J P Roustan

École Nationale Supérieure Agronomique de Toulouse, Tolosa de Llenguadoc, Midi-Pyrénées, France

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Publications (13)73.37 Total impact

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    ABSTRACT: Eutypine, 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzyl aldehyde, is a toxin produced byEutypa lata,the causal agent of eutypa dieback of grapevines. It has previously been demonstrated that tolerance of some cultivars to this disease was correlated with their capacity to convert eutypine to the corresponding alcohol, eutypinol, which lacks phytotoxicity. We have thus purified to homogeneity a protein fromVigna radiatathat exhibited eutypine-reducing activity and have isolated the corresponding cDNA. This encodes an NADPH-dependent reductase of 36 kDa that we have namedVigna radiataeutypine-reducing enzyme (VR-ERE), based on the capacity of a recombinant form of the protein to reduce eutypine into eutypinol. The strongest homologies (86.8%) of VR-ERE at the amino acid level were found with CPRD14, a drought-inducible gene of unknown function, isolated fromVigna unguiculataand with an aromatic alcohol dehydrogenase (71.7%) fromEucalyptus gunnii. Biochemical characterization of VR-ERE revealed that a variety of compounds containing an aldehyde group can act as substrates. However, the highest affinity was observed with 3-substituted benzaldehydes. Expression of aVR-EREtransgene inVitis viniferacells culturedin vitroconferred resistance to the toxin. This discovery opens up new biotechnological approaches for the generation of grapevines resistant to eutypa dieback.
    The Plant Journal 10/2008; 16(3):335 - 343. · 6.58 Impact Factor
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    ABSTRACT: Eutypine (4-hydroxy-3-[3-methyl-3-butene-1-ynyl] benzaldehyde) is a toxin produced by Eutypa lata, the causal agent of eutypa dieback in the grapevine (Vitis vinifera). Eutypine is enzymatically converted by numerous plant tissues into eutypinol (4-hydroxy-3-[3-methyl-3-butene-1-ynyl] benzyl alcohol), a metabolite that is nontoxic to grapevine. We report a four-step procedure for the purification to apparent electrophoretic homogeneity of a eutypine-reducing enzyme (ERE) from etiolated mung bean (Vigna radiata) hypocotyls. The purified protein is a monomer of 36 kD, uses NADPH as a cofactor, and exhibits a Km value of 6.3 &mgr;M for eutypine and a high affinity for 3- and 4-nitro-benzaldehyde. The enzyme failed to catalyze the reverse reaction using eutypinol as a substrate. ERE detoxifies eutypine efficiently over a pH range from 6.2 to 7.5. These data strongly suggest that ERE is an aldehyde reductase that could probably be classified into the aldo-keto reductase superfamily. We discuss the possible role of this enzyme in eutypine detoxification.
    Plant physiology 02/1999; 119(2):621-6. · 6.56 Impact Factor
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    ABSTRACT: Eutypine, 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzaldehyde, is a toxin produced by Eutypa lata (Pers.: Fr.) Tul., the causal agent of dying arm disease of Vitis vinifera L. (grapevine). Previously, we have shown that eutypine is involved in the development of disease symptoms. In the present study, the effects of V. vinifera cell-suspension cultures on the biological activity of the toxin were investigated. Eutypine was converted by grapevine tissues into a single compound, identified by mass spectrometry and nuclear magnetic resonance as 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzyl alcohol, designated eutypinol. This compound was found to be non-toxic for grapevine tissues. Unlike eutypine, eutypinol failed to affect the oxidation rate or membrane potential of isolated mitochondria. In grapevine cells, reduction of eutypine into the corresponding alcohol is an NADPH-dependent enzymatic reaction. An enzyme which reduced eutypine was partially purified, over 1000-fold, using a five-step purification procedure. By gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the protein was found to have a molecular mass of 54–56 kDa. The enzyme exhibited an apparent K m for eutypine of 44 μM, and was active between pH 6.8 and 7.5 with a maximum at pH 7.0. The eutypine reductase activity was improved by Mn2+ and Mg2+ and inhibited by disulfiram and p-hydroxymercuribenzoate. The possible role of the eutypine-detoxification mechanism in the defense reactions of V. vinifera cells is discussed.
    Planta 01/1999; 207(4):544-550. · 3.38 Impact Factor
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    ABSTRACT: Eutypine, 4-hydroxy-3-(3-methyl-3-butene-1-ynyl) benzyl aldehyde, is a toxin produced by Eutypa lata, the causal agent of eutypa dieback of grapevines. It has previously been demonstrated that tolerance of some cultivars to this disease was correlated with their capacity to convert eutypine to the corresponding alcohol, eutypinol, which lacks phytotoxicity. We have thus purified to homogeneity a protein from Vigna radiata that exhibited eutypine-reducing activity and have isolated the corresponding cDNA. This encodes an NADPH-dependent reductase of 36 kDa that we have named Vigna radiata eutypine-reducing enzyme (VR-ERE), based on the capacity of a recombinant form of the protein to reduce eutypine into eutypinol. The strongest homologies (86.8%) of VR-ERE at the amino acid level were found with CPRD14, a drought-inducible gene of unknown function, isolated from Vigna unguiculata and with an aromatic alcohol dehydrogenase (71.7%) from Eucalyptus gunnii. Biochemical characterization of VR-ERE revealed that a variety of compounds containing an aldehyde group can act as substrates. However, the highest affinity was observed with 3-substituted benzaldehydes. Expression of a VR-ERE transgene in Vitis vinifera cells cultured in vitro conferred resistance to the toxin. This discovery opens up new biotechnological approaches for the generation of grapevines resistant to eutypa dieback.
    The Plant Journal 12/1998; 16(3):335-43. · 6.58 Impact Factor
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    ABSTRACT: The role of ethylene in shoot regeneration was investigated using transgenic Cucumis melo plants expressing an antisense 1-aminocyclopropane-1-carboxylate (ACC) oxidase gene. ACC oxidase catalyses the last step of ethylene biosynthesis. Leaf and cotyledon explants from the transgenic plants exhibited low ACC oxidase activity and ethylene production, whereas the regeneration capacity of the tissues was greatly enhanced (3.5- and 2.8-fold, respectively) compared to untransformed control tissues. Addition of ethylene released by 50 or 100 µm 2-chloroethylphosphonic acid dramatically reduced the shoot regeneration rate of the transgenic tissues. The results clearly demonstrate that ethylene plays an important role in C. melo morphogenesis in vitro.
    Plant Cell Reports 03/1998; 17(6):586-589. · 2.94 Impact Factor
  • Comptes Rendus de l Académie des Sciences - Series III - Sciences de la Vie 01/1997; 320(2):149-158.
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    ABSTRACT: Eutypine is a toxin produced by Eutypa lata, the causal agent of the dying-arm disease of grapevine. We have previously shown that this toxin behaves as a lipophylic weak acid (pK = 6.2) and induces drastic changes in both the respiration and energy balance of grapevine cells. In the present study, the molecular mode of action of eutypine at the mitochondrial level, using methyl-eutypine, the unprotonable derivative of the toxin, has been investigated. The effects of these molecules on mitochondrial respiration and membrane potential were compared using isolated mitochondria from grapevine cells in suspension cultures or potato tuber mitochondria. Eutypine induces marked stimulation of oxygen consumption and a depolarizing effect, while methyl-eutypine exhibits a very small effect on both the rate of oxygen uptake and membrane potential. For high eutypine concentrations, a mixed effect corresponding to a direct inhibition of electron transport and uncoupling can be observed. In addition, below 200 microM, eutypine displays a linear relationship between oxidation rate and membrane potential similar to that of the classical protonophore carbonyl cyanide-m-chlorophenylhydrazone (CCCP). However, unlike CCCP, eutypine induces a potential-dependent proton conductance that can be due to the potential-dependent migration of the dissociated form of the toxin across the membrane. It is concluded that eutypine uncouples mitochondrial oxidative phosphorylation and decreases the ADP/O ratio in grapevine cells by increasing the proton leaks via a cyclic protonophore mechanism. The physiological aspects of these results are discussed.
    Archives of Biochemistry and Biophysics 11/1996; 334(2):200-5. · 3.37 Impact Factor
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    ABSTRACT: The plant hormone ethylene plays a major role in the ripening of climacteric fruit. We have generated transgenic cantaloupe Charentais melons expressing an antisense ACC oxidase gene; ACC oxidase catalyzes the last step of ethylene biosynthesis. Ethylene production of transgenic fruit was < 1% of control untransformed fruit, and the ripening process was blocked both on and off the vine. The antisense phenotype could be reversed by exogenous ethylene treatment. Analysis of antisense ACC oxidase melons indicated that the ripening process includes ethylene-dependent and ethylene-independent pathways. Because the transgenic line we generated displays extended storage life and improved quality, it has a promising potential for commercial development.
    Nature Biotechnology 08/1996; 14(7):862-6. · 32.44 Impact Factor
  • A Sarrafi, J P Roustan, J Fallot, G Alibert
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    ABSTRACT: Crosses were made between five cytoplasmic male-sterile and five restorer sunflower inbred lines. Twenty-five F1 hybrids and their parents were studied for their organogenesis ability in a randomized block design with four replications. Each replication per genotype consisted of ten petri dishes with four expiants. Regeneration medium consisted of full MS medium modified by the addition of hormones and solidified with 6 g/l agar. Statistical analysis showed that both general and specific combining abilities were significant for all of the organogenesis parameters studied, and both showed several significant positive or negative values. General combining ability values were usually higher than those of specific combining ability, indicating the importance of additive genetic control for organogenesis parameters in sunflower. Narrow-sense heritability for the number of explants producing shoots and roots was 65.8%, which suggests that organogenesis of currently inferior inbred lines in sunflower should be improved in a crossing program.
    Theoretical and Applied Genetics 02/1996; 92(2):225-9. · 3.66 Impact Factor
  • Cytologia 01/1994; · 0.31 Impact Factor
  • K M B, A Latche, J P Roustan, J Fallot
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    ABSTRACT: The effects of CoCl2, AgNO3 and ethylene released by exogenous 2-chloroethylphosphonic acid (Ethephon), were studied on shoot regeneration from cotyledons of Helianthus annuus cv. E8206R, a poorly regenerative cultivar. Inhibition of ethylene biosynthesis by CoCl2, at concentrations of 20 μK, provoked a substantial enhancement of shoot regeneration (30 %): the control was poorly regenerative. However, CoCl2 had no effect when Ethephon was supplied. Inhibition of ethylene action by AgNO3, at concentrations of 10-25 μM, caused a significant increase in plant regeneration: 25 % instead of 1.2 % in the control. Furthermore, addition of Ethephon to AgNO3-treated tissues failed to reduce the stimulation of shoot regeneration caused by AgNO3. On the basis of these findings, it is suggested that ethylene inhibits the regeneration process from cotyledons of sunflower.
    Plant Cell Reports 07/1991; 10(4):204-7. · 2.94 Impact Factor
  • J P Roustan, A Latche, J Fallot
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    ABSTRACT: The effects of Co(2+) and Ni(2+) on ethylene production and somatic embryogenesis by carrot (Daucus carota L.) cell cultures were studied. At concentrations of 10 μM to 50 μM, CoCl2 effectively inhibited ethylene production by embryogenic cultures and significantly stimulated somatic embryogenesis. The observed increase of embryo number was proportional to the inhibition level of ethylene production. However, CoCl2 had no effect when Ethephon was supplied. Nickel also reduced ethylene production, but to a slightly lesser extent than CoCl2, bringing about a lower increase in the number of somatic embryos. The role of ethylene on somatic embryogenesis is discussed.
    Plant Cell Reports 03/1989; 8(3):182-185. · 2.94 Impact Factor
  • J.-P. Roustan, A. Latche, J. Fallot
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    ABSTRACT: The effects of salicylic acid (SA) and other phenolic compounds, acetylsalicylic acid (ASA), benzoic add (BA) and sulfosalicylic acid (SSA), on ethylene production and somatic embryogenesis by carrot (Daucus carota L.) cell cultures were studied. SA and ASA, at concentrations of 10 μM and 100 μM, significantly stimulated somatic embryogenesis and effectively inhibited ethylene production by carrot cell suspension cultures. The observed increase of embryo number was proportional to the inhibition rate of ethylene production. However, BA and SSA affected neither ethylene production nor somatic embryogenesis. The role of SA in somatic embryogenesis is discussed.
    Biologia Plantarum 32(4):273-276. · 1.69 Impact Factor