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Publikationen - Molekulare Signalverarbeitung

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Preprints

Mik, V.; Poslíšil, T.; Brunoni, F.; Grúz, J.; Nožková, V.; Wasternack, C.; Miersch, O.; Strnad, M.; Floková, K.; Novák, O.; Široká, J.; Synthetic and analytical routes to the L-amino acid conjugates of cis-OPDA and their identification and quantification in plants ChemRxiv (2023) DOI: 10.26434/chemrxiv-2023-qlzj4

Cis-(+)-12-oxophytodienoic acid (cis-(+)-OPDA) is a bioactive jasmonate, a precursor of jasmonic acid, which also displays signaling activity on its own. Modulation of cis-(+)-OPDA actions may be carried out via biotransformation leading to metabolites of various functions, similar to other phytohormones. This work introduces a methodology for the synthesis of racemic cis-OPDA conjugates with amino acids (OPDA-aa) and their deuterium-labeled analogs, which enables the identification and accurate quantification of these compounds in plants. We have developed a highly sensitive liquid chromatography-tandem mass spectrometry-based method for the reliable determination of seven OPDA-aa (OPDA-Alanine, OPDA-Aspartate, OPDA-Glutamate, OPDA-Glycine, OPDA-Isoleucine, OPDA-Phenylalanine, and OPDA-Valine) from minute amount of plant material. The extraction from 10 mg of fresh plant tissue by 10% aqueous methanol followed by single-step sample clean-up on hydrophilic–lipophilic balanced columns prior to final analysis was optimized. The method was validated in terms of accuracy and precision, and the method parameters such as process efficiency, recovery and matrix effects were evaluated. In mechanically wounded 30-day-old Arabidopsis thaliana leaves, five endogenous (+)-OPDA-aa were identified and their endogenous levels reached a maximum of pmol/g. The time-course accumulation revealed a peak 60 min after the wounding, roughly corresponding to the accumulation of cis-(+)-OPDA. Current synthetic and analytical methodologies support studies on cis-(+)-OPDA conjugation with amino acids and research into the biological significance of these metabolites in plants.
Publikation

Mik, V.; Pospíšil, T.; Brunoni, F.; Grúz, J.; Nožková, V.; Wasternack, C.; Miersch, O.; Strnad, M.; Floková, K.; Novák, O.; Široká, J.; Synthetic and analytical routes to the L-amino acid conjugates of cis-OPDA and their identification and quantification in plants Phytochemistry 215, 113855, (2023) DOI: 10.1016/j.phytochem.2023.113855

Cis-(+)-12-oxophytodienoic acid (cis-(+)-OPDA) is a bioactive jasmonate, a precursor of jasmonic acid, which also displays signaling activity on its own. Modulation of cis-(+)-OPDA actions may be carried out via biotransformation leading to metabolites of various functions. This work introduces a methodology for the synthesis of racemic cis-OPDA conjugates with amino acids (OPDA-aa) and their deuterium-labeled analogs, which enables the unambiguous identification and accurate quantification of these compounds in plants. We have developed a highly sensitive liquid chromatography-tandem mass spectrometry-based method for the reliable determination of seven OPDA-aa (OPDA-Alanine, OPDA-Aspartate, OPDA-Glutamate, OPDA-Glycine, OPDA-Isoleucine, OPDA-Phenylalanine, and OPDA-Valine) from minute amount of plant material. The extraction from 10 mg of fresh plant tissue by 10% aqueous methanol followed by single-step sample clean-up on hydrophilic–lipophilic balanced columns prior to final analysis was optimized. The method was validated in terms of accuracy and precision, and the method parameters such as process efficiency, recovery and matrix effects were evaluated. In mechanically wounded 30-day-old Arabidopsis thaliana leaves, five endogenous (+)-OPDA-aa were identified and their endogenous levels were estimated. The time-course accumulation revealed a peak 60 min after the wounding, roughly corresponding to the accumulation of cis-(+)-OPDA. Our synthetic and analytical methodologies will support studies on cis-(+)-OPDA conjugation with amino acids and research into the biological significance of these metabolites in plants.
Publikation

Meena, S. K.; Heidecker, M.; Engelmann, S.; Jaber, A.; de Vries, T.; Triller, S.; Baumann‐Kaschig, K.; Abel, S.; Behrens, S.; Gago-Zachert, S.; Altered expression levels of long noncoding natural antisense transcripts overlapping the UGT73C6 gene affect rosette size in Arabidopsis thaliana Plant J. 113, 460-477, (2023) DOI: 10.1111/tpj.16058

Natural antisense long noncoding RNAs (lncNATs) are involved in the regulation of gene expression in plants, modulating different relevant developmental processes and responses to various stimuli. We have identified and characterized two lncNATs (NAT1UGT73C6 and NAT2UGT73C6, collectively NATsUGT73C6) from Arabidopsis thaliana that are transcribed from gene fully overlapping UGT73C6, a member of the UGT73C subfamily of genes encoding UDP-glycosyltransferases (UGTs). Expression of both NATsUGT73C6 is developmentally controlled and occurs independently of the transcription of UGT73C6 in cis. Downregulation of NATsUGT73C6 levels through artificial microRNAs results in a reduction of the rosette area, while constitutive overexpression of NAT1UGT73C6 or NAT2UGT73C6 leads to the opposite phenotype, an increase in rosette size. This activity of NATsUGT73C6 relies on its RNA sequence, and, although modulation of UGT73C6 in cis cannot be excluded, the observed phenotypes are not a consequence of the regulation of UGT73C6 in trans. The NATsUGT73C6 levels were shown to affect cell proliferation and thus individual leaf size. Consistent with this concept, our data suggest that the NATsUGT73C6 influence the expression levels of key transcription factors involved in regulating leaf growth by modulating cell proliferation. These findings thus reveal an additional regulatory layer on the process of leaf growth.
Publikation

Kuhn, J. H.; Adkins, S.; Alioto, D.; Gago-Zachert, S.; et al., .; 2020 taxonomic update for phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales Arch. Virol. 165, 3023–3072, (2020) DOI: 10.1007/s00705-020-04731-2

In March 2020, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. At the genus rank, 20 new genera were added, two were deleted, one was moved, and three were renamed. At the species rank, 160 species were added, four were deleted, ten were moved and renamed, and 30 species were renamed. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV.
Publikation

Serra, P.; Carbonell, A.; Navarro, B.; Gago-Zachert, S.; Li, S.; Di Serio, F.; Flores, R.; Symptomatic plant viroid infections in phytopathogenic fungi: A request for a critical reassessment Proc. Natl. Acad. Sci. U.S.A. 117, 10126-10128, (2020) DOI: 10.1073/pnas.1922249117

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Publikation

Gago-Zachert, S.; Schuck, J.; Weinholdt, C.; Knoblich, M.; Pantaleo, V.; Grosse, I.; Gursinsky, T.; Behrens, S.-E.; Highly efficacious antiviral protection of plants by small interfering RNAs identified in vitro Nucleic Acids Res. 47, 9343-9357, (2019) DOI: 10.1093/nar/gkz678

In response to a viral infection, the plant’s RNA silencing machinery processes viral RNAs into a huge number of small interfering RNAs (siRNAs). However, a very few of these siRNAs actually interfere with viral replication. A reliable approach to identify these immunologically effective siRNAs (esiRNAs) and to define the characteristics underlying their activity has not been available so far. Here, we develop a novel screening approach that enables a rapid functional identification of antiviral esiRNAs. Tests on the efficacy of such identified esiRNAs of a model virus achieved a virtual full protection of plants against a massive subsequent infection in transient applications. We find that the functionality of esiRNAs depends crucially on two properties: the binding affinity to Argonaute proteins and the ability to access the target RNA. The ability to rapidly identify functional esiRNAs could be of great benefit for all RNA silencing-based plant protection measures against viruses and other pathogens.
Publikation

García, M. L.; Bó, E. D.; da Graça, J. V.; Gago-Zachert, S.; Hammond, J.; Moreno, P.; Natsuaki, T.; Pallás, V.; Navarro, J. A.; Reyes, C. A.; Luna, G. R.; Sasaya, T.; Tzanetakis, I. E.; Vaira, A. M.; Verbeek, M.; ICTV Report Consortium, .; Corrigendum: ICTV Virus Taxonomy Profile: Ophioviridae J. Gen. Virol. 99, 949-949, (2018) DOI: 10.1099/jgv.0.001093

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Publikation

García, M. L.; Bó, E. D.; da Graça, J. V.; Gago-Zachert, S.; Hammond, J.; Moreno, P.; Natsuaki, T.; Pallás, V.; Navarro, J. A.; Reyes, C. A.; Luna, G. R.; Sasaya, T.; Tzanetakis, I. E.; Vaira, A. M.; Verbeek, M.; ICTV Report Consortium, .; ICTV Virus Taxonomy Profile: Ophioviridae J. Gen. Virol. 98, 1161-1162, (2017) DOI: 10.1099/jgv.0.000836

The Ophioviridae is a family of filamentous plant viruses, with single-stranded negative, and possibly ambisense, RNA genomes of 11.3–12.5 kb divided into 3–4 segments, each encapsidated separately. Virions are naked filamentous nucleocapsids, forming kinked circles of at least two different contour lengths. The sole genus, Ophiovirus, includes seven species. Four ophioviruses are soil-transmitted and their natural hosts include trees, shrubs, vegetables and bulbous or corm-forming ornamentals, both monocots and dicots. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Ophioviridae, which is available at http://www.ictv.global/report/ophioviridae.
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.
Bücher und Buchkapitel

Flores, R.; Gago-Zachert, S.; Serra, P.; De la Peña, M.; Navarro, B.; Chrysanthemum Chlorotic Mottle Viroid (Hadidi, A., et al., eds.). 331-338, (2017) DOI: 10.1016/B978-0-12-801498-1.00031-0

Chrysanthemum chlorotic mottle viroid (CChMVd) (398–401 nt) belongs to genus Pelamoviroid, family Avsunviroidae and, like other members of this family, replicates in plastids through a rolling-circle mechanism involving hammerhead ribozymes. CChMVd RNA adopts a branched conformation stabilized by a kissing-loop interaction, resembling peach latent mosaic viroid in this respect. Chrysanthemum is the only natural and experimental host for CChMVd, which in the most sensitive varieties induces leaf mottling and chlorosis, delay in flowering, and dwarfing. The viroid has been found in major chrysanthemum growing areas including Europe and Asia. There are natural variants in which the change (UUUC→GAAA) mapping at a tetraloop in the CChMVd branched conformation is sufficient to change the symptomatic phenotype into a nonsymptomatic one without altering the viroid titer. Preinfection with nonsymptomatic variants prevents challenge inoculation with symptomatic ones. Moreover, experimental coinoculation with symptomatic and nonsymptomatic CChMVd variants results in symptomless phenotypes only when the latter is in vast excess, thus indicating its lower fitness.
  • Die Leibniz-Gemeinschaft
  • Digitalisierung in der Landwirtschaft
  • Martin-Luther Universität Halle
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