseekh, Saleh; Aarabi, Fayezeh; Tiwari, Arjun; Blokhina, Olga; Panzarova, Klara; Benedikty, Zuzana; Tyystjärvi, Esa; Fernie, Alisdair R.; Trtilek, Martin; Aro, Eva-Mari; Rintamäki, Eevi; Kangasjarvi, Jaakko (2020)
    The Arabidopsis mutant rcd1 is tolerant to methyl viologen (MV). MV enhances the Mehler reaction, i.e. electron transfer from Photosystem I (PSI) to O-2, generating reactive oxygen species (ROS) in the chloroplast. To study the MV tolerance of rcd1, we first addressed chloroplast thiol redox enzymes potentially implicated in ROS scavenging. NADPH-thioredoxin oxidoreductase type C (NTRC) was more reduced in rcd1. NTRC contributed to the photosynthetic and metabolic phenotypes of rcd1, but did not determine its MV tolerance. We next tested rcd1 for alterations in the Mehler reaction. In rcd1, but not in the wild type, the PSI-to-MV electron transfer was abolished by hypoxic atmosphere. A characteristic feature of rcd1 is constitutive expression of mitochondrial dysfunction stimulon (MDS) genes that affect mitochondrial respiration. Similarly to rcd1, in other MDS-overexpressing plants hypoxia also inhibited the PSI-to-MV electron transfer. One possible explanation is that the MDS gene products may affect the Mehler reaction by altering the availability of O-2. In green tissues, this putative effect is masked by photosynthetic O-2 evolution. However, O-2 evolution was rapidly suppressed in MV-treated plants. Transcriptomic meta-analysis indicated that MDS gene expression is linked to hypoxic response not only under MV, but also in standard growth conditions. This article is part of the theme issue 'Retrograde signalling from endosymbiotic organelles'.
  • Kaeppel, Sandra; Eggeling, Ralf; Ruempler, Florian; Groth, Marco; Melzer, Rainer; Theissen, Guenter (2021)
    Key message We studied the DNA-binding profile of the MADS-domain transcription factor SEPALLATA3 and mutant variants by SELEX-seq. DNA-binding characteristics of SEPALLATA3 mutant proteins lead us to propose a novel DNA-binding mode. MIKC-type MADS-domain proteins, which function as essential transcription factors in plant development, bind as dimers to a 10-base-pair AT-rich motif termed CArG-box. However, this consensus motif cannot fully explain how the abundant family members in flowering plants can bind different target genes in specific ways. The aim of this study was to better understand the DNA-binding specificity of MADS-domain transcription factors. Also, we wanted to understand the role of a highly conserved arginine residue for binding specificity of the MADS-domain transcription factor family. Here, we studied the DNA-binding profile of the floral homeotic MADS-domain protein SEPALLATA3 by performing SELEX followed by high-throughput sequencing (SELEX-seq). We found a diverse set of bound sequences and could estimate the in vitro binding affinities of SEPALLATA3 to a huge number of different sequences. We found evidence for the preference of AT-rich motifs as flanking sequences. Whereas different CArG-boxes can act as SEPALLATA3 binding sites, our findings suggest that the preferred flanking motifs are almost always the same and thus mostly independent of the identity of the central CArG-box motif. Analysis of SEPALLATA3 proteins with a single amino acid substitution at position 3 of the DNA-binding MADS-domain further revealed that the conserved arginine residue, which has been shown to be involved in a shape readout mechanism, is especially important for the recognition of nucleotides at positions 3 and 8 of the CArG-box motif. This leads us to propose a novel DNA-binding mode for SEPALLATA3, which is different from that of other MADS-domain proteins known.
  • Johansson, Karin S. L.; El-Soda, Mohamed; Pagel, Ellen; Meyer, Rhonda C.; Toldsepp, Kadri; Nilsson, Anders K.; Brosche, Mikael; Kollist, Hannes; Uddling, Johan; Andersson, Mats X. (2020)
    Background and Aims The stomatal conductance (g(s)) of most plant species decreases in response to elevated atmospheric CO2 concentration. This response could have a significant impact on plant water use in a future climate. However, the regulation of the CO2 induced stomatal closure response is not fully understood. Moreover, the potential genetic links between short-term (within minutes to hours) and long-term (within weeks to months) responses of g(s) to increased atmospheric CO2 have not been explored. Methods We used Arabidopsis thaliana recombinant inbred lines originating from accessions Col-0 (strong CO2 response) and C24 (weak CO2 response) to study short- and long-term controls of g(s) Quantitative trait locus (QTL) mapping was used to identify loci controlling short- and long-term g(s) responses to elevated CO2 as well as other stomata-related traits. Key Results Short- and long-term stomatal responses to elevated CO2 were significantly correlated. Both short-and long-term responses were associated with a QTL, at the end of chromosome 2. The location of this QTL was confirmed using near-isogonic lines and it was fine-mapped to a 410-kb region. The QTL did not correspond to any known gene involved in stomatal closure and had no effect on the responsiveness to abscisic acid. Additionally, we identified numerous other loci associated with stomatal regulation. Conclusions We identified and confirmed the effect of a strong QTL corresponding to a yet unknown regulator of stomatal closure in response to elevated CO2 concentration. The correlation between short- and long-term stomatal CO2 responses and the genetic link between these traits highlight the importance of understanding guard cell CO2 signalling to predict and manipulate plant water use in a world with increasing atmospheric CO2 concentration. This study demonstrates the power of using natural variation to unravel the genetic regulation of complex traits.
  • Deger, Aysin Guzel; Scherzer, Sönke; Nuhkat, Maris; Kedzierska, Justyna; Kollist, Hannes; Brosche, Mikael; Unyayar, Serpil; Boudsocq, Marie; Hedrich, Rainer; Roelfsema, M. Rob G. (2015)
    During infection plants recognize microbe-associated molecular patterns (MAMPs), and this leads to stomatal closure. This study analyzes the molecular mechanisms underlying this MAMP response and its interrelation with ABA signaling. Stomata in intact Arabidopsis thaliana plants were stimulated with the bacterial MAMP flg22, or the stress hormone ABA, by using the noninvasive nanoinfusion technique. Intracellular double-barreled microelectrodes were applied to measure the activity of plasma membrane ion channels. Flg22 induced rapid stomatal closure and stimulated the SLAC1 and SLAH3 anion channels in guard cells. Loss of both channels resulted in cells that lacked flg22-induced anion channel activity and stomata that did not close in response to flg22 or ABA. Rapid flg22-dependent stomatal closure was impaired in plants that were flagellin receptor (FLS2)-deficient, as well as in the ost1-2 (Open Stomata 1) mutant, which lacks a key ABA-signaling protein kinase. By contrast, stomata of the ABA protein phosphatase mutant abi1-1 (ABscisic acid Insensitive 1) remained flg22-responsive. These data suggest that the initial steps in flg22 and ABA signaling are different, but that the pathways merge at the level of OST1 and lead to activation of SLAC1 and SLAH3 anion channels.
  • Rai, Neha; Neugart, Susanne; Yan, Yan; Wang, Fang; Siipola, Sari M.; Lindfors, Anders V.; Winkler, Jana Barbro; Albert, Andreas; Brosche, Mikael; Lehto, Tarja; Morales, Luis O.; Aphalo, Pedro J. (2019)
    Cryptochromes (CRYs) and UV RESISTANCE LOCUS 8 (UVR8) photoreceptors perceive UV-A/blue (315-500 nm) and UV-B (280-315 nm) radiation in plants, respectively. While the roles of CRYs and UVR8 have been studied in separate controlled-environment experiments, little is known about the interaction between these photoreceptors. Here, Arabidopsis wild-type Ler, CRYs and UVR8 photoreceptor mutants (uvr8-2, cry1cry2 and cry1cry2uvr8-2), and a flavonoid biosynthesis-defective mutant (tt4) were grown in a sun simulator. Plants were exposed to filtered radiation for 17 d or for 6 h, to study the effects of blue, UV-A, and UV-B radiation. Both CRYs and UVR8 independently enabled growth and survival of plants under solar levels of UV, while their joint absence was lethal under UV-B. CRYs mediated gene expression under blue light. UVR8 mediated gene expression under UV-B radiation, and in the absence of CRYs, also under UV-A. This negative regulation of UVR8-mediated gene expression by CRYs was also observed for UV-B. The accumulation of flavonoids was also consistent with this interaction between CRYs and UVR8. In conclusion, we provide evidence for an antagonistic interaction between CRYs and UVR8 and a role of UVR8 in UV-A perception.
  • Cui, Fuqiang; Brosche, Mikael; Shapiguzov, Alexey; He, Xin-Qiang; Vainonen, Julia P.; Leppala, Johanna; Trotta, Andrea; Kangasjarvi, Saijaliisa; Salojarvi, Jarkko; Kangasjarvi, Jaakko; Overmyer, Kirk (2019)
    Reactive oxygen species (ROS) are key signalling intermediates in plant metabolism, defence, and stress adaptation. In plants, both the chloroplast and mitochondria are centres of metabolic control and ROS production, which coordinate stress responses in other cell compartments. The herbicide and experimental tool, methyl viologen (MV) induces ROS generation in the chloroplast under illumination, but is also toxic in non-photosynthetic organisms. We used MV to probe plant ROS signalling in compartments other than the chloroplast. Taking a genetic approach in the model plant Arabidopsis (Arabidopsis thaliana), we used natural variation, QTL mapping, and mutant studies with MV in the light, but also under dark conditions, when the chloroplast electron transport is inactive. These studies revealed a light-independent MV-induced ROS-signalling pathway, suggesting mitochondrial involvement. Mitochondrial Mn SUPEROXIDE DISMUTASE was required for ROS-tolerance and the effect of MV was enhanced by exogenous sugar, providing further evidence for the role of mitochondria. Mutant and hormone feeding assays revealed roles for stress hormones in organellar ROS-responses. The radical-induced cell death1 mutant, which is tolerant to MV-induced ROS and exhibits altered mitochondrial signalling, was used to probe interactions between organelles. Our studies suggest that mitochondria are involved in the response to ROS induced by MV in plants.
  • Zamora, Olena; Schulze, Sebastian; Azoulay-Shemer, Tamar; Parik, Helen; Unt, Jaanika; Brosché, Mikael; Schroeder, Julian I.; Yarmolinsky, Dmitry; Kollist, Hannes (2021)
    Jasmonic acid (JA) and salicylic acid (SA) regulate stomatal closure, preventing pathogen invasion into plants. However, to what extent abscisic acid (ABA), SA and JA interact, and what the roles of SA and JA are in stomatal responses to environmental cues, remains unclear. Here, by using intact plant gas-exchange measurements in JA and SA single and double mutants, we show that stomatal responsiveness to CO2, light intensity, ABA, high vapor pressure deficit and ozone either did not or, for some stimuli only, very slightly depended upon JA and SA biosynthesis and signaling mutants, including dde2, sid2, coi1, jai1, myc2 and npr1 alleles. Although the stomata in the mutants studied clearly responded to ABA, CO2, light and ozone, ABA-triggered stomatal closure in npr1-1 was slightly accelerated compared with the wild type. Stomatal reopening after ozone pulses was quicker in the coi1-16 mutant than in the wild type. In intact Arabidopsis plants, spraying with methyl-JA led to only a modest reduction in stomatal conductance 80 min after treatment, whereas ABA and CO2 induced pronounced stomatal closure within minutes. We could not document a reduction of stomatal conductance after spraying with SA. Coronatine-induced stomatal opening was initiated slowly after 1.5-2.0 h, and reached a maximum by 3 h after spraying intact plants. Our results suggest that ABA, CO2 and light are major regulators of rapid guard cell signaling, whereas JA and SA could play only minor roles in the whole-plant stomatal response to environmental cues in Arabidopsis and Solanum lycopersicum (tomato).
  • Biniaz, Yaser; Tahmasebi, Aminallah; Afsharifar, Alireza; Tahmasebi, Ahmad; Poczai, Peter (2022)
    Environmental stresses adversely affect crop growth and yield, resulting in major losses to plants. These stresses occur simultaneously in nature, and we therefore conducted a meta-analysis in this study to identify differential and shared genes, pathways, and transcriptomic mechanisms involved in Arabidopsis response to biotic and abiotic stresses. The results showed a total of 436/21 significant up-/downregulated differentially expressed genes (DEGs) in response to biotic stresses, while 476 and 71 significant DEGs were respectively up- and downregulated in response to abiotic stresses in Arabidopsis thaliana. In addition, 21 DEGs (2.09%) were commonly regulated in response to biotic and abiotic stresses. Except for WRKY45 and ATXTH22, which were respectively up-/down- and down-/upregulated in response to biotic and abiotic stresses, other common DEGs were upregulated in response to all biotic and abiotic treatments. Moreover, the transcription factors (TFs) bHLH, MYB, and WRKY were the common TFs in response to biotic and abiotic stresses. In addition, ath-miR414 and ath-miR5658 were identified to be commonly expressed in response to both biotic and abiotic stresses. The identified common genes and pathways during biotic and abiotic stresses may provide potential candidate targets for the development of stress resistance breeding programs and for the genetic manipulation of crop plants.
  • Zinchenko, Ganna (Helsingfors universitet, 2016)
    The characterization of flower-specific ubiquitin-proteasome system (UPS) components and identification of their functional molecular networks will help to elucidate the involvement of UPS in regulating flower development and/or flowering time and, therefore, reproductive success of the plant. UPS component COP1 ubiquitin E3 ligase is known to regulate flowering time. The earlier data suggests that COP1 could be involved in regulating cytokinin signaling possibly through Arabidopsis Response Regulator1 (ARR1) ubiquitination. ARR1 is a B-type cytokinin response regulator, and it has recently been shown to be an unstable protein. Furthermore, KMD, F-box protein in SCF E3 ligase complex, has been shown to interact with ARR1 as well. The aim of this study is the characterization of COP1 interaction with novel target proteins ARR1, ARR2, ARR10 and ARR12 that appear to be regulated in different ways. Moreover, KMD proteins were included within the study as a possible competitor of COP1 for interaction with ARR1. In order to perform interactome studies, yeast two-hybrid assay with a preceding molecular cloning of the genes of interest was used. The results can be used to unravel the role of ubiquitin mediated regulation of cytokinin pathway.
  • Järvi, Sari; Isojärvi, Janne; Kangasjärvi, Saijaliisa; Salojärvi, Jarkko; Mamedov, Fikret; Suorsa, Marjaana; Aro, Eva-Mari (2016)
    Chloroplasts play an important role in the cellular sensing of abiotic and biotic stress. Signals originating from photosynthetic light reactions, in the form of redox and pH changes, accumulation of reactive oxygen and electrophile species or stromal metabolites are of key importance in chloroplast retrograde signaling. These signals initiate plant acclimation responses to both abiotic and biotic stresses. To reveal the molecular responses activated by rapid fluctuations in growth light intensity, gene expression analysis was performed with Arabidopsis thaliana wild type and the tlp18.3 mutant plants, the latter showing a stunted growth phenotype under fluctuating light conditions (Biochem. J, 406, 415-425). Expression pattern of genes encoding components of the photosynthetic electron transfer chain did not differ between fluctuating and constant light conditions, neither in wild type nor in tlp18.3 plants, and the composition of the thylakoid membrane protein complexes likewise remained unchanged. Nevertheless, the fluctuating light conditions repressed in wild-type plants a broad spectrum of genes involved in immune responses, which likely resulted from shade-avoidance responses and their intermixing with hormonal signaling. On the contrary, in the tlp18.3 mutant plants there was an imperfect repression of defense-related transcripts upon growth under fluctuating light, possibly by signals originating from minor malfunction of the photosystem II (PSII) repair cycle, which directly or indirectly modulated the transcript abundances of genes related to light perception via phytochromes. Consequently, a strong allocation of resources to defense reactions in the tlp18.3 mutant plants presumably results in the stunted growth phenotype under fluctuating light.