Mitochondrial respiratory pathways modulate nitrate sensing and nitrogen-dependent regulation of plant architecture in Nicotiana sylvestris

Mitochondrial electron transport pathways exert effects on carbon–nitrogen (C/N) relationships. To examine whether mitochondria–N interactions also influence plant growth and development, we explored the responses of roots and shoots to external N supply in wild-type (WT) Nicotiana sylvestris and the cytoplasmic male sterile II (CMSII) mutant, which has a N-rich phenotype. Root architecture in N. sylvestris seedlings showed classic responses to nitrate and sucrose availability. In contrast, CMSII showed an altered ‘nitrate-sensing’ phenotype with decreased sensitivity to C and N metabolites. The WT growth phenotype was restored in CMSII seedling roots by high nitrate plus sugars and in shoots by gibberellic acid (GA). Genome-wide cDNA-amplified fragment length polymorphism (AFLP) analysis of leaves from mature plants revealed that only a small subset of transcripts was altered in CMSII. Tissue abscisic acid content was similar in CMSII and WT roots and shoots, and growth responses to zeatin were comparable. However, the abundance of key transcripts associated with GA synthesis was modified both by the availability of N and by the CMSII mutation. The CMSII mutant maintained a much higher shoot/root ratio at low N than WT, whereas no difference was observed at high N. Shoot/root ratios were strikingly correlated with root amines/nitrate ratios, values of <1 being characteristic of high N status. We propose a model in which the amine/nitrate ratio interacts with GA signalling and respiratory pathways to regulate the partitioning of biomass between shoots and roots

Phytosphingosine-phosphate is a signal for AtMPK6 activation and Arabidopsis response to chilling

Long-chain bases (LCBs) are pleiotropic sphingolipidic signals in eukaryotes. We investigated the source and function of phytosphingosine-1-phosphate (PHS-P), a phospho-LCB rapidly and transiently formed in Arabidopsis thaliana on chilling.PHS-P was analysed by thin-layer chromatography following in vivo metabolic radiolabelling. Pharmacological and genetic approaches were used to identify the sphingosine kinase isoforms involved in cold-responsive PHS-P synthesis. Gene expression, mitogen-activated protein kinase activation and growth phenotypes of three LCB kinase mutants (lcbk1, sphk1 and lcbk2) were studied following cold exposure. Chilling provoked the rapid and transient formation of PHS-P in Arabidopsis cultured cells and plantlets. Cold-evoked PHS-P synthesis was reduced by LCB kinase inhibitors and abolished in the LCB kinase lcbk2 mutant, but not in lcbk1 and sphk1 mutants. lcbk2 presented a constitutive AtMPK6 activation at 22°C. AtMPK6 activation was also triggered by PHS-P treatment independently of PHS/PHS-P balance. lcbk2 mutants grew comparably with wild-type plants at 22 and 4°C, but exhibited a higher root growth at 12°C, correlated with an altered expression of the cold-responsive DELLA gene RGL3. Together, our data indicate a function for LCBK2 in planta. Furthermore, they connect PHS-P formation with plant response to cold, expanding the field of LCB signalling in plants

Répercussions d'un défaut mitochondrial sur les interactions subcellulaires dans le métabolisme foliaire (analyse d'un mutant respiratoire de Nicotiana Sylvestris)

LE DIALOGUE EXISTANT ENTRE LES DIFFERENTS COMPARTIMENTS SUBCELLULAIRES EST BASE SUR DES ECHANGES MASSIFS DE METABOLITES, D'ATP ET DE POUVOIR REDUCTEUR. LA CHAINE DE TRANSPORT D'ELECTRONS MITOCHONDRIALE JOUE UN ROLE IMPORTANT DANS CETTE COORDINATION INTER-COMPARTIMENTALE. CE TRAVAIL PRESENTE UNE ETUDE DE L'IMPLICATION DU PROCESSUS MITOCHONDRIAL DANS LES GRANDES FONCTIONS FOLIAIRES. LE MUTANT DE NICOTIANA SYLVESTRIS, CMSII (CYTOPLASMIC MALE-STERILE II), EST L'UN DES RARES MUTANTS DISPONIBLES AFFECTES DANS LE FONCTIONNEMENT D'UN COMPLEXE DE LA CHAINE RESPIRATOIRE (PERTE DU COMPLEXE I) SUITE A LA DELETION DU GENE MITOCHONDRIAL NAD7. CE MUTANT EST DONC UN EXCELLENT SYSTEME POUR DETERMINER LES CONSEQUENCES DE LA PERTE D'UNE DESHYDROGENASE MITOCHONDRIALE MAJEURE, AU NIVEAU DES PROCESSUS FOLIAIRES. IL EST MONTRE QUE LE DYSFONCTIONNEMENT DU COMPLEXE I INDUIT UN AJUSTEMENT DES SYSTEMES ANTIOXYDANTS, METTANT EN EVIDENCE UNE SIGNALISATION MITOCHONDRIE-NOYAU IMPLIQUEE DANS LA DETERMINATION DE LA RESISTANCE AU STRESS. DE PLUS, LE COMPLEXE I EST IMPORTANT DANS L'OPTIMISATION DE L'ASSIMILATION DU CO2 LORSQUE LA PHOTORESPIRATION EST ACTIVE, ET PARTICULIEREMENT PENDANT LA PERIODE D'INDUCTION QUI SUIT UNE TRANSITION OBSCURITE-LUMIERE. NOS DONNEES SUGGERENT QUE CE ROLE EST LIE A UNE OPTIMISATION DES ECHANGES REDOX ENTRE CHLOROPLASTE ET MITOCHONDRIE. EN PLUS DE CET EFFET SUR LES FLUX PHOTOSYNTHETIQUES, L'ABSENCE DU COMPLEXE I A DE FORTES REPERCUSSIONS SUR L'ASSIMILATION DE L'AZOTE. UNE DIMINUTION DANS LA DISPONIBILITE EN 2-OXOGLUTARATE, SQUELETTE CARBONE REQUIS POUR L'ASSIMILATION DE L'AZOTE, EST ASSOCIEE A UNE MODIFICATION DE LA BALANCE C/N, NOTAMMENT UNE ACCUMULATION DES ACIDES AMINES RICHES EN AZOTE. L'ENSEMBLE DES RESULTATS MONTRE QUE L'ETAT REDOX CELLULAIRE EST INFLUENCE PAR LA CHAINE DE TRANSPORT D'ELECTRONS MITOCHONDRIALE, ET SUGGERE QUE L'ETAT REDOX POURRAIT ETRE UN ACTEUR CLE DANS LE CONTROLE DES INTERACTIONS C/N.PLANT DEVELOPMENT AND FUNCTION REQUIRES DIALOGUE BETWEEN DIFFERENT SUBCELLULLAR COMPARTMENTS. INVOLVING EXCHANGE OF METABOLITES, NUCLEOTIDES AND REDUCING POWER. THE MITOCHONDRIAL ELECTRON TRANSPORT CHAIN PLAYS AN IMPORTANT PART IN THIS INTERCOMPARTMENTAL CO-ORDINATION. THIS WORK PRESENTS A STUDY OF THE INTEGRATION OF MITOCHONDRIAL PROCESSES IN WHOLE LEAF PHYSIOLOGY AND MET ABOLISM. THE NICOTIANA SYLVESTRIS MUTANT, CMSII (CYTOPLASMIC MALE-STERILE II). IS ONE OF THE FEW AVAILABLE MUTANTS WITH A DYSFUNCTIONAL RESPIRATORY COMPLEX (LOSS OF COMPLEX I ACTIVITY) DUE TO A DELETION IN THE MITOCHONDRIAL GENE, NAD7. THIS MUTANT REPRESENTS, THEREFORE, AN EXCELLENT SYSTEM IN WHICH TO DISSECT THE CONSEQUENCES OF LOSS OF A MAJOR MITOCHONDRIAL DEHYDROGENASE FOR WHOLE LEAF PROCESSES. IT IS SHOWN THAT COMPLEX I DYSFUNCTION CAUSES ADJUSTMENT OF LEAF ANTIOXIDANT SYSTEMS. IMPLICA TING MITOCHONDRIA TO NUCLEUS SIGNALLING IN DETERMINATION OF STRESS RESISTANCE. SECOND, FLUX ANALYSIS OF PHOTOSYNTHESIS DEMONSTRATES THE IMPORTANCE OF COMPLEX I FUNCTION IN OPTIMISING CARBON ASSIMILATION WHEN PHOTORESPIRATION IS ACTIVE AND, PARTICULARLY, DURING THE INDUCTION PERIOD THAT FOLLOWS A DARK/LIGHT TRANSITION. EVIDENCE IS PRESENTED THAT THIS ROLE IS LINKED TO AN OPTIMISATION OF CHLOROPLAST-MITOCHONDRIA REDUCTANT EXCHANGE. IN ADDITION TO THIS EFFECT ON OVERALL PHOTOSYNTHETIC FLUXES, THE ABSENCE OF COMPLEX I HAS DRAMATIC REPERCUSSIONS FOR THE INTEGRATION OF NITROGEN ASSIMILATION WITH CO2 FIXATION. A SUBSTANTIAL MODIFICATION OF LEAF ORGANIC ACID PROFILES INVOLVES DECREASED AVAILABILITY OF 2-OXOGLUTARATE. THE CARBON SKELETON FOR AMMONIA ASSIMILATION. THIS OBSERVATION IS LINKED TO A MARKED CHANGE OF LEAF C/N BALANCE, EVIDENCED BY THE ACCUMULATION OF AMINO ACIDS RICH IN NITROGEN. TAKEN TOGETHER, THE RESULTS INDICATE THAT CELLULAR REDOX STATE IS GREATLY INFLUENCED BY MITOCHONDRIAL ELECTRON TRANSPORT STATUS. AND LEAD TO THE CONCLUSION THAT REDOX STATE MAY BE A KEY PLAYER IN CONTROLLING C/N RELATIONSHIPS.ORSAY-PARIS 11-BU Sciences (914712101) / SudocSudocFranceF

The Arabidopsis Putative Selenium-Binding Protein Family: Expression Study and Characterization of SBP1 as a Potential New Player in Cadmium Detoxification Processes

International audienceIn Arabidopsis thaliana, the putative selenium-binding protein gene family is made of 3 members (SBP 1 to 3). RT-PCR analyses showed that SBP1 expression was ubiquitous. SBP2 was expressed at a lower level in flowers and roots whereas SBP3 transcripts were only detected in young seedling tissues. In Cd-treated seedlings, SBP1 level of expression was rapidly increased in roots. In shoots, SBP1 transcripts accumulated later and for higher Cd doses. SBP2 and SBP3 expression showed delayed or no responsiveness to Cd. In addition, luciferase (LUC) activity recorded on Arabidopsis lines expressing the LUC gene under the control of SBP1 promoter further showed dynamic regulation of SBP1 expression during development and in response to Cd stress. Western blot analysis using polyclonal antibodies raised against SBP1 showed that SBP1 protein accumulated in Cd-exposed tissues in correlation with SBP1 transcript amount. The sbp1 null mutant displayed no visible phenotype under normal and stress conditions that was explained by the up regulation of SBP2 expression. SBP1 over-expression enhanced Cd accumulation in roots and reduced sensitivity to Cd in WT and more significantly in Cd-hypersensitive cad mutants that lack phytochelatins. Similarly in Saccharomyces cerevisiae, SBP1 expression led to increased Cd tolerance of the Cd-hypersensitive ycf1 mutant. In vitro experiments showed that SBP1 has the ability to bind Cd. These data highlight the importance of maintaining the adequate SBP protein level under healthy and stress conditions and suggest that during Cd stress, SBP1 accumulation efficiently helps to detoxify Cd potentially through direct binding

The Arabidopsis protein farnesylation era1 mutant displays an altered hormonal-dependent nitrate regulation of root architecture

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Tagging and Capture of Prenylated CaaX-Proteins from Plant Cell Cultures

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Evidence for ACD5 ceramide kinase activity involvement in Arabidopsis response to cold stress

International audienceAlthough sphingolipids emerged as important signals for plant response to low temperature, investigations have been limited so far to the function of long-chain base intermediates. The formation and function of ceramide phosphates (Cer-Ps) in chilled Arabidopsis were explored. Cer-Ps were analysed by thin layer chromatography (TLC) following in vivo metabolic radiolabelling. Ceramide kinase activity, gene expression and growth phenotype were determined in unstressed and cold-stressed wild type (WT) and Arabidopsis ceramide kinase mutant acd5. A rapid and transient formation of Cer-P occurs in cold-stressed WT Arabidopsis plantlets and cultured cells, which is strongly impaired in acd5 mutant. Although concomitant, Cer-P formation is independent of long-chain base phosphate (LCB-P) formation. No variation of ceramide kinase activity was measured in vitro in WT plantlets upon cold stress but the activity in acd5 mutant was further reduced by cold stress. At the seedling stage, acd5 response to cold was similar to that of WT. Nevertheless, acd5 seed germination was hypersensitive to cold and abscisic acid (ABA), and ABA-dependent gene expression was modified in acd5 seeds when germinated at low temperature. Our data involve for the first time Cer-P and ACD5 in low temperature response and further underline the complexity of sphingolipid signalling operating during cold stress

Arabidopsis Putative Selenium-Binding Protein1 Expression Is Tightly Linked to Cellular Sulfur Demand and Can Reduce Sensitivity to Stresses Requiring Glutathione for Tolerance1[W]

Selenium-Binding Protein1 (SBP1) gene expression was studied in Arabidopsis (Arabidopsis thaliana) seedlings challenged with several stresses, including cadmium (Cd), selenium {selenate [Se(VI)] and selenite [Se(IV)]}, copper (Cu), zinc (Zn), and hydrogen peroxide (H2O2) using transgenic lines expressing the luciferase (LUC) reporter gene under the control of the SBP1 promoter. In roots and shoots of SBP1∷LUC lines, LUC activity increased in response to Cd, Se(VI), Cu, and H2O2 but not in response to Se(IV) or Zn. The pattern of expression of SBP1 was similar to that of PRH43, which encodes the 5′-Adenylylphosphosulfate Reductase2, a marker for the induction of the sulfur assimilation pathway, suggesting that an enhanced sulfur demand triggers SBP1 up-regulation. Correlated to these results, SBP1 promoter showed enhanced activity in response to sulfur starvation. The sulfur starvation induction of SBP1 was abolished by feeding the plants with glutathione (GSH) and was enhanced when seedlings were treated simultaneously with buthionine sulfoxide, which inhibits GSH synthesis, indicating that GSH level participates in the regulation of SBP1 expression. Changes in total GSH level were observed in seedlings challenged with Cd, Se(VI), and H2O2. Accordingly, cad2-1 seedlings, affected in GSH synthesis, were more sensitive than wild-type plants to these three stresses. Moreover, wild-type and cad2-1 seedlings overexpressing SBP1 showed a significant enhanced tolerance to Se(VI) and H2O2 in addition to the previously described resistance to Cd, highlighting that SBP1 expression decreases sensitivity to stress requiring GSH for tolerance. These results are discussed with regard to the potential regulation and function of SBP1 in plants

The Arabidopsis DUF239 gene family encodes Neprosin‐like proteins that are widely expressed in seed endosperm

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Functional Mitochondrial Complex I Is Required by Tobacco Leaves for Optimal Photosynthetic Performance in Photorespiratory Conditions and during Transients

The importance of the mitochondrial electron transport chain in photosynthesis was studied using the tobacco (Nicotiana sylvestris) mutant CMSII, which lacks functional complex I. Rubisco activities and oxygen evolution at saturating CO(2) showed that photosynthetic capacity in the mutant was at least as high as in wild-type (WT) leaves. Despite this, steady-state photosynthesis in the mutant was reduced by 20% to 30% at atmospheric CO(2) levels. The inhibition of photosynthesis was alleviated by high CO(2) or low O(2). The mutant showed a prolonged induction of photosynthesis, which was exacerbated in conditions favoring photorespiration and which was accompanied by increased extractable NADP-malate dehydrogenase activity. Feeding experiments with leaf discs demonstrated that CMSII had a lower capacity than the WT for glycine (Gly) oxidation in the dark. Analysis of the postillumination burst in CO(2) evolution showed that this was not because of insufficient Gly decarboxylase capacity. Despite the lower rate of Gly metabolism in CMSII leaves in the dark, the Gly to Ser ratio in the light displayed a similar dependence on photosynthesis to the WT. It is concluded that: (a) Mitochondrial complex I is required for optimal photosynthetic performance, despite the operation of alternative dehydrogenases in CMSII; and (b) complex I is necessary to avoid redox disruption of photosynthesis in conditions where leaf mitochondria must oxidize both respiratory and photorespiratory substrates simultaneously