Joëlle Gérard

Ozone‐induced changes in photosynthesis and photorespiration of hybrid poplar in relation to the developmental stage of the leaves

Young poplar trees (Populus tremula Michx. x Populus alba L. clone INRA 717-1B4) were subjected to 120 ppb of ozone for 35 days in phytotronic chambers. Treated trees displayed precocious leaf senescence and visible symptoms of injury (dark brown/black upper surface stippling) exclusively observed on fully expanded leaves. In these leaves, ozone reduced parameters related to photochemistry (Chl content and maximum rate of photosynthetic electron transport) and photosynthetic CO(2) fixation [net CO(2) assimilation, Rubisco (ribulose-1,5-bisphosphate carboxylase oxygenase) activity and maximum velocity of Rubisco for carboxylation]. In fully expanded leaves, the rate of photorespiration as estimated from Chl fluorescence was markedly impaired by the ozone treatment together with the activity of photorespiratory enzymes (Rubisco and glycolate oxidase). Immunoblot analysis revealed a decrease in the content of serine hydroxymethyltransferase in treated mature leaves, while the content of the H subunit of the glycine decarboxylase complex was not modified. Leaves in the early period of expansion were exempt from visible symptoms of injury and remained unaffected as regards all measured parameters. Leaves reaching full expansion under ozone exposure showed potential responses of protection (stimulation of mitochondrial respiration and transitory stomatal closure). Our data underline the major role of leaf phenology in ozone sensitivity of photosynthetic processes and reveal a marked ozone-induced inhibition of photorespiration.

The CHAP domain of Cse functions as an endopeptidase that acts at mature septa to promote Streptococcus thermophilus cell separation

Cell separation is dependent on cell wall hydrolases that cleave the peptidoglycan shared between daughter cells. In Streptococcus thermophilus, this step is performed by the Cse protein whose depletion resulted in the formation of extremely long chains of cells. Cse, a natural chimeric enzyme created by domain shuffling, carries at least two important domains for its activity: the LysM expected to be responsible for the cell wall‐binding and the CHAP domain predicted to contain the active centre. Accordingly, the localization of Cse on S. thermophilus cell surface has been undertaken by immunogold electron and immunofluorescence microscopies using of antibodies raised against the N‐terminal end of this protein. Immunolocalization shows the presence of the Cse protein at mature septa. Moreover, the CHAP domain of Cse exhibits a cell wall lytic activity in zymograms performed with cell walls of Micrococcus lysodeikticus, Bacillus subtilis and S. thermophilus. Additionally, RP‐HPLC analysis of muropeptides released from B. subtilis and S. thermophilus cell wall after digestion with the CHAP domain shows that Cse is an endopeptidase. Altogether, these results suggest that Cse is a cell wall hydrolase involved in daughter cell separation of S. thermophilus.

Distinct responses to ozone of abaxial and adaxial stomata in three Euramerican poplar genotypes

Ozone induces stomatal sluggishness, which impacts photosynthesis and transpiration. Stomatal responses to variation of environmental parameters are slowed and reduced by ozone and may be linked to difference of ozone sensitivity. Here we determine the ozone effects on stomatal conductance of each leaf surface. Potential causes of this sluggish movement, such as ultrastructural or ionic fluxes modification, were studied independently on both leaf surfaces of three Euramerican poplar genotypes differing in ozone sensitivity and in stomatal behaviour. The element contents in guard cells were linked to the gene expression of ion channels and transporters involved in stomatal movements, directly in microdissected stomata. In response to ozone, we found a decrease in the stomatal conductance of the leaf adaxial surface correlated with high calcium content in guard cells compared with a slight decrease on the abaxial surface. No ultrastructural modifications of stomata were shown except an increase in the number of mitochondria. The expression of vacuolar H(+) /Ca(2+) -antiports (CAX1 and CAX3 homologs), β-carbonic anhydrases (βCA1 and βCA4) and proton H(+) -ATPase (AHA11) genes was strongly decreased under ozone treatment. The sensitive genotype characterized by constitutive slow stomatal response was also characterized by constitutive low expression of genes encoding vacuolar H(+) /Ca(2+) -antiports.

Ozone exposure and flux-based response functions for photosynthetic traits in wheat, maize and poplar.

Ozone exposure- and dose-response relationships based on photosynthetic leaf traits (CO2 assimilation, chlorophyll content, Rubisco and PEPc activities) were established for wheat, maize and poplar plants grown in identical controlled conditions, providing a comparison between crop and tree species, as well as between C3 and C4 plants. Intra-specific variability was addressed by comparing two wheat cultivars with contrasting ozone tolerance. Depending on plant models and ozone levels, first-order, second-order and segmented linear regression models were used to derive ozone response functions. Overall, flux-based functions appeared superior to exposure-based functions in describing the data, but the improvement remained modest. The best fit was obtained using the POD0.5 for maize and POD3 for poplar. The POD6 appeared relevant for wheat, although intervarietal differences were found. Our results suggest that taking into account the dynamics of leaf antioxidant capacity could improve current methods for ozone risk assessment for plants.

Properties of mitochondria isolated from greening soybean and lupin tissues

Abstract In germinating soybean and lupin seedlings differing in their amount of lipid reserves, the activity of cotyledon mitochondria was found to be successively associated with the glyoxysomal and the peroxisomal systems. However, the level of glyoxysomal activity was lower in lipid poor lupin tissues. A peak in succinate oxidation by purified mitochondria was observed in the fully green cotyledons of the two species. A marked cyanide resistance developed in soybean cotyledon mitochondria during the greening of the tissue whereas lupin cotyledon mitochondria remained highly cyanides sensitive. This difference between the two species was also found in purified leaf mitochondria oxidizing succinate, malate or glycine. Moreover, the reoxidation of NADH provided during malate and glycine oxidation mainly used the rotenone and cyanide-resistant pathways in soybean mitochondria whereas the complex I and the cytochrome pathway were largely involved in lupin mitochondria.

Antioxidative responses of three oak species under ozone and water stress conditions

Plants are frequently exposed to adverse environmental conditions such as drought and ozone (O3). Under these conditions, plants can survive due to their ability to adjust their metabolism. The aim of the present study was to compare the detoxification mechanisms of three oak species showing different O3 sensitivity and water use strategy. Two-year-old seedlings of Quercus ilex, Q. pubescens and Q. robur were grown under the combination of three levels of O3 (1.0, 1.2 and 1.4 times the ambient O3 concentration) and three levels of water availability (on average 100, 80 and 42% of field capacity i.e. well-watered, moderate drought and severe drought, respectively) in an O3 Free Air Controlled Exposure facility. Ozone and drought induced the accumulation of reactive oxygen species (ROS) and this phenomenon was species-specific. Sometimes, ROS accumulation was not associated with membrane injury suggesting that several antioxidative defence mechanisms inhibited or alleviated the oxidative damage. Both O3 and drought increased total carotenoids that were able to prevent the peroxidation action by free radicals in Q. ilex, as confirmed by unchanged malondialdehyde by-product values. The concomitant decrease of total flavonoids may be related to the consumption of these compounds by the cell to inhibit the accumulation of hydrogen peroxide. Unchanged total phenols confirmed that Q. ilex has a superior ability to counteract oxidative conditions. Similar responses were found in Q. pubescens, although the negative impact of both factors was less efficiently faced than in the sympatric Q. ilex. In Q. robur, high O3 concentrations and severe drought induced a partial rearrangement of the phenylpropanoid pathways. These antioxidative mechanisms were not able to protect the cell structure (as confirmed by ROS accumulation) suggesting that Q. robur showed a lower degree of tolerance than the other two species.

Integrated analysis of the detoxification responses of two Euramerican poplar genotypes exposed to ozone and water deficit: Focus on the ascorbate-glutathione cycle

Ozone (O3) and drought increase tree oxidative stress. To protect forest health, we need to improve risk assessment, using metric model such as the phytotoxic O3 dose above a threshold of y nmol·m-2·s-1 (PODy), while taking into account detoxification mechanisms and interacting stresses. The impact of drought events on the effect of O3 pollution deserves special attention. Water deficit may decrease O3 entrance into the leaves by reducing stomatal opening; however, water deficit also induces changes in cell redox homeostasis. Besides, the behaviour of the cell antioxidative charge in case of stress combination (water deficit and O3) still remains poorly investigated. To decipher the response of detoxification mechanisms relatively to the Halliwell-Asada-Foyer cycle (HAF), we exposed poplar saplings (Populus nigra × deltoides) composed of two genotypes (Carpaccio and Robusta), to various treatments for 17 days, i.e. i) mild water deficit, ii) 120 ppb O3, and iii) a combination of these two treatments. Ozone similarly impacted the growth of the two genotypes, with an important leaf loss. Water deficit decreased growth by almost one third as compared to the control plants. As for the combined treatment, water deficit protected the saplings from leaf ozone injury, but with an inhibitory effect on growth. The pool of total ascorbate was not modified by the different treatments, while the pool of total glutathione increased with POD0. We noticed a few differences between the two genotypes, particularly concerning the activity of monodehydroascorbate reductase and glutathione reductase relatively to POD0. The expression profiles of genes coding for the dehydroascorbate reductase and glutathione reductase isoforms differed, probably in link with the putative localisation of ROS production in response to water deficit and ozone, respectively. Our result would argue for a major role of MDHAR, GR and glutathione in the preservation of the redox status.

Both gravistimulation onset and removal trigger an increase of cytoplasmic free calcium in statocytes of roots grown in microgravity

Gravity is a permanent environmental signal guiding plant growth and development. Gravity sensing in plants starts with the displacement of starch-filled plastids called statoliths, ultimately leading to auxin redistribution and organ curvature. While the involvement in gravity sensing of several actors such as calcium is known, the effect of statolith displacement on calcium changes remains enigmatic. Microgravity is a unique environmental condition offering the opportunity to decipher this link. In this study, roots of Brassica napus were grown aboard the International Space Station (ISS) either in microgravity or in a centrifuge simulating Earth gravity. The impact of short simulated gravity onset and removal was measured on statolith positioning and intracellular free calcium was assessed using pyroantimonate precipitates as cytosolic calcium markers. Our findings show that a ten-minute onset or removal of gravity induces very low statolith displacement, but which is, nevertheless, associated with an increase of the number of pyroantimonate precipitates. These results highlight that a change in the cytosolic calcium distribution is triggered in absence of a significant statolith displacement.