A global non-invasive methodology for the phenotyping of potato under water deficit conditions using imaging, physiological and molecular tools.

BACKGROUND: Drought is a major consequence of global heating that has negative impacts on agriculture. Potato is a drought-sensitive crop; tuber growth and dry matter content may both be impacted. Moreover, water deficit can induce physiological disorders such as glassy tubers and internal rust spots. The response of potato plants to drought is complex and can be affected by cultivar type, climatic and soil conditions, and the point at which water stress occurs during growth. The characterization of adaptive responses in plants presents a major phenotyping challenge. There is therefore a demand for the development of non-invasive analytical techniques to improve phenotyping. RESULTS: This project aimed to take advantage of innovative approaches in MRI, phenotyping and molecular biology to evaluate the effects of water stress on potato plants during growth. Plants were cultivated in pots under different water conditions. A control group of plants were cultivated under optimal water uptake conditions. Other groups were cultivated under mild and severe water deficiency conditions (40 and 20% of field capacity, respectively) applied at different tuber growth phases (initiation, filling). Water stress was evaluated by monitoring soil water potential. Two fully-equipped imaging cabinets were set up to characterize plant morphology using high definition color cameras (top and side views) and to measure plant stress using RGB cameras. The response of potato plants to water stress depended on the intensity and duration of the stress. Three-dimensional morphological images of the underground organs of potato plants in pots were recorded using a 1.5 T MRI scanner. A significant difference in growth kinetics was observed at the early growth stages between the control and stressed plants. Quantitative PCR analysis was carried out at molecular level on the expression patterns of selected drought-responsive genes. Variations in stress levels were seen to modulate ABA and drought-responsive ABA-dependent and ABA-independent genes. CONCLUSIONS: This methodology, when applied to the phenotyping of potato under water deficit conditions, provides a quantitative analysis of leaves and tubers properties at microstructural and molecular levels. The approaches thus developed could therefore be effective in the multi-scale characterization of plant response to water stress, from organ development to gene expression.

Embolismo graso cerebral como una rara complicación de liposucción con abdominoplastia

El síndrome de embolismo graso (SEG) se presenta más frecuentemente después de una fractura de huesos largos o de un procedimientoquirúrgico ortopédico. Se describen, sin embargo, casos del SEG en condiciones no traumáticas como la resucitación cardiopulmonar, pancreatitis, quemaduras, nutrición parenteraly hasta en las crisis del siclémico. A pesar de que la liposucción y la abdominoplastia son procedimientos corrientes en la cirugía estética, los casos bien documentados delSEG después de éstos son muy raros. Sebdescribe el caso de una paciente que desarrolló un embolismo graso cerebral (EGC) con fallo multiorgánico después de una liposucción con abdominoplastia...

The lysine-ketoglutarate reductase-saccharopine dehydrogenase is involved in the osmo-induced synthesis of pipecolic acid in rapeseed leaf tissues.

Higher plant responses to abiotic stresses are associated with physiological and biochemical changes triggering a number of metabolic adjustments. We focused on L-lysine catabolism, and have previously demonstrated that degradation of this amino acid is osmo-regulated at the level of lysine-ketoglutarate reductase (LKR, EC 1.5.1.8) and saccharopine dehydrogenase (SDH, EC 1.5.1.9) in Brassica napus. LKR and SDH activities are enhanced by decreasing osmotic potential and decrease when the upshock osmotic treatment is followed by a downshock osmotic one. Moreover we have shown that the B. napus LKR/SDH gene is up-regulated in osmotically-stressed tissues. The LKR/SDH activity produces alpha-aminoadipate semialdehyde which could be further converted into alpha-aminoadipate and acetyl CoA. Alternatively alpha-aminoadipate could behave as a precursor for pipecolic acid. Pipecolic acid is described as an osmoprotectant in bacteria and is co-accumulated with proline in halophytic plants. We suggest that osmo-induction of the LKR/SDH activity could be partly responsible for pipecolic acid accumulation. This proposal has been assessed in this study through pipecolic acid amounts determination in rape leaf discs subjected to various upshift and downshift osmotic treatments. Changes in pipecolic acid level actually behave as those observed for LKR and SDH activities, since it increases or decreases in rape leaf discs treated under hyper- or hypo-osmotic conditions, respectively. In addition we show that pipecolic acid level is positively correlated with the external osmotic potential as well as with the duration of the applied treatment. On the other hand pipecolic acid level is related to the availability of L-lysine and not to that of D-lysine. Collectively the results obtained demonstrate that lysine catabolism through LKR/SDH activity is involved in osmo-induced synthesis of pipecolic acid.

Hawthorn extracts inhibit LDL oxidation.

Polyphenol-rich diet decreases cardiovascular risk. LDL oxidation is the primary event in atherosclerosis plaque formation and antioxidants such as polyphenols were shown to inhibit LDL oxidation and atherosclerosis development. Hawthorn (Crataegus) and derived pharmaceuticals are rich in polyphenols and already prescribed to treat moderate heart failure, nervousness and sleep disorders. Extracts either from fresh plant parts (flower buds, flowers, young leaves or green fruits) or from dried pharmaceutical parts (flowers and flowering tops) were previously shown to be effective inhibitors of lipoperoxidation and scavengers of oxygen species. In this study, the capacity of total and ethyl-acetate extracts from dried pharmaceutical flowers, tops and fruits to inhibit Cu(2+)-induced LDL oxidation was tested. This capacity was positively linked to their content in total polyphenols, proanthocyanidins (global and oligomeric forms), as well as to their content in two individual phenolics: a flavanol, the dimeric procyanidin B2 and a flavonol glycoside, hyperoside. Flavanol-type phenolics showed to be higher active than the majority of the flavonoids tested in inhibiting Cu(2+)-induced LDL peroxidation. This study suggests that hawthorn could be a source of polyphenols able to inhibit LDL oxidation.

Phenolic compounds and antioxidant activities of buckwheat (Fagopyrum esculentum Moench) hulls and flour.

The interest of polyphenolics as therapeutic agents against diseases involving radical damage is growing. The phenolic contents of the hulls and flour from the seeds of Fagopyrum esculentum (French variety 'La Harpe') (total phenols, flavonoids, total flavanols, oligomeric proanthocyanidins) are compared with the antioxidative effects of the extracts against reactive oxygen species: hydrogen peroxide, hypochlorous acid, superoxide anion. The higher efficiency of the flour extract can be related to its higher flavanolic content rather than to flavonoids which are predominant in the hull extract.

[Proline accumulation by tomato leaf tissue in response to salinity]. / Accumulation de proline dans les tissus foliaires de tomate en réponse à la salinité.

The capacity of tomato leaf tissues to accumulate proline in response to a salt shock (150 mM NaCl) applied to excised shoots, leaves, leaflets or leaf discs was determined and compared to that of whole plants grown at the same salinity. The associated changes in free amino acids, Na+, K+ and Cl- contents were also investigated. In excised organs treated for 80 h, up to 200 mumol g-1 DW of proline were accumulated, whereas the amount of proline in leaf discs did not exceed a value ten-fold lower. In the whole plants subjected to salinity the Na+, Cl- and K+ contents remained low in comparison to that observed in excised organs. Proline and other amino acids increased more slowly in whole plants than in excised shoots. The contribution of roots and vascular tissues to the control of Na+ and Cl- accumulation and to the regulation of proline metabolism are discussed.

Mutational analysis of the [Het-s] prion analog of Podospora anserina. A short N-terminal peptide allows prion propagation.

The het-s locus is one of nine known het (heterokaryon incompatibility) loci of the fungus Podospora anserina. This locus exists as two wild-type alleles, het-s and het-S, which encode 289 amino acid proteins differing at 13 amino acid positions. The het-s and het-S alleles are incompatible as their coexpression in the same cytoplasm causes a characteristic cell death reaction. We have proposed that the HET-s protein is a prion analog. Strains of the het-s genotype exist in two phenotypic states, the neutral [Het-s*] and the active [Het-s] phenotype. The [Het-s] phenotype is infectious and is transmitted to [Het-s*] strains through cytoplasmic contact. het-s and het-S were associated in a single haploid nucleus to generate a self-incompatible strain that displays a restricted and abnormal growth. In the present article we report the molecular characterization of a collection of mutants that restore the ability of this self-incompatible strain to grow. We also describe the functional analysis of a series of deletion constructs and site-directed mutants. Together, these analyses define positions critical for reactivity and allele specificity. We show that a 112-amino-acid-long N-terminal peptide of HET-s retains [Het-s] activity. Moreover, expression of a mutant het-s allele truncated at position 26 is sufficient to allow propagation of the [Het-s] prion analog.

Combination of biomolecular and stable isotope techniques to determine the origin of organic matter used by bacterial communities: application to sediment.

Natural isotopic composition is a good tool to trace organic matter in ecosystems. Recent studies used a combination of molecular and stable isotope techniques to determine the origin of the organic carbon used by bacteria in the water column. In our study, we show that this procedure can be used for analysis of sediment bacterial communities with few modifications. In the water column, bacterial recovery is done before DNA extraction. In the sediment, we tested qualitatively and quantitatively a direct and indirect extraction of DNA. The direct extraction was the most efficient. It recovered between 3.1 and 15.8 microg DNA g(-1) dry sediment and the contamination of field samples by eucaryotic DNA was less than 13%. In this preliminary study of the salt marsh ecosystem, the delta(13)C values of DNA (-26 to - 24%) recovered from the sediment were close to the delta(13)C values of halophytic plants (-26.4 and - 25.3%) showing a relationship between plants and microorganisms. Thus, this procedure can be used to trace the flow of carbon through the sediment microbial biomass and to understand the variation of bacterial activity according to the inputs of allocthonous and autochtonous organic matter.

Suppression of the osmoinduced proline response of rapeseed leaf discs by polyamines.

Rapeseed leaf discs (RLD) subjected to upshock osmotic stress accumulate proline (Pro). Di- and polyamines (PA) supplied to the external medium suppressed Pro accumulation. These effects were dependent not only on diamine and PA concentrations but also on their cationic charge. The suppression of Pro accumulation required that diamine and PA be taken up and further accumulated in the leaf tissues. Glycine betaine (GB) also inhibited Pro accumulation, with the effects of GB and PA being additive. Experiments to elucidate the mechanism(s) responsible for the inhibitory effect of spermine (Spm) indicated that it could be simulated with methionine sulfoximine (MSO), a potent inhibitor of glutamine synthetase. The inhibitory effects of Spm and MSO were both alleviated by supplying glutamine to the RLD. In addition, Spm as well as MSO increased glutamate content, indicating that these compounds could inhibit the conversion of glutamate to proline. A comparison of the changes in chlorophyll and protein content of RLD osmotreated with or without added Spm indicates that this PA behaves as an antisenescent compound, preventing chlorophyll breakdown and proteolysis and hence the conversion of amino acids to Pro. Since the PA concentrations used in this work were much higher than the endogenous concentrations in RLD, the significance of PA under osmotic stress remains unclear. This study shows, however, that PA can suppress Pro accumulation.

The protein product of the het-s heterokaryon incompatibility gene of the fungus Podospora anserina behaves as a prion analog.

The het-s locus of Podospora anserina is a heterokaryon incompatibility locus. The coexpression of the antagonistic het-s and het-S alleles triggers a lethal reaction that prevents the formation of viable heterokaryons. Strains that contain the het-s allele can display two different phenotypes, [Het-s] or [Het-s*], according to their reactivity in incompatibility. The detection in these phenotypically distinct strains of a protein expressed from the het-s gene indicates that the difference in reactivity depends on a posttranslational difference between two forms of the polypeptide encoded by the het-s gene. This posttranslational modification does not affect the electrophoretic mobility of the protein in SDS/PAGE. Several results suggest a similarity of behavior between the protein encoded by the het-s gene and prions. The [Het-s] character can propagate in [Het-s*] strains as an infectious agent, producing a [Het-s*] --> [Het-s] transition, independently of protein synthesis. Expression of the [Het-s] character requires a functional het-s gene. The protein present in [Het-s] strains is more resistant to proteinase K than that present in [Het-s*] mycelium. Furthermore, overexpression of the het-s gene increases the frequency of the transition from [Het-s*] to [Het-s]. We propose that this transition is the consequence of a self-propagating conformational modification of the protein mediated by the formation of complexes between the two different forms of the polypeptide.

A single amino acid difference is sufficient to elicit vegetative incompatibility in the fungus Podospora anserina.

Vegetative incompatibility is known to limit heterokaryosis in filamentous fungi. It results from genetic differences between incompatible strains at specific loci. The proteins encoded by the two incompatible alleles het-s and het-S of the fungus Podospora anserina differ from each other by 14 amino acids. Two approaches have been used to identify how many and which of these differences are necessary to elicit incompatibility. Twelve alleles of the het-s locus of wild-type isolates of P. anserina and of the related species Podospora comata have been sequenced to determine the extent of the variability of genes controlling s and S specificities. Expression of hybrid het-s/het-S genes and site-specific mutagenesis revealed that the specificities of het-s and het-S are under the control of a limited number of amino acid differences. The results show that vegetative incompatibility between s and S strains can be attributed to a single amino acid difference in the proteins encoded by the het-s locus.

Two allelic genes responsible for vegetative incompatibility in the fungus Podospora anserina are not essential for cell viability.

Vegetative incompatibility is a lethal reaction that destroys the heterokaryotic cells formed by the fusion of hyphae of non-isogenic strains in many fungi. That incompatibility is genetically determined is well known but the function of the genes triggering this rapid cell death is not. The two allelic incompatibility genes, s and S, of the fungus Podospora anserina were characterized. Both encode 30 kDa polypeptides, which differ by 14 amino acids between the two genes. These two proteins are responsible for the incompatibility reaction that results when cells containing s and S genes fuse. Inactivation of the s or S gene by disruption suppresses incompatibility but does not affect the growth or the sexual cycle of the mutant strains. This suggests that these incompatibility genes have no essential function in the life cycle of the fungus.

repa, a repetitive and dispersed DNA sequence of the filamentous fungus Podospora anserina.

The sequences of homologous DNA regions of two wild-type strains of the fungus Podospora anserina, revealed in one strain the presence of a 349bp insertion leading to a RFLP. This DNA sequence is repeated in the genome and some of its locations are different in various wild-type strains. This DNA element exhibits structural similarities with the yeast solo delta, sigma or tau elements.