Christoph Dockter

Stanford University, Palo Alto, California, United States

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Publications (14)72.23 Total impact

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    ABSTRACT: The erectoides-m anthocyanin-less 1 (ert-m ant1) double mutants are among the very few examples of induced double mutants in barley. From phenotypic observations of mutant plants it is known that the Ert-m gene product regulates plant architecture whereas the Ant1 gene product is involved in anthocyanin biosynthesis. We used a near-isogenic line of the cultivar Bowman, BW316 (ert-m.34), to create four F2-mapping populations by crosses to the barley cultivars Barke, Morex, Bowman and Quench. We phenotyped and genotyped 460 plants, allowing the ert-m mutation to be mapped to an interval of 4.7 cM on the short arm of barley chromosome 7H. Bioinformatic searches identified 21 candidate gene models in the mapped region. One gene was orthologous to a regulator of Arabidopsis thaliana plant architecture, ERECTA, encoding a leucine-rich repeat receptor-like kinase. Sequencing of HvERECTA in barley ert-m mutant accessions identified severe DNA changes in 15 mutants, including full gene deletions in ert-m.40 and ert-m.64. Both deletions, additionally causing anthocyanin deficiency, were found to stretch over a large region including two putative candidate genes for the anthocyanin biosynthesis locus Ant1. Analyses of ert-m and ant1 single- and double-deletion mutants suggest Ant1 as a closely linked gene encoding a R2R3 myeloblastosis transcription factor.
    Plant Molecular Biology 07/2015; 88(6). DOI:10.1007/s11103-015-0350-x · 4.07 Impact Factor
  • Christoph Dockter · Mats Hansson
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    ABSTRACT: The Green Revolution combined advancements in breeding and agricultural practice, and provided food security to millions of people. Daily food supply is still a major issue in many parts of the world and is further challenged by future climate change. Fortunately, life science research is currently making huge progress, and the development of future crop plants will be explored. Today, plant breeding typically follows one gene per trait. However, new scientific achievements have revealed that many of these traits depend on different genes and complex interactions of proteins reacting to various external stimuli. These findings open up new possibilities for breeding where variations in several genes can be combined to enhance productivity and quality. In this review we present an overview of genes determining plant architecture in barley, with a special focus on culm length. Many genes are currently known only through their mutant phenotypes, but emerging genomic sequence information will accelerate their identification. More than 1000 different short-culm barley mutants have been isolated and classified in different phenotypic groups according to culm length and additional pleiotropic characters. Some mutants have been connected to deficiencies in biosynthesis and reception of brassinosteroids and gibberellic acids. Still other mutants are unlikely to be connected to these hormones. The genes and corresponding mutations are of potential interest for development of stiff-straw crop plants tolerant to lodging, which occurs in extreme weather conditions with strong winds and heavy precipitation. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.
    Journal of Experimental Botany 01/2015; 66(12). DOI:10.1093/jxb/eru521 · 5.79 Impact Factor
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    ABSTRACT: Reduced plant height and culm robustness are quantitative characteristics important for assuring cereal crop yield and quality under adverse weather conditions. A very limited number of short-culm mutant alleles were introduced into commercial crop cultivars during the “Green Revolution”. We identified phenotypic traits, including sturdy culm, specific for deficiencies in brassinosteroid biosynthesis and signaling in semi-dwarf mutants of barley (Hordeum vulgare L.). The phenotypic traits were explored to perform a phenotypic screen of near-isogenic short-culm mutant lines from the brachytic, breviaristatum, dense spike, erectoides, semi-brachytic, semidwarf, and slender dwarf mutant groups. In silico mapping of brassinosteroid-related genes in the barley genome in combination with sequencing of barley mutant lines assigned more than 20 historic mutants to three brassinosteroid-biosynthesis genes (Hv-BRD, Hv-CPD, Hv-DIM) and one brassinosteroid-signaling gene (Hv-BRI1). Analyses of F2- and M2-populations, allelic crosses, and modeling of non-synonymous amino-acid exchanges in protein crystal structures gave a further understanding of the control of barley plant architecture and sturdiness by brassinosteroid-related genes. Alternatives to the widely used but highly temperature-sensitive uzu1.a allele of Hv-BRI1 are presented as genetic building blocks for breeding strategies with sturdy and climate-tolerant barley cultivars.
    Plant physiology 12/2014; 166:1912. DOI:10.1104/pp.114.250738 · 7.39 Impact Factor
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    ABSTRACT: Within the cereal grasses, variation in inflorescence architecture results in a conspicuous morphological diversity that in crop species influences the yield of cereal grains. Although significant progress has been made in identifying some of the genes underlying this variation in maize and rice, in the temperate cereals, a group that includes wheat, barley, and rye, only the dosage-dependent and highly pleiotropic Q locus in hexaploid wheat has been molecularly characterized. Here we show that the characteristic variation in the density of grains along the inflorescence, or spike, of modern cultivated barley (Hordeum vulgare) is largely the consequence of a perturbed interaction between microRNA172 and its corresponding binding site in the mRNA of an APELATA2 (AP2)-like transcription factor, HvAP2. We used genome-wide association and biparental mapping to identify HvAP2. By comparing inflorescence development and HvAP2 transcript abundance in an extreme dense-spike mutant and its nearly isogenic WT line, we show that HvAP2 turnover driven by microRNA 172 regulates the length of a critical developmental window that is required for elongation of the inflorescence internodes. Our data indicate that this heterochronic change, an altered timing of developmental events caused by specific temporal variation in the efficiency of HvAP2 turnover, leads to the striking differences in the size and shape of the barley spike.
    Proceedings of the National Academy of Sciences 09/2013; 110(41). DOI:10.1073/pnas.1311681110 · 9.81 Impact Factor
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    ABSTRACT: The barley (Hordeum vulgare L.) mutants fch2 and clo-f2 comprise an allelic group of 14 Chl b-deficient lines. The genetic map position of fch2 corresponds to the physical map position of the gene encoding chlorophyllide a oxygenase. This enzyme converts chlorophyllide a to chlorophyllide b and it is essential for Chl b biosynthesis. The fch2 and clo-f2 barley lines were shown to be mutated in the gene for chlorophyllide a oxygenase. A five-base insertion was found in fch2 and base deletions in clo-f2.101, clo-f2.105, clo-f2.2800 and clo-f2.3613. In clo-f2.105 and clo-f2.108, nonsense base exchanges were discovered. All of these mutations led to a premature stop of translation and none of the mutants formed Chl b. The mutant clo-f2.2807 was transcript deficient and formed no Chl b. Missense mutations in clo-f2.102 (leading to the amino acid exchange D495N) and clo-f2.103 (G280D) resulted in a total lack of Chl b, whereas in the missense mutants clo-f2.107 (P419L), clo-f2.109 (A94T), clo-f2.122 (C320Y), clo-f2.123 (A94T), clo-f2.133 (A376V) and clo-f2.181 (L373F) intermediate contents of Chl b were determined. The missense mutations affect conserved residues, and their effect on chlorophyllide a oxygenase is discussed. The mutations in clo-f2.102, clo-f2.103, clo-f2.133 and clo-f2.181 may influence electron transfer as illustrated in the active site of a structural model protein. The changes in clo-f2.107, clo-f2.109, clo-f2.122 and clo-f2.123 may lead to Chlb deficiency by interfering with the regulation of chlorophyllide a oxygenase. The correlation of mutations and phenotypes strongly supports that the barley locus fch2 encodes chlorophyllide a oxygenase.
    Plant and Cell Physiology 04/2012; 53(7):1232-46. DOI:10.1093/pcp/pcs062 · 4.98 Impact Factor
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    ABSTRACT: Time to flowering has an important impact on yield and has been a key trait in the domestication of crop plants and the spread of agriculture. In 1961, the cultivar Mari (mat-a.8) was the very first induced early barley (Hordeum vulgare L.) mutant to be released into commercial production. Mari extended the range of two-row spring barley cultivation as a result of its photoperiod insensitivity. Since its release, Mari or its derivatives have been used extensively across the world to facilitate short-season adaptation and further geographic range extension. By exploiting an extended historical collection of early-flowering mutants of barley, we identified Praematurum-a (Mat-a), the gene responsible for this key adaptive phenotype, as a homolog of the Arabidopsis thaliana circadian clock regulator Early Flowering 3 (Elf3). We characterized 87 induced mat-a mutant lines and identified >20 different mat-a alleles that had clear mutations leading to a defective putative ELF3 protein. Expression analysis of HvElf3 and Gigantea in mutant and wild-type plants demonstrated that mat-a mutations disturb the flowering pathway, leading to the early phenotype. Alleles of Mat-a therefore have important and demonstrated breeding value in barley but probably also in many other day-length-sensitive crop plants, where they may tune adaptation to different geographic regions and climatic conditions, a critical issue in times of global warming.
    Proceedings of the National Academy of Sciences 02/2012; 109(11):4326-31. DOI:10.1073/pnas.1113009109 · 9.81 Impact Factor
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    ABSTRACT: The structure of the major light-harvesting chlorophyll a/b complex (LHCII) was analyzed by pulsed EPR measurements and compared with the crystal structure. Site-specific spin labeling of the recombinant protein allowed the measurement of distance distributions over several intra- and intermolecular distances in monomeric and trimeric LHCII, yielding information on the protein structure and its local flexibility. A spin label rotamer library based on a molecular dynamics simulation was used to take the local mobility of spin labels into account. The core of LHCII in solution adopts a structure very similar or identical to the one seen in crystallized LHCII trimers with little motional freedom as indicated by narrow distance distributions along and between α helices. However, distances comprising the lumenal loop domain show broader distance distributions, indicating some mobility of this loop structure. Positions in the hydrophilic N-terminal domain, upstream of the first trans-membrane α helix, exhibit more and more mobility the closer they are to the N terminus. The nine amino acids at the very N terminus that have not been resolved in any of the crystal structure analyses give rise to very broad and possibly bimodal distance distributions, which may represent two families of preferred conformations.
    Journal of Biological Chemistry 01/2012; 287(4):2915-2925. · 4.57 Impact Factor
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    ABSTRACT: The structure of the major light-harvesting chlorophyll a/b complex (LHCII) was analyzed by pulsed EPR measurements and compared with the crystal structure. Site-specific spin labeling of the recombinant protein allowed the measurement of distance distributions over several intra- and intermolecular distances in monomeric and trimeric LHCII, yielding information on the protein structure and its local flexibility. A spin label rotamer library based on a molecular dynamics simulation was used to take the local mobility of spin labels into account. The core of LHCII in solution adopts a structure very similar or identical to the one seen in crystallized LHCII trimers with little motional freedom as indicated by narrow distance distributions along and between α helices. However, distances comprising the lumenal loop domain show broader distance distributions, indicating some mobility of this loop structure. Positions in the hydrophilic N-terminal domain, upstream of the first trans-membrane α helix, exhibit more and more mobility the closer they are to the N terminus. The nine amino acids at the very N terminus that have not been resolved in any of the crystal structure analyses give rise to very broad and possibly bimodal distance distributions, which may represent two families of preferred conformations.
    Journal of Biological Chemistry 12/2011; 287(4):2915-25. DOI:10.1074/jbc.M111.307728 · 4.57 Impact Factor
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    ABSTRACT: Compared to folding of soluble proteins, folding of membrane proteins is complicated by the fact that it requires an amphiphilic environment. Few existing techniques can provide structurally resolved information on folding kinetics. For the major plant light harvesting complex LHCII, it is demonstrated that changes in water accessibility of a particular amino acid residue can be followed during folding by measuring the hyperfine interaction of spin labels with deuterium nuclei of heavy water. The incorporation of residue 196 into the hydrophobic core of a detergent micelle was investigated. The technique provides a time constant that is similar to the one found with fluorescence spectroscopy for the slower folding step of the whole protein and with electron paramagnetic resonance for change of the distance between residues 90 and 196. If applied to several residues, this technique should provide information on the sequence of events during membrane protein folding.
    Journal of Physical Chemistry Letters 02/2010; 1(3). DOI:10.1021/jz900424n · 7.46 Impact Factor
  • Harald Paulsen · Christoph Dockter · Aleksei Volkov · Gunnar Jeschke
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    ABSTRACT: The major light-harvesting chlorophyll a/b protein (LHCIIb) is one of the most abundant proteins of the chloroplast in green plants. It contains roughly half of the chlorophylls involved in photosynthesis, and exhibits an unusual ability to self-organize in vitro. Simply mixing the apoprotein, native or recombinant, with its pigments, chlorophyll a, chlorophyll b, and xanthophylls, in detergent solution, suffices to trigger protein folding and the assembly of about 18 pigments in their correct binding sites. A study of the mechanism of this self-organization seems worthwhile since (1) our knowledge about membrane protein folding is scarce compared to what we know about the folding of water-soluble proteins, (2) the mechanism of LHCIIb formation in vitro may give useful clues about the so-far unknown pathway of its assembly in the chloroplast, and (3) a thorough understanding of the process may facilitate the application of recombinant LHCIIb in hybrid constructs such as photovoltaic devices or the construction of potentially useful proteins or other polymers that spontaneously bind other dyes at a similarly high density. During the assembly of recombinant LHCIIb, the formation of protein secondary structure is triggered by the binding of pigments. Chlorophylls are bound in two apparent kinetic phases. A faster one in the range of tens of seconds reflects the binding of chlorophyll a along with xanthophylls. During the slower step in the range of minutes, mostly chlorophyll b is bound. The intermediate complex lacking chlorophyll b is unstable and susceptible to proteases. The resulting two-step model of LHCIIb formation is able to explain why LHCIIb in vivo exhibits an apparently constant chlorophyll a:b ratio although several binding sites have been found to be accessible to both of the two chlorophylls in vitro and in plants over-producing chlorophyll b. Time-resolved electron paramagnetic resonance (EPR) techniques have been established to assess protein folding beyond secondary structure formation during LHCIIb assembly, and first results of EPR-monitored kinetics are shown.
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    ABSTRACT: The major light-harvesting chlorophyll a/b complex (LHCII) of the photosynthetic apparatus in plants self-organizes in vitro. The recombinant apoprotein, denatured in dodecyl sulfate, spontaneously folds when it is mixed with its pigments, chlorophylls, and carotenoids in detergent solution, and assembles into structurally authentic LHCII in the course of several minutes. Pulse EPR techniques, specifically double-electron-electron resonance (DEER), have been used to analyze protein folding during this process. Pairs of nitroxide labels were introduced site-specifically into recombinant LHCII and shown not to affect the stability and function of the pigment-protein complex. Interspin distance distributions between two spin pairs were measured at various time points, one pair located on either end of the second transmembrane helix (helix 3), the other one located near the luminal ends of the intertwined transmembrane helices 1 and 4. In the dodecyl sulfate-solubilized apoprotein, both distance distributions were consistent with a random-coil protein structure. A rapid freeze-quench experiment on the latter spin pair indicated that 1 s after initiating reconstitution the protein structure is virtually unchanged. Subsequently, both distance distributions monitored protein folding in the same time range in which the assembly of chlorophylls into the complex had been observed. The positioning of the spin pair spanning the hydrophobic core of LHCII clearly preceded the juxtaposition of the spin pair on the luminal side of the complex. This indicates that superhelix formation of helices 1 and 4 is a late step in LHCII assembly.
    Proceedings of the National Academy of Sciences 11/2009; 106(44):18485-90. DOI:10.1073/pnas.0906462106 · 9.81 Impact Factor
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    A Volkov · C Dockter · T Bund · H Paulsen · G Jeschke
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    ABSTRACT: Membrane proteins reside in a structured environment in which some of their residues are accessible to water, some are in contact with alkyl chains of lipid molecules, and some are buried in the protein. Water accessibility of residues may change during folding or function-related structural dynamics. Several techniques based on the combination of pulsed electron paramagnetic resonance (EPR) with site-directed spin labeling can be used to quantify such water accessibility. Accessibility parameters for different residues in major plant light-harvesting complex IIb are determined by electron spin echo envelope modulation spectroscopy in the presence of deuterated water, deuterium contrast in transversal relaxation rates, analysis of longitudinal relaxation rates, and line shape analysis of electron-spin-echo-detected EPR spectra as well as by the conventional techniques of measuring the maximum hyperfine splitting and progressive saturation in continuous-wave EPR. Systematic comparison of these parameters allows for a more detailed characterization of the environment of the spin-labeled residues. These techniques are applicable independently of protein size and require approximately 10-20 nmol of singly spin-labeled protein per sample. For a residue close to the N-terminus, in a domain unresolved in the existing x-ray structures of light-harvesting complex IIb, all methods indicate high water accessibility.
    Biophysical Journal 03/2009; 96(3):1124-41. DOI:10.1016/j.bpj.2008.09.047 · 3.97 Impact Factor
  • Christoph Dockter · Mats Hansson · Damian Gruszka
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    ABSTRACT: Plant height and culm sturdiness are quantitative characters important for securing cereal crop yield in adverse weather conditions with strong winds and torrential rain. Brassinosteroids, a class of growth-promoting hormones, are known to regulate culm architecture and height in plants. We recently identified a unique combination of phenotypic characters, including short and sturdy straw, specific for brassinosteroid deficiency in the barley BRASSINOSTEROID-INSENSITIVE 1 (BRI1) mutant uzu1.a. Identical combinations of phenotypes (short culm, dense spike basis, erect leaves, and undulated leaf margins) were found in wild type barley grown in presence of the brassinosteroid inhibitor propiconazole. A comprehensive phenotyping screen of historic barley mutant collections identified more than 20 morphologically identical mutants in the barley complementation groups BRACHYTIC, BREVIARISTATUM and ERECTOIDES. In silico genome mapping of brassinosteroid biosynthesis and signaling genes in combination with re-sequencing of historic barley mutants, identified HvBRI1, HvBRD (BRASSINOSTEROID-6-OXIDASE), HvCPD (CONSTITUTIVE PHOTOMORPHOGENIC DWARF) and HvDIM (DIMINUTO DWARF) as major controllers of plant height and culm robustness in barley. The identified mutants represent alternatives to the widely used but highly temperature-sensitive uzu1.a allele of HvBRI1 for the development of sturdy and climate-tolerant barley cultivars.
    International Plant and Animal Genome Conference XXII 2014;
  • Christoph Dockter · Damian Gruszka · Mats Hansson
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    ABSTRACT: Mutants are excellent tools for gaining insights into biochemical pathways, enzymatic reaction mechanisms, signaling networks and molecular processes in general. The Scandinavian barley (Hordeum vulgare L.) mutant collection located at the Nordic Genetic Resource Center, Sweden, is one of the largest barley mutant collections. With more than 11,000 entries, it is a fantastic resource for molecular studies connected to traits of applied and basic scientific interest. In our research, we are especially interested in mutants with differences in day-length response, maturity, ripeness, chloroplast development, general plant size, culm length, culm stiffness and kernel size. The talk will focus on our recent results related to identification of genes causing a dwarf or semi-dwarf phenotype. Mutations in these genes are of importance for breeding of lodging resistant crop plants, an issue of growing importance in times of global warming. Brassinosteroids effect cell elongation. Therefore, mutants deficient in genes encoding brassinosteroid receptors and brassinosteroid biosynthetic enzymes show a short-culm phenotype. We screened a collection of dwarf and semi-dwarf barley mutants for their response to exogenous brassinolide in a so-called leaf-unrolling test. We identified novel alleles of Uzu as well as new barley genes deficient in brassinosteroid mutants, which might be of importance for breeding of robust and lodging resistant barley cultivars.
    International Plant and Animal Genome Conference XXI 2013;