zur Suche springenzur Navigation springenzum Inhalt springen

Publikationen - Molekulare Signalverarbeitung

Sortieren nach: Erscheinungsjahr Typ der Publikation

Zeige Ergebnisse 1 bis 10 von 10.

Publikation

López-Carrasco, A.; Ballesteros, C.; Sentandreu, V.; Delgado, S.; Gago-Zachert, S.; Flores, R.; Sanjuán, R.; Different rates of spontaneous mutation of chloroplastic and nuclear viroids as determined by high-fidelity ultra-deep sequencing PLOS Pathog. 13, e1006547, (2017) DOI: 10.1371/journal.ppat.1006547

Mutation rates vary by orders of magnitude across biological systems, being higher for simpler genomes. The simplest known genomes correspond to viroids, subviral plant replicons constituted by circular non-coding RNAs of few hundred bases. Previous work has revealed an extremely high mutation rate for chrysanthemum chlorotic mottle viroid, a chloroplast-replicating viroid. However, whether this is a general feature of viroids remains unclear. Here, we have used high-fidelity ultra-deep sequencing to determine the mutation rate in a common host (eggplant) of two viroids, each representative of one family: the chloroplastic eggplant latent viroid (ELVd, Avsunviroidae) and the nuclear potato spindle tuber viroid (PSTVd, Pospiviroidae). This revealed higher mutation frequencies in ELVd than in PSTVd, as well as marked differences in the types of mutations produced. Rates of spontaneous mutation, quantified in vivo using the lethal mutation method, ranged from 1/1000 to 1/800 for ELVd and from 1/7000 to 1/3800 for PSTVd depending on sequencing run. These results suggest that extremely high mutability is a common feature of chloroplastic viroids, whereas the mutation rates of PSTVd and potentially other nuclear viroids appear significantly lower and closer to those of some RNA viruses.
Publikation

López-Carrasco, A.; Gago-Zachert, S.; Mileti, G.; Minoia, S.; Flores, R.; Delgado, S.; The transcription initiation sites of eggplant latent viroid strands map within distinct motifs in their in vivo RNA conformations RNA Biol. 13, 83-97, (2016) DOI: 10.1080/15476286.2015.1119365

Eggplant latent viroid (ELVd), like other members of family Avsunviroidae, replicates in plastids through a symmetric rolling-circle mechanism in which elongation of RNA strands is most likely catalyzed by a nuclear-encoded polymerase (NEP) translocated to plastids. Here we have addressed where NEP initiates transcription of viroid strands. Because this step is presumably directed by sequence/structural motifs, we have previously determined the conformation of the monomeric linear (+) and (−) RNAs of ELVd resulting from hammerhead-mediated self-cleavage. In silico predictions with 3 softwares led to similar bifurcated conformations for both ELVd strands. In vitro examination by non-denaturing PAGE showed that they migrate as prominent single bands, with the ELVd (+) RNA displaying a more compact conformation as revealed by its faster electrophoretic mobility. In vitro SHAPE analysis corroborated the ELVd conformations derived from thermodynamics-based predictions in silico. Moreover, sequence analysis of 94 full-length natural ELVd variants disclosed co-variations, and mutations converting canonical into wobble pairs or vice versa, which confirmed in vivo most of the stems predicted in silico and in vitro, and additionally helped to introduce minor structural refinements. Therefore, results from the 3 experimental approaches were essentially consistent among themselves. Application to RNA preparations from ELVd-infected tissue of RNA ligase-mediated rapid amplification of cDNA ends, combined with pretreatments to modify the 5′ ends of viroid strands, mapped the transcription initiation sites of ELVd (+) and (−) strands in vivo at different sequence/structural motifs, in contrast with the situation previously observed in 2 other members of the family Avsunviroidae.
Publikation

Fonseca, S.; Chini, A.; Hamberg, M.; Adie, B.; Porzel, A.; Kramell, R.; Miersch, O.; Wasternack, C.; Solano, R.; (+)-7-iso-Jasmonoyl-L-isoleucine is the endogenous bioactive jasmonate Nat. Chem. Biol. 5, 344-350, (2009) DOI: 10.1038/nchembio.161

Hormone-triggered activation of the jasmonate signaling pathway in Arabidopsis thaliana requires SCFCOI1-mediated proteasome degradation of JAZ repressors. (−)-JA-L-Ile is the proposed bioactive hormone, and SCFCOI1 is its likely receptor. We found that the biological activity of (−)-JA-L-Ile is unexpectedly low compared to coronatine and the synthetic isomer (+)-JA-L-Ile, which suggests that the stereochemical orientation of the cyclopentanone-ring side chains greatly affects receptor binding. Detailed GC-MS and HPLC analyses showed that the (−)-JA-L-Ile preparations currently used in ligand binding studies contain small amounts of the C7 epimer (+)-7-iso-JA-L-Ile. Purification of each of these molecules demonstrated that pure (−)-JA-L-Ile is inactive and that the active hormone is (+)-7-iso-JA-L-Ile, which is also structurally more similar to coronatine. In addition, we show that pH changes promote conversion of (+)-7-iso-JA-L-Ile to the inactive (−)-JA-L-Ile form, thus providing a simple mechanism that can regulate hormone activity through epimerization.
Publikation

BERGER, S.; Weichert, H.; Porzel, A.; Wasternack, C.; Kühn, H.; Feussner, I.; Enzymatic and non-enzymatic lipid peroxidation in leaf development BBA-Mol. Cell Biol. Lipids 1533, 266-276, (2001) DOI: 10.1016/S1388-1981(01)00161-5

Enzymatic and non-enzymatic lipid peroxidation has been implicated in programmed cell death, which is a major process of leaf senescence. To test this hypothesis we developed a high-performance liquid chromatography (HPLC) method for a simultaneous analysis of the major hydro(pero)xy polyenoic fatty acids. Quantities of lipid peroxidation products in leaves of different stages of development including natural senescence indicated a strong increase in the level of oxygenated polyenoic fatty acids (PUFAs) during the late stages of leaf senescence. Comprehensive structural elucidation of the oxygenation products by means of HPLC, gas chromatography/mass spectrometry and 1H nuclear magnetic resonance suggested a non-enzymatic origin. However, in some cases a small share of specifically oxidized PUFAs was identified suggesting involvement of lipid peroxidizing enzymes. To inspect the possible role of enzymatic lipid peroxidation in leaf senescence, we analyzed the abundance of lipoxygenases (LOXs) in rosette leaves of Arabidopsis. LOXs and their product (9Z,11E,13S,15Z)-13-hydroperoxy-9,11,15-octadecatrienoic acid were exclusively detected in young green leaves. In contrast, in senescing leaves the specific LOX products were overlaid by large amounts of stereo-random lipid peroxidation products originating from non-enzymatic oxidation. These data indicate a limited contribution of LOXs to total lipid peroxidation, and a dominant role of non-enzymatic lipid peroxidation in late stages of leaf development.
Publikation

Miersch, O.; Porzel, A.; Wasternack, C.; Microbial conversion of jasmonates - hydroxylations by Aspergillus niger Phytochemistry 50, 1147-1152, (1999) DOI: 10.1016/S0031-9422(98)00698-0

Aspergillus niger is able to hydroxylate the pentenyl side chain of (−)-jasmonic acid (JA) leading to (11S)- (−)-hydroxy-JA/ (11R)- (−)-hydroxy-JA (2:1) and (−)-11,12-didehydro-JA. Methyl (−)-jasmonate (JA-Me) is converted upon hydrolysis. During prolonged cultivation or at non-optimized isolation procedures, the 11-hydroxy- (9Z)-pentenyl side chain may isomerize to (10E)-9-hydroxy- and (9E)-11-hydroxy-compounds by allylic rearrangement. The fungus hydroxylates (±)-9,10-dihydro-JA at position C-11 into 11j-hydroxy-9,10-dihydro-JA. As JA-Me, the methyl dihydro-JA is hydroxylated only upon hydrolysis into the free acid.
Publikation

Kramell, R.; Porzel, A.; Miersch, O.; Schneider, G.; Wasternack, C.; Chromatographic resolution of peptide-like conjugates of jasmonic acid and of cucurbic acid isomers J. Chromatogr. A 847, 103-107, (1999) DOI: 10.1016/S0021-9673(99)00335-0

The chiral separation of peptide-like conjugates of jasmonic acid and of cucurbic acid isomers was investigated by liquid chromatography on Chiralpak AS and Nucleodex β-PM. The retention sequences reflect distinct chromatographic properties with respect to the chirality of the jasmonic acid part or of the cucurbic acid isomers. The chromatographic behaviour of the amide conjugates on a reversed-phase C18 column provides evidence for the resolution of diastereomeric conjugates depending on the chirality of both constituents of the conjugate molecule. The chromatographic procedures are suitable for the analytical and preparative separation of such conjugates.
Bücher und Buchkapitel

Kramell, R.; Porzel, A.; Miersch, O.; Schneider, G.; Characterization of Isoleucine Conjugates of Cucurbic Acid Isomers by Reversed-Phase and Chiral High-Performance Liquid Chromatography 77-78, (1998)

0
Bücher und Buchkapitel

Feussner, I.; Balkenhohl, T. J.; Porzel, A.; Kühn, H.; Wasternack, C.; Structural Elucidation of Oxygenated Triacylglycerols in Cucumber and Sunflower Cotyledons 57-58, (1998)

0
Publikation

Feussner, I.; Balkenhohl, T. J.; Porzel, A.; Kühn, H.; Wasternack, C.; Structural Elucidation of Oxygenated Storage Lipids in Cucumber Cotyledons J. Biol. Chem. 272, 21635-21641, (1997) DOI: 10.1074/jbc.272.34.21635

At early stages of germination, a special lipoxygenase is expressed in cotyledons of cucumber and several other plants. This enzyme is localized at the lipid storage organelles and oxygenates their storage triacylglycerols. We have isolated this lipid body lipoxygenase from cucumber seedlings and found that it is capable of oxygenating in vitro di- and trilinolein to the corresponding mono-, di-, and trihydroperoxy derivatives. To investigate the in vivo activity of this enzyme during germination, lipid bodies were isolated from cucumber seedlings at different stages of germination, and the triacylglycerols were analyzed for oxygenated derivatives by a combination of high pressure liquid chromatography, gas chromatography/mass spectrometry, and nuclear magnetic resonance spectroscopy. We identified as major oxygenation products triacylglycerols that contained one, two, or three 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid residues. During germination, the amount of oxygenated lipids increased strongly, reaching a maximum after 72 h and declining afterward. The highly specific pattern of hydroperoxy lipids formed suggested the involvement of the lipid body lipoxygenase in their biosynthesis.These data suggest that this lipoxygenase may play an important role during the germination process of cucumber and other plants and support our previous hypothesis that the specific oxygenation of the storage lipids may initiate their mobilization as a carbon and energy source for the growing seedling.
Publikation

Feussner, I.; Porzel, A.; Wasternack, C.; Kühn, H.; Quantitative Analyse von Lipoxygenase-Metaboliten in Lipiden durch NMR-Spektroskopie BIOspektrum 3, 54-58, (1997)

0
  • Die Leibniz-Gemeinschaft
  • Digitalisierung in der Landwirtschaft
  • Martin-Luther Universität Halle
IPB Mainnav Search