Publications by year
2022
Skeffington AW, Gentzel M, Ohara A, Milentyev A, Heintze C, Böttcher L, Görlich S, Shevchenko A, Poulsen N, Kröger N, et al (2022). Shedding light on silica biomineralization by comparative analysis of the silica-associated proteomes from three diatom species.
The Plant journal : for cell and molecular biology,
110(6), 1700-1716.
Abstract:
Shedding light on silica biomineralization by comparative analysis of the silica-associated proteomes from three diatom species.
Morphogenesis of the intricate patterns of diatom silica cell walls is a protein-guided process, yet to date only very few such silica biomineralization proteins have been identified. Therefore, it is currently unknown whether all diatoms share conserved proteins of a basal silica forming machinery, and whether unique proteins are responsible for the morphogenesis of species-specific silica patterns. To answer these questions, we extracted proteins from the silica of three diatom species (Thalassiosira pseudonana, Thalassiosira oceanica, and Cyclotella cryptica) by complete demineralization of the cell walls. Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) analysis of the extracts identified 92 proteins that we name 'soluble silicome proteins' (SSPs). Surprisingly, no SSPs are common to all three species, and most SSPs showed very low similarity to one another in sequence alignments. In-depth bioinformatics analyses revealed that SSPs could be grouped into distinct classes based on short unconventional sequence motifs whose functions are yet unknown. The results from the in vivo localization of selected SSPs indicates that proteins, which lack sequence homology but share unconventional sequence motifs may exert similar functions in the morphogenesis of the diatom silica cell wall.
Abstract.
2021
Skeffington A, Gentzel M, Ohara A, Milentyev A, Heintze C, Böttcher L, Görlich S, Shevchenko A, Poulsen N, Kröger N, et al (2021). Shedding light on biosilica morphogenesis by comparative analysis of the silica-associated proteomes from three diatom species.
Abstract:
Shedding light on biosilica morphogenesis by comparative analysis of the silica-associated proteomes from three diatom species
Summary Morphogenesis of the intricate patterns of diatom silica cell walls is a protein-guided process, yet to date only very few such silica morphogenetic proteins have been identified. Therefore, it is unknown whether all diatoms share conserved proteins of a basal silica forming machinery, and whether unique proteins are responsible for the morphogenesis of species specific silica patterns. To answer these questions, we extracted proteins from the silica of three diatom species ( Thalassiosira pseudonana, Thalassiosira oceanica and Cyclotella cryptica ) by complete demineralization of the cell walls. LC-MS/MS analysis of the extracts identified 92 proteins that we name ‘Soluble Silicome Proteins’ (SSPs). Surprisingly, no SSPs are common to all three species, and most SSPs showed very low similarity to one another in sequence alignments. In depth bioinformatics analyses revealed that SSPs can be grouped into distinct classes bases on short unconventional sequence motifs whose functions are yet unknown. The results from in vivo localization of selected SSPs indicates that proteins, which lack sequence homology but share unconventional sequence motifs may exert similar functions in the morphogenesis of the diatom silica cell wall.
Abstract.
2020
Skeffington AW, Grimm A, Schönefeld S, Petersen K, Scheffel A (2020). An Efficient Method for the Plating of Haploid and Diploid Emiliania huxleyi on Solid Medium1.
Journal of phycology,
56(1), 238-242.
Abstract:
An Efficient Method for the Plating of Haploid and Diploid Emiliania huxleyi on Solid Medium1.
Emiliania huxleyi is a globally important coccolithophore and one of the most successful eukaryotic organisms in the modern oceans. Despite a large body of work on this organism, including the sequencing of its genome, the tools required for forward and reverse functional genetic studies are still undeveloped. Here we present an optimized method for the clonal isolation of E. huxleyi by plating on solid medium. We demonstrate the utility of this method for a variety of strains including haploid, calcifying-diploid, and noncalcifying diploid strains. We show that, in contrast to previous studies, no changes in cell ploidy status occur when the cells are plated. Our method will greatly aid attempts to elucidate the genetic basis of the remarkable physiology of E. huxleyi by forward and reverse genetic approaches.
Abstract.
Skeffington A, Donath A (2020). ProminTools: Shedding light on proteins of unknown function in biomineralization with user friendly tools illustrated using mollusc shell matrix protein sequences.
Abstract:
ProminTools: Shedding light on proteins of unknown function in biomineralization with user friendly tools illustrated using mollusc shell matrix protein sequences
Biominerals are crucial to the fitness of many organism and studies of the mechanisms of biomineralization are driving research into novel materials. Biomineralization is generally controlled by a matrix of organic molecules including proteins, so proteomic studies of biominerals are important for understanding biomineralization mechanisms. Many such studies identify large numbers of proteins of unknown function, which are often of low sequence complexity and biased in their amino acid composition. A lack of user-friendly tools to find patterns in such sequences and robustly analyse their statistical properties relative to the background proteome means that they are often neglected in follow-up studies. Here we present ProminTools, a user-friendly package for comparison of two sets of protein sequences in terms of their global properties and motif content. Outputs include data tables, graphical summaries in an html file and an R-script as a starting point for data-set specific visualizations. We demonstrate the utility of ProminTools using a previously published shell matrix proteome of the giant limpet Lottia gigantea.
Abstract.
Skeffington AW, Donath A (2020). ProminTools: shedding light on proteins of unknown function in biomineralization with user friendly tools illustrated using mollusc shell matrix protein sequences.
PeerJ,
8Abstract:
ProminTools: shedding light on proteins of unknown function in biomineralization with user friendly tools illustrated using mollusc shell matrix protein sequences.
Biominerals are crucial to the fitness of many organism and studies of the mechanisms of biomineralization are driving research into novel materials. Biomineralization is generally controlled by a matrix of organic molecules including proteins, so proteomic studies of biominerals are important for understanding biomineralization mechanisms. Many such studies identify large numbers of proteins of unknown function, which are often of low sequence complexity and biased in their amino acid composition. A lack of user-friendly tools to find patterns in such sequences and robustly analyse their statistical properties relative to the background proteome means that they are often neglected in follow-up studies. Here we present ProminTools, a user-friendly package for comparison of two sets of protein sequences in terms of their global properties and motif content. Outputs include data tables, graphical summaries in an html file and an R-script as a starting point for data-set specific visualizations. We demonstrate the utility of ProminTools using a previously published shell matrix proteome of the giant limpet Lottia gigantea.
Abstract.
2018
Skeffington AW, Scheffel A (2018). Exploiting algal mineralization for nanotechnology: bringing coccoliths to the fore.
Current opinion in biotechnology,
49, 57-63.
Abstract:
Exploiting algal mineralization for nanotechnology: bringing coccoliths to the fore.
Complex mineral structures are produced by many microalgal species. Pioneering work on diatom silica has demonstrated the potential of such structures in nanotechnology. The calcified scales of coccolithophores (coccoliths) have received less attention, but the large diversity of architectures make coccoliths attractive as parts for nano-devices. Currently coccolith calcite can be modified by the incorporation of metal ions or adsorption of enzymes to the surface, but genetic modification of coccolithophores may permit the production of coccoliths with customized architectures and surface properties. Further work on the laboratory cultivation of diverse species, the physiochemical properties of coccoliths and on genetic tools for coccolithophores will be necessary to realize the full potential of coccoliths in nanotechnology.
Abstract.
2014
Skeffington AW, Graf A, Duxbury Z, Gruissem W, Smith AM (2014). Glucan, Water Dikinase Exerts Little Control over Starch Degradation in Arabidopsis Leaves at Night.
Plant physiology,
165(2), 866-879.
Abstract:
Glucan, Water Dikinase Exerts Little Control over Starch Degradation in Arabidopsis Leaves at Night.
The first step on the pathway of starch degradation in Arabidopsis (Arabidopsis thaliana) leaves at night is the phosphorylation of starch polymers, catalyzed by glucan, water dikinase (GWD). It has been suggested that GWD is important for the control of starch degradation, because its transcript levels undergo strong diel fluctuations, its activity is subject to redox regulation in vitro, and starch degradation is strongly decreased in gwd mutant plants. To test this suggestion, we analyzed changes in GWD protein abundance in relation to starch levels in wild-type plants, in transgenic plants in which GWD transcripts were strongly reduced by induction of RNA interference, and in transgenic plants overexpressing GWD. We found that GWD protein levels do not vary over the diel cycle and that the protein has a half-life of 2 d. Overexpression of GWD does not accelerate starch degradation in leaves, and starch degradation is not inhibited until GWD levels are reduced by 70%. Surprisingly, this degree of reduction also inhibits starch synthesis in the light. To discover the importance of redox regulation, we generated transgenic plants expressing constitutively active GWD. These plants retained normal control of degradation. We conclude that GWD exerts only a low level of control over starch degradation in Arabidopsis leaves.
Abstract.
2013
Scialdone A, Mugford ST, Feike D, Skeffington A, Borrill P, Graf A, Smith AM, Howard M (2013). Arabidopsis plants perform arithmetic division to prevent starvation at night.
Elife,
2Abstract:
Arabidopsis plants perform arithmetic division to prevent starvation at night.
Photosynthetic starch reserves that accumulate in Arabidopsis leaves during the day decrease approximately linearly with time at night to support metabolism and growth. We find that the rate of decrease is adjusted to accommodate variation in the time of onset of darkness and starch content, such that reserves last almost precisely until dawn. Generation of these dynamics therefore requires an arithmetic division computation between the starch content and expected time to dawn. We introduce two novel chemical kinetic models capable of implementing analog arithmetic division. Predictions from the models are successfully tested in plants perturbed by a night-time light period or by mutations in starch degradation pathways. Our experiments indicate which components of the starch degradation apparatus may be important for appropriate arithmetic division. Our results are potentially relevant for any biological system dependent on a food reserve for survival over a predictable time period. DOI:http://dx.doi.org/10.7554/eLife.00669.001.
Abstract.
Author URL.
2009
Robertson FC, Skeffington AW, Gardner MJ, Webb AAR (2009). Interactions between circadian and hormonal signalling in plants.
Plant molecular biology,
69(4), 419-427.
Abstract:
Interactions between circadian and hormonal signalling in plants.
Growth and development of plants is controlled by external and internal signals. Key internal signals are those generated by hormones and the circadian clock. We highlight interactions between the circadian clock and hormonal signalling networks in regulating the physiology and growth of plants. Microarray analysis has shown that a significant proportion of transcripts involved in hormonal metabolism, catabolism, perception and signalling are also regulated by the circadian clock. In particular, there are interactions between the clock and abscisic acid, auxin, cytokinin and ethylene signalling. We discuss the role of circadian modulation ('gating') of hormonal signals in preventing temporally inappropriate responses. A consideration of the daily changes in physiology provides evidence that circadian gating of hormonal signalling couples the rhythmic regulation of carbon and water utilisation to rhythmic patterns of growth.
Abstract.
Karunakaran R, Ramachandran VK, Seaman JC, East AK, Mouhsine B, Mauchline TH, Prell J, Skeffington A, Poole PS (2009). Transcriptomic analysis of Rhizobium leguminosarum biovar viciae in symbiosis with host plants Pisum sativum and Vicia cracca.
Journal of bacteriology,
191(12), 4002-4014.
Abstract:
Transcriptomic analysis of Rhizobium leguminosarum biovar viciae in symbiosis with host plants Pisum sativum and Vicia cracca.
Rhizobium leguminosarum bv. viciae forms nitrogen-fixing nodules on several legumes, including pea (Pisum sativum) and vetch (Vicia cracca), and has been widely used as a model to study nodule biochemistry. To understand the complex biochemical and developmental changes undergone by R. leguminosarum bv. viciae during bacteroid development, microarray experiments were first performed with cultured bacteria grown on a variety of carbon substrates (glucose, pyruvate, succinate, inositol, acetate, and acetoacetate) and then compared to bacteroids. Bacteroid metabolism is essentially that of dicarboxylate-grown cells (i.e. induction of dicarboxylate transport, gluconeogenesis and alanine synthesis, and repression of sugar utilization). The decarboxylating arm of the tricarboxylic acid cycle is highly induced, as is gamma-aminobutyrate metabolism, particularly in bacteroids from early (7-day) nodules. To investigate bacteroid development, gene expression in bacteroids was analyzed at 7, 15, and 21 days postinoculation of peas. This revealed that bacterial rRNA isolated from pea, but not vetch, is extensively processed in mature bacteroids. In early development (7 days), there were large changes in the expression of regulators, exported and cell surface molecules, multidrug exporters, and heat and cold shock proteins. fix genes were induced early but continued to increase in mature bacteroids, while nif genes were induced strongly in older bacteroids. Mutation of 37 genes that were strongly upregulated in mature bacteroids revealed that none were essential for nitrogen fixation. However, screening of 3,072 mini-Tn5 mutants on peas revealed previously uncharacterized genes essential for nitrogen fixation. These encoded a potential magnesium transporter, an AAA domain protein, and proteins involved in cytochrome synthesis.
Abstract.
