TY - JOUR A1 - Thirumalaikumar, Venkatesh P. A1 - Gorka, Michal A1 - Schulz, Karina A1 - Masclaux-Daubresse, Celine A1 - Sampathkumar, Arun A1 - Skirycz, Aleksandra A1 - Vierstra, Richard D. A1 - Balazadeh, Salma T1 - Selective autophagy regulates heat stress memory in Arabidopsis by NBR1-mediated targeting of HSP90.1 and ROF1 JF - Autophagy N2 - In nature, plants are constantly exposed to many transient, but recurring, stresses. Thus, to complete their life cycles, plants require a dynamic balance between capacities to recover following cessation of stress and maintenance of stress memory. Recently, we uncovered a new functional role for macroautophagy/autophagy in regulating recovery from heat stress (HS) and resetting cellular memory of HS inArabidopsis thaliana. Here, we demonstrated that NBR1 (next to BRCA1 gene 1) plays a crucial role as a receptor for selective autophagy during recovery from HS. Immunoblot analysis and confocal microscopy revealed that levels of the NBR1 protein, NBR1-labeled puncta, and NBR1 activity are all higher during the HS recovery phase than before. Co-immunoprecipitation analysis of proteins interacting with NBR1 and comparative proteomic analysis of annbr1-null mutant and wild-type plants identified 58 proteins as potential novel targets of NBR1. Cellular, biochemical and functional genetic studies confirmed that NBR1 interacts with HSP90.1 (heat shock protein 90.1) and ROF1 (rotamase FKBP 1), a member of the FKBP family, and mediates their degradation by autophagy, which represses the response to HS by attenuating the expression ofHSPgenes regulated by the HSFA2 transcription factor. Accordingly, loss-of-function mutation ofNBR1resulted in a stronger HS memory phenotype. Together, our results provide new insights into the mechanistic principles by which autophagy regulates plant response to recurrent HS. KW - Arabidopsis thaliana KW - heat stress KW - HSFA2 KW - HSP90.1 KW - NBR1 KW - ROF1 KW - selective autophagy KW - stress memory KW - stress recovery Y1 - 2020 U6 - https://doi.org/10.1080/15548627.2020.1820778 SN - 1554-8635 VL - 17 IS - 9 SP - 2184 EP - 2199 PB - Taylor & Francis CY - Abingdon ER - TY - JOUR A1 - Calderan-Rodrigues, Maria Juliana A1 - Luzarowski, Marcin A1 - Monte-Bello, Carolina Cassano A1 - Minen, Romina Ines A1 - Zühlke, Boris M. A1 - Nikoloski, Zoran A1 - Skirycz, Aleksandra A1 - Caldana, Camila T1 - Proteogenic dipeptides are characterized by diel fluctuations and target of rapamycin complex-signaling dependency in the model plant Arabidopsis thaliana JF - Frontiers in plant science : FPLS N2 - As autotrophic organisms, plants capture light energy to convert carbon dioxide into ATP, nicotinamide adenine dinucleotide phosphate (NADPH), and sugars, which are essential for the biosynthesis of building blocks, storage, and growth. At night, metabolism and growth can be sustained by mobilizing carbon (C) reserves. In response to changing environmental conditions, such as light-dark cycles, the small-molecule regulation of enzymatic activities is critical for reprogramming cellular metabolism. We have recently demonstrated that proteogenic dipeptides, protein degradation products, act as metabolic switches at the interface of proteostasis and central metabolism in both plants and yeast. Dipeptides accumulate in response to the environmental changes and act via direct binding and regulation of critical enzymatic activities, enabling C flux distribution. Here, we provide evidence pointing to the involvement of dipeptides in the metabolic rewiring characteristics for the day-night cycle in plants. Specifically, we measured the abundance of 13 amino acids and 179 dipeptides over short- (SD) and long-day (LD) diel cycles, each with different light intensities. Of the measured dipeptides, 38 and eight were characterized by day-night oscillation in SD and LD, respectively, reaching maximum accumulation at the end of the day and then gradually falling in the night. Not only the number of dipeptides, but also the amplitude of the oscillation was higher in SD compared with LD conditions. Notably, rhythmic dipeptides were enriched in the glucogenic amino acids that can be converted into glucose. Considering the known role of Target of Rapamycin (TOR) signaling in regulating both autophagy and metabolism, we subsequently investigated whether diurnal fluctuations of dipeptides levels are dependent on the TOR Complex (TORC). The Raptor1b mutant (raptor1b), known for the substantial reduction of TOR kinase activity, was characterized by the augmented accumulation of dipeptides, which is especially pronounced under LD conditions. We were particularly intrigued by the group of 16 dipeptides, which, based on their oscillation under SD conditions and accumulation in raptor1b, can be associated with limited C availability or photoperiod. By mining existing protein-metabolite interaction data, we delineated putative protein interactors for a representative dipeptide Pro-Gln. The obtained list included enzymes of C and amino acid metabolism, which are also linked to the TORC-mediated metabolic network. Based on the obtained results, we speculate that the diurnal accumulation of dipeptides contributes to its metabolic adaptation in response to changes in C availability. We hypothesize that dipeptides would act as alternative respiratory substrates and by directly modulating the activity of the focal enzymes. KW - dipeptide KW - diel cycle KW - metabolism KW - TOR signaling KW - protein-metabolite KW - interactions KW - carbon limitation KW - amino acid Y1 - 2021 U6 - https://doi.org/10.3389/fpls.2021.758933 SN - 1664-462X VL - 12 PB - Frontiers Media CY - Lausanne ER - TY - JOUR A1 - Stobiecki, Maciej A1 - Skirycz, Aleksandra A1 - Kerhoas, L. A1 - Kachlicki, P. A1 - Muth, D. A1 - Einhorn, J. A1 - Mueller-Roeber, Bernd T1 - Profiling of phenolic glycosidic conjugates in leaves of Arabidopsis thaliana using LC/MS JF - Metabolomics : the official journal of the Metabolomics Society N2 - Profiling of plant secondary metabolites is still a very difficult task. Liquid chromatography (LC) or capillary electrophoresis hyphenated with different kinds of detectors are methods of choice for analysis of polar, thermo labile compounds with high molecular masses. We demonstrate the applicability of LC combined with UV diode array or/and mass spectrometric detectors for the unambiguous identification and quantification of flavonoid conjugates isolated from Arahidopsis thaliana leaves of different genotypes and grown in different environmental conditions. During LC/UV/MS/MS analyses we were able to identify tetra-, tri, and di-glycosides of kaempferol, quercetin and isorhamnetin. Based on our results we can conclude that due to the co-elution of different chemical compounds in reversed phase H PLC systems the application of UV detectors does not allow to precisely profile all flavonoid conjugates existing in A. thaliana genotypes. Using MS detection it was possible to unambiguously recognize the glycosylation patterns of the aglycones. However, from the mass spectra we could not conclude neither the anomeric form of the C-1 carbon atoms of sugar moieties in glycosidic bonds between sugars or sugar and aglycone nor the position of the second carbon involved in disaccharides. The applicability of collision induced dissociation techniques (CID MS/MS) for structural analyses of the studied group of plant secondary metabolites with two types of analyzers (triple quadrupole or ion trap) was demonstrated. KW - liquid chromatography-mass spectrometry KW - metabolite profiling KW - metabolomics KW - flavonoid glycosides Y1 - 2006 U6 - https://doi.org/10.1007/s11306-006-0031-5 SN - 1573-3882 VL - 2 SP - 197 EP - 219 PB - Springer CY - New York ER - TY - JOUR A1 - Zupok, Arkadiusz A1 - Górka, Michał Jakub A1 - Siemiatkowska, Beata A1 - Skirycz, Aleksandra A1 - Leimkühler, Silke T1 - Iron-Dependent Regulation of Molybdenum Cofactor Biosynthesis Genes in Escherichia coli JF - Journal of bacteriology N2 - Molybdenum cofactor (Moco) biosynthesis is a complex process that involves the coordinated function of several proteins. In recent years it has become obvious that the availability of iron plays an important role in the biosynthesis of Moco. First, the MoaA protein binds two (4Fe-4S] clusters per monomer. Second, the expression of the moaABCDE and moeAB operons is regulated by FNR, which senses the availability of oxygen via a functional NFe-4S) cluster. Finally, the conversion of cyclic pyranopterin monophosphate to molybdopterin requires the availability of the L-cysteine desulfurase IscS, which is a shared protein with a main role in the assembly of Fe-S clusters. In this report, we investigated the transcriptional regulation of the moaABCDE operon by focusing on its dependence on cellular iron availability. While the abundance of selected molybdoenzymes is largely decreased under iron-limiting conditions, our data show that the regulation of the moaABCDE operon at the level of transcription is only marginally influenced by the availability of iron. Nevertheless, intracellular levels of Moco were decreased under iron-limiting conditions, likely based on an inactive MoaA protein in addition to lower levels of the L-cysteine desulfurase IscS, which simultaneously reduces the sulfur availability for Moco production. IMPORTANCE FNR is a very important transcriptional factor that represents the master switch for the expression of target genes in response to anaerobiosis. Among the FNR-regulated operons in Escherichia coli is the moaABCDE operon, involved in Moco biosynthesis. Molybdoenzymes have essential roles in eukaryotic and prokaryotic organisms. In bacteria, molybdoenzymes are crucial for anaerobic respiration using alternative electron acceptors. This work investigates the connection of iron availability to the biosynthesis of Moco and the production of active molybdoenzymes. KW - Escherichia coli KW - FNR KW - iron regulation KW - iron-sulfur cluster KW - anaerobic respiration KW - molybdenum cofactor Y1 - 2019 U6 - https://doi.org/10.1128/JB.00382-19 SN - 0021-9193 SN - 1098-5530 VL - 201 IS - 17 PB - American Society for Microbiology CY - Washington ER - TY - JOUR A1 - Friedrich, Thomas A1 - Oberkofler, Vicky A1 - Trindade, Inês A1 - Altmann, Simone A1 - Brzezinka, Krzysztof A1 - Lämke, Jörn S. A1 - Gorka, Michal A1 - Kappel, Christian A1 - Sokolowska, Ewelina A1 - Skirycz, Aleksandra A1 - Graf, Alexander A1 - Bäurle, Isabel T1 - Heteromeric HSFA2/HSFA3 complexes drive transcriptional memory after heat stress in Arabidopsis JF - Nature Communications N2 - Adaptive plasticity in stress responses is a key element of plant survival strategies. For instance, moderate heat stress (HS) primes a plant to acquire thermotolerance, which allows subsequent survival of more severe HS conditions. Acquired thermotolerance is actively maintained over several days (HS memory) and involves the sustained induction of memory-related genes. Here we show that FORGETTER3/ HEAT SHOCK TRANSCRIPTION FACTOR A3 (FGT3/HSFA3) is specifically required for physiological HS memory and maintaining high memory-gene expression during the days following a HS exposure. HSFA3 mediates HS memory by direct transcriptional activation of memory-related genes after return to normal growth temperatures. HSFA3 binds HSFA2, and in vivo both proteins form heteromeric complexes with additional HSFs. Our results indicate that only complexes containing both HSFA2 and HSFA3 efficiently promote transcriptional memory by positively influencing histone H3 lysine 4 (H3K4) hyper-methylation. In summary, our work defines the major HSF complex controlling transcriptional memory and elucidates the in vivo dynamics of HSF complexes during somatic stress memory. Moderate heat stress primes plants to acquire tolerance to subsequent, more severe heat stress. Here the authors show that the HSFA3 transcription factor forms a heteromeric complex with HSFA2 to sustain activated transcription of genes required for acquired thermotolerance by promoting H3K4 hyper-methylation. Y1 - 2021 U6 - https://doi.org/10.1038/s41467-021-23786-6 SN - 2041-1723 VL - 12 IS - 1 PB - Nature Publishing Group UK CY - [London] ER - TY - JOUR A1 - Skirycz, Aleksandra A1 - Reichelt, Michael A1 - Burow, Meike A1 - Birkemeyer, Claudia Sabine A1 - Rolcik, Jacub A1 - Kopka, Joachim A1 - Zanor, Maria Ines A1 - Gershenzon, Jonathan A1 - Strnad, Miroslav A1 - Szopa, Jan A1 - Müller-Röber, Bernd A1 - Witt, Isabell T1 - DOF transcription factor AtDof1.1 (OBP2) is part of a regulatory network controlling glucosinolate biosynthesis in Arabidopsis N2 - Glucosinolates are a group of secondary metabolites that function as defense substances against herbivores and micro-organisms in the plant order Capparales. Indole glucosinolates (IGS), derivatives of tryptophan, may also influence plant growth and development. In Arabidopsis thaliana, indole-3-acetaldoxime (IAOx) produced from tryptophan by the activity of two cytochrome P450 enzymes, CYP79B2 and CYP79B3, serves as a precursor for IGS biosynthesis but is also an intermediate in the biosynthetic pathway of indole-3-acetic acid (IAA). Another cytochrome P450 enzyme, CYP83B1, funnels IAOx into IGS. Although there is increasing information about the genes involved in this biochemical pathway, their regulation is not fully understood. OBP2 has recently been identified as a member of the DNA-binding-with-one- finger (DOF) transcription factors, but its function has not been studied in detail so far. Here we report that OBP2 is expressed in the vasculature of all Arabidopsis organs, including leaves, roots, flower stalks and petals. OBP2 expression is induced in response to a generalist herbivore, Spodoptera littoralis, and by treatment with the plant signalling molecule methyl jasmonate, both of which also trigger IGS accumulation. Constitutive and inducible over- expression of OBP2 activates expression of CYP83B1. In addition, auxin concentration is increased in leaves and seedlings of OBP2 over-expression lines relative to wild-type, and plant size is diminished due to a reduction in cell size. RNA interference-mediated OBP2 blockade leads to reduced expression of CYP83B1. Collectively, these data provide evidence that OBP2 is part of a regulatory network that regulates glucosinolate biosynthesis in Arabidopsis Y1 - 2006 UR - http://onlinelibrary.wiley.com/doi/10.1111/j.1365-313X.2006.02767.x/full ER - TY - JOUR A1 - Gomez-Merino, Fernando Carlos A1 - Arana-Ceballos, Fernando Alberto A1 - Trejo-Tellez, L. I. A1 - Skirycz, Aleksandra A1 - Brearley, C. A. A1 - Dormann, P. A1 - Müller-Röber, Bernd T1 - Arabidopsis AtDGK7, the smallest member of plant diacylglycerol kinases (DGKs), displays unique biochemical features and saturates at low substrate concentration : the DGK inhibitor R59022 differentially affects AtDGK2 and AtDGK7 activity in vitro and alters plant growth and development N2 - Diacylglycerol kinase (DGK) regulates the level of the second messenger diacylglycerol and produces phosphatidic acid (PA), another signaling molecule. The Arabidopsis thaliana genome encodes seven putative diacylglycerol kinase isozymes (named AtDGK1 to -7), structurally falling into three major clusters. So far, enzymatic activity has not been reported for any plant Cluster II DGK. Here, we demonstrate that a representative of this cluster, AtDGK7, is biochemically active when expressed as a recombinant protein in Escherichia coli. AtDGK7, encoded by gene locus At4g30340, contains 374 amino acids with an apparent molecular mass of 41.2 kDa. AtDGK7 harbors an N-terminal catalytic domain, but in contrast to various characterized DGKs (including AtDGK2), it lacks a cysteine-rich domain at its N terminus, and, importantly, its C-terminal DGK accessory domain is incomplete. Recombinant AtDGK7 expressed in E. coli exhibits Michaelis-Menten type kinetics with 1,2-dioleoyl-sn-glycerol as substrate. AtDGK7 activity was affected by pH, detergents, and the DGK inhibitor R59022. We demonstrate that both AtDGK2 and AtDGK7 phosphorylate diacylglycerol molecular species that are typically found in plants, indicating that both enzymes convert physiologically relevant substrates. AtDGK7 is expressed throughout the Arabidopsis plant, but expression is strongest in flowers and young seedlings. Expression of AtDGK2 is transiently induced by wounding. R59022 at similar to 80 mu M inhibits root elongation and lateral root formation and reduces plant growth, indicating that DGKs play an important role in plant development Y1 - 2005 SN - 0021-9258 ER -