@article{NakamuraSteupColleonietal.2022, author = {Nakamura, Yasunori and Steup, Martin and Colleoni, Christophe and Iglesias, Alberto A. and Bao, Jinsong and Fujita, Naoko and Tetlow, Ian}, title = {Molecular regulation of starch metabolism}, series = {Plant molecular biology : an international journal of fundamental research and genetic engineering}, volume = {108}, journal = {Plant molecular biology : an international journal of fundamental research and genetic engineering}, number = {4-5}, publisher = {Springer}, address = {Dordrecht}, issn = {0167-4412}, doi = {10.1007/s11103-022-01253-0}, pages = {289 -- 290}, year = {2022}, language = {en} } @misc{CisekTokarzKontenisetal.2018, author = {Cisek, Richard and Tokarz, Danielle and Kontenis, Lukas and Barzda, Virginijus and Steup, Martin}, title = {Polarimetric second harmonic generation microscopy}, series = {Starch-Starke}, volume = {70}, journal = {Starch-Starke}, number = {1-2}, publisher = {Wiley-VCH}, address = {Weinheim}, issn = {0038-9056}, doi = {10.1002/star.201700031}, pages = {15}, year = {2018}, abstract = {Second harmonic generation (SHG) is a nonlinear optical process that inherently generates signal in non-centrosymmetric materials, such as starch granules, and therefore can be used for label-free imaging. Both intensity and polarization of SHG are determined by material properties that are characterized by the nonlinear susceptibility tensor, ((2)). Examination of the tensor is performed for each focal volume of the image by measuring the outgoing polarization state of the SHG signal for a set of incoming laser beam polarizations. Mapping of nonlinear properties expressed as the susceptibility ratio reveals structural features including the organization of crystalline material within a single starch granule, and the distribution of structural properties in a population of granules. Isolated granules, as well as in situ starch, can be analyzed using polarimetric SHG microscopy. Due to the fast sample preparation and short imaging times, polarimetric SHG microscopy allows for a quick assessment of starch structure and permits rapid feedback for bioengineering applications. This article presents the basics of SHG theory and microscopy applications for starch-containing materials. Quantification of ultrastructural features within individual starch granules is described. New results obtained by polarization resolved SHG microscopy of starch granules are presented for various maize genotypes revealing heterogeneity within a single starch particle and between various granules.}, language = {en} } @article{SmirnovaFernieSpahnetal.2017, author = {Smirnova, Julia and Fernie, Alisdair R. and Spahn, Christian M. T. and Steup, Martin}, title = {Photometric assay of maltose and maltose-forming enzyme activity by using 4-alpha-glucanotransferase (DPE2) from higher plants}, series = {Analytical biochemistry : methods in the biological sciences}, volume = {532}, journal = {Analytical biochemistry : methods in the biological sciences}, publisher = {Elsevier}, address = {San Diego}, issn = {0003-2697}, doi = {10.1016/j.ab.2017.05.026}, pages = {72 -- 82}, year = {2017}, abstract = {Maltose frequently occurs as intermediate of the central carbon metabolism of prokaryotic and eukaryotic cells. Various mutants possess elevated maltose levels. Maltose exists as two anomers, (alpha- and beta-form) which are rapidly interconverted without requiring enzyme-mediated catalysis. As maltose is often abundant together with other oligoglucans, selective quantification is essential. In this communication, we present a photometric maltose assay using 4-alpha-glucanotransferase (AtDPE2) from Arabidopsis thaliana. Under in vitro conditions, AtDPE2 utilizes maltose as glucosyl donor and glycogen as acceptor releasing the other hexosyl unit as free glucose which is photometrically quantified following enzymatic phosphorylation and oxidation. Under the conditions used, DPE2 does not noticeably react with other di- or oligosaccharides. Selectivity compares favorably with that of maltase frequently used in maltose assays. Reducing end interconversion of the two maltose anomers is in rapid equilibrium and, therefore, the novel assay measures total maltose contents. Furthermore, an AtDPE2-based continuous photometric assay is presented which allows to quantify beta-amylase activity and was found to be superior to a conventional test. Finally, the AtDPE2-based maltose assay was used to quantify leaf maltose contents of both Arabidopsis wild type and AtDPE2-deficient plants throughout the light-dark cycle. These data are presented together with assimilatory starch levels. (C) 2017 Published by Elsevier Inc.}, language = {en} } @article{JueppnerMubeenLeisseetal.2017, author = {J{\"u}ppner, Jessica and Mubeen, Umarah and Leisse, Andrea and Caldana, Camila and Brust, Henrike and Steup, Martin and Herrmann, Marion and Steinhauser, Dirk and Giavalisco, Patrick}, title = {Dynamics of lipids and metabolites during the cell cycle of Chlamydomonas reinhardtii}, series = {The plant journal}, volume = {92}, journal = {The plant journal}, publisher = {Wiley}, address = {Hoboken}, issn = {0960-7412}, doi = {10.1111/tpj.13642}, pages = {331 -- 343}, year = {2017}, abstract = {Metabolites and lipids are the final products of enzymatic processes, distinguishing the different cellular functions and activities of single cells or whole tissues. Understanding these cellular functions within a well-established model system requires a systemic collection of molecular and physiological information. In the current report, the green alga Chlamydomonas reinhardtii was selected to establish a comprehensive workflow for the detailed multi-omics analysis of a synchronously growing cell culture system. After implementation and benchmarking of the synchronous cell culture, a two-phase extraction method was adopted for the analysis of proteins, lipids, metabolites and starch from a single sample aliquot of as little as 10-15million Chlamydomonas cells. In a proof of concept study, primary metabolites and lipids were sampled throughout the diurnal cell cycle. The results of these time-resolved measurements showed that single compounds were not only coordinated with each other in different pathways, but that these complex metabolic signatures have the potential to be used as biomarkers of various cellular processes. Taken together, the developed workflow, including the synchronized growth of the photoautotrophic cell culture, in combination with comprehensive extraction methods and detailed metabolic phenotyping has the potential for use in in-depth analysis of complex cellular processes, providing essential information for the understanding of complex biological systems.}, language = {en} } @article{NakamuraOnoSawadaetal.2017, author = {Nakamura, Yasunori and Ono, Masami and Sawada, Takayuki and Crofts, Naoko and Fujita, Naoko and Steup, Martin}, title = {Characterization of the functional interactions of plastidial starch phosphorylase and starch branching enzymes from rice endosperm during reserve starch biosynthesis}, series = {Plant science : an international journal of experimental plant biology}, volume = {264}, journal = {Plant science : an international journal of experimental plant biology}, publisher = {Elsevier}, address = {Clare}, issn = {0168-9452}, doi = {10.1016/j.plantsci.2017.09.002}, pages = {83 -- 95}, year = {2017}, abstract = {Functional interactions of plastidial phosphorylase (Phol) and starch branching enzymes (BEs) from the developing rice endosperm are the focus of this study. In the presence of both Phol and BE, the same branched primer molecule is elongated and further branched almost simultaneously even at very low glucan concentrations present in the purified enzyme preparations. By contrast, in the absence of any BE, glucans are not, to any significant extent, elongated by Phol. Based on our in vitro data, in the developing rice endosperm, Phol appears to be weakly associated with any of the BE isozymes. By using fluorophore-labeled malto-oligosaccharides, we identified maltose as the smallest possible primer for elongation by Phol. Linear dextrins act as carbohydrate substrates for BEs. By functionally interacting with a BE, Phol performs two essential functions during the initiation of starch biosynthesis in the rice endosperm: First, it elongates maltodextrins up to a degree of polymerization of at least 60. Second, by closely interacting with BEs, Phol is able to elongate branched glucans efficiently and thereby synthesizes branched carbohydrates essential for the initiation of amylopectin biosynthesis.}, language = {en} } @misc{SullivanNitschkeSteupetal.2017, author = {Sullivan, Mitchell A. and Nitschke, Silvia and Steup, Martin and Minassian, Berge A. and Nitschke, Felix}, title = {Pathogenesis of Lafora disease}, series = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe}, journal = {Postprints der Universit{\"a}t Potsdam : Mathematisch-Naturwissenschaftliche Reihe}, number = {1080}, issn = {1866-8372}, doi = {10.25932/publishup-47462}, url = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-474622}, pages = {18}, year = {2017}, abstract = {Lafora disease (LD, OMIM \#254780) is a rare, recessively inherited neurodegenerative disease with adolescent onset, resulting in progressive myoclonus epilepsy which is fatal usually within ten years of symptom onset. The disease is caused by loss-of-function mutations in either of the two genes EPM2A (laforin) or EPM2B (malin). It characteristically involves the accumulation of insoluble glycogen-derived particles, named Lafora bodies (LBs), which are considered neurotoxic and causative of the disease. The pathogenesis of LD is therefore centred on the question of how insoluble LBs emerge from soluble glycogen. Recent data clearly show that an abnormal glycogen chain length distribution, but neither hyperphosphorylation nor impairment of general autophagy, strictly correlates with glycogen accumulation and the presence of LBs. This review summarizes results obtained with patients, mouse models, and cell lines and consolidates apparent paradoxes in the LD literature. Based on the growing body of evidence, it proposes that LD is predominantly caused by an impairment in chain-length regulation affecting only a small proportion of the cellular glycogen. A better grasp of LD pathogenesis will further develop our understanding of glycogen metabolism and structure. It will also facilitate the development of clinical interventions that appropriately target the underlying cause of LD.}, language = {en} } @article{SchwarteWegnerHavensteinetal.2015, author = {Schwarte, Sandra and Wegner, Fanny and Havenstein, Katja and Groth, Detlef and Steup, Martin and Tiedemann, Ralph}, title = {Sequence variation, differential expression, and divergent evolution in starch-related genes among accessions of Arabidopsis thaliana}, series = {Plant molecular biology : an international journal of fundamental research and genetic engineering}, volume = {87}, journal = {Plant molecular biology : an international journal of fundamental research and genetic engineering}, number = {4-5}, publisher = {Springer}, address = {Dordrecht}, issn = {0167-4412}, doi = {10.1007/s11103-015-0293-2}, pages = {489 -- 519}, year = {2015}, abstract = {Transitory starch metabolism is a nonlinear and highly regulated process. It originated very early in the evolution of chloroplast-containing cells and is largely based on a mosaic of genes derived from either the eukaryotic host cell or the prokaryotic endosymbiont. Initially located in the cytoplasm, starch metabolism was rewired into plastids in Chloroplastida. Relocation was accompanied by gene duplications that occurred in most starch-related gene families and resulted in subfunctionalization of the respective gene products. Starch-related isozymes were then evolutionary conserved by constraints such as internal starch structure, posttranslational protein import into plastids and interactions with other starch-related proteins. 25 starch-related genes in 26 accessions of Arabidopsis thaliana were sequenced to assess intraspecific diversity, phylogenetic relationships, and modes of selection. Furthermore, sequences derived from additional 80 accessions that are publicly available were analyzed. Diversity varies significantly among the starch-related genes. Starch synthases and phosphorylases exhibit highest nucleotide diversities, while pyrophosphatases and debranching enzymes are most conserved. The gene trees are most compatible with a scenario of extensive recombination, perhaps in a Pleistocene refugium. Most genes are under purifying selection, but disruptive selection was inferred for a few genes/substitutiones. To study transcript levels, leaves were harvested throughout the light period. By quantifying the transcript levels and by analyzing the sequence of the respective accessions, we were able to estimate whether transcript levels are mainly determined by genetic (i.e., accession dependent) or physiological (i.e., time dependent) parameters. We also identified polymorphic sites that putatively affect pattern or the level of transcripts.}, language = {en} } @article{RadingSandmannSteupetal.2015, author = {Rading, M. Michael and Sandmann, Michael and Steup, Martin and Chiarugi, Davide and Valleriani, Angelo}, title = {Weak correlation of starch and volume in synchronized photosynthetic cells}, series = {Physical review : E, Statistical, nonlinear and soft matter physics}, volume = {91}, journal = {Physical review : E, Statistical, nonlinear and soft matter physics}, number = {1}, publisher = {American Physical Society}, address = {College Park}, issn = {1539-3755}, doi = {10.1103/PhysRevE.91.012711}, pages = {11}, year = {2015}, abstract = {In cultures of unicellular algae, features of single cells, such as cellular volume and starch content, are thought to be the result of carefully balanced growth and division processes. Single-cell analyses of synchronized photoautotrophic cultures of the unicellular alga Chlamydomonas reinhardtii reveal, however, that the cellular volume and starch content are only weakly correlated. Likewise, other cell parameters, e.g., the chlorophyll content per cell, are only weakly correlated with cell size. We derive the cell size distributions at the beginning of each synchronization cycle considering growth, timing of cell division and daughter cell release, and the uneven division of cell volume. Furthermore, we investigate the link between cell volume growth and starch accumulation. This work presents evidence that, under the experimental conditions of light-dark synchronized cultures, the weak correlation between both cell features is a result of a cumulative process rather than due to asymmetric partition of biomolecules during cell division. This cumulative process necessarily limits cellular similarities within a synchronized cell population.}, language = {en} } @article{CisekTokarzSteupetal.2015, author = {Cisek, Richard and Tokarz, Danielle and Steup, Martin and Tetlow, Ian J. and Emes, Michael J. and Hebelstrup, Kim H. and Blennow, Andreas and Barzda, Virginijus}, title = {Second harmonic generation microscopy investigation of the crystalline ultrastructure of three barley starch lines affected by hydration}, series = {Biomedical optics express}, volume = {6}, journal = {Biomedical optics express}, number = {10}, publisher = {Optical Society of America}, address = {Washington}, issn = {2156-7085}, doi = {10.1364/BOE.6.003694}, pages = {3694 -- 3700}, year = {2015}, abstract = {Second harmonic generation (SHG) microscopy is employed to study changes in crystalline organization due to altered gene expression and hydration in barley starch granules. SHG intensity and susceptibility ratio values (R'(SHG)) are obtained using reduced Stokes-Mueller polarimetric microscopy. The maximum R'(SHG) values occur at moderate moisture indicating the narrowest orientation distribution of nonlinear dipoles from the cylindrical axis of glucan helices. The maximum SHG intensity occurs at the highest moisture and amylopectin content. These results support the hypothesis that SHG is caused by ordered hydrogen and hydroxyl bond networks which increase with hydration of starch granules. (C) 2015 Optical Society of America}, language = {en} } @article{HemmeVeyelMuehlhausetal.2014, author = {Hemme, Dorothea and Veyel, Daniel and Muehlhaus, Timo and Sommer, Frederik and Jueppner, Jessica and Unger, Ann-Katrin and Sandmann, Michael and Fehrle, Ines and Schoenfelder, Stephanie and Steup, Martin and Geimer, Stefan and Kopka, Joachim and Giavalisco, Patrick and Schroda, Michael}, title = {Systems-wide analysis of acclimation responses to long-term heat stress and recovery in the photosynthetic model organism Chlamydomonas reinhardtii}, series = {The plant cell}, volume = {26}, journal = {The plant cell}, number = {11}, publisher = {American Society of Plant Physiologists}, address = {Rockville}, issn = {1040-4651}, doi = {10.1105/tpc.114.130997}, pages = {4270 -- 4297}, year = {2014}, abstract = {We applied a top-down systems biology approach to understand how Chlamydomonas reinhardtii acclimates to long-term heat stress (HS) and recovers from it. For this, we shifted cells from 25 to 42 degrees C for 24 h and back to 25 degrees C for >= 8 h and monitored abundances of 1856 proteins/protein groups, 99 polar and 185 lipophilic metabolites, and cytological and photosynthesis parameters. Our data indicate that acclimation of Chlamydomonas to long-term HS consists of a temporally ordered, orchestrated implementation of response elements at various system levels. These comprise (1) cell cycle arrest; (2) catabolism of larger molecules to generate compounds with roles in stress protection; (3) accumulation of molecular chaperones to restore protein homeostasis together with compatible solutes; (4) redirection of photosynthetic energy and reducing power from the Calvin cycle to the de novo synthesis of saturated fatty acids to replace polyunsaturated ones in membrane lipids, which are deposited in lipid bodies; and (5) when sinks for photosynthetic energy and reducing power are depleted, resumption of Calvin cycle activity associated with increased photorespiration, accumulation of reactive oxygen species scavengers, and throttling of linear electron flow by antenna uncoupling. During recovery from HS, cells appear to focus on processes allowing rapid resumption of growth rather than restoring pre-HS conditions.}, language = {en} }