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A systems genetics study of seed quality and seedling vigour in Brassica rapa
Basnet, R.K. - \ 2015
University. Promotor(en): Richard Visser, co-promotor(en): Guusje Bonnema; Chris Maliepaard. - Wageningen : Wageningen University - ISBN 9789462574250 - 177
brassica campestris - zaadkwaliteit - groeikracht - zaden - zaadontwikkeling - zaadkieming - zaailingen - plantengenetica - genomica - seed quality - vigour - seeds - seed development - seed germination - seedlings - plant genetics - genomics
Seed is the basic and most critical input for seed propagated agricultural crops: seed quality and seedling vigour determine plant establishment, growth and development in both natural and agricultural ecosystems. Seed quality and seedling vigour are mainly determined by the interactions of the following three components: genetic background, physiological quality and the environmental conditions during seed set, seed ripening, storage, seed germination and early seedling development. In the past, many efforts have been made to improve seed germination and seedling vigour by optimizing physiological and environmental factors (non-genetic factors); however, the paradigm has shifted to investigate genetic factors and to use these to improve crop performance by plant breeding. The aim of this thesis is to unravel the genetics of seed germination and seedling vigour under different conditions in Brassica rapa, using a systems genetics approach. Studies in many crop species have reported that seed germination and seedling vigour traits are governed by many genes and are strongly affected by environmental conditions. As salinity stress is becoming one of the most important abiotic stresses affecting crop growth and yield, we studied the genetics of seed germination and seedling vigour under neutral and salt stress conditions. For a number of crops, it has been established that larger seed size and higher seed weight indicate more reserve food and contribute positively to seedling establishment. Therefore, our hypothesis for this thesis is that transcriptional regulation of genes during seed development determines the composition and content of seed reserves, and that these seed reserves play a major role in seed germination and seedling growth, especially at the heterotrophic stage under optimal and sub-optimal conditions.
B. rapa is an extremely diverse Brassica species which includes, besides many diverse leafy vegetable types and turnips, also oilseed crops. Brassica seeds are of high economic importance for several reasons. They are the starting point of the life cycle of the crop, but also they are directly used as sources of vegetable oil or condiments. At present, B. napus is the most important source of vegetable oil worldwide, but B. rapa is often used for introgression breeding to broaden its narrow genetic base resulting in genetic improvements. Therefore, the acquired knowledge is also useful for the scientific community and plant breeders working in B. napus and other Brassica species.
In Chapter 2 we evaluated the genetic diversity of a B. rapa core collection of 168 accessions representing different crop types and geographic origins. Using the Bayesian cluster analysis software STRUCTURE, we identified four subpopulations: subpopulation 1 with accessions of Indian origin, spring oil, yellow sarson and rapid cycling; subpopulation 2 consisting of several types from Asian origins: pak choi, winter oil, mizuna, mibuna, komasuna, turnip green, oil rape and Asian turnip; subpopulation 3, which included mainly accessions of Chinese cabbage and subpopulation 4 with mostly vegetable turnip, fodder turnip and brocoletto accessions from European origin. The geographical distribution of the accessions was very much congruent with genetic, metabolic and morphological diversity. This initial study was followed by association studies for secondary metabolites from the tocopherol and carotenoids pathways, using the population structure of these four subpopulations as a correction term to control for spurious marker-trait associations (Chapter 2). Additionally, we used a machine learning approach, Random Forest (RF) regression, to find marker-trait associations. We chose the RF approach as it can handle large numbers of variables (markers, metabolites, transcript abundance) in combination with relatively small sample sets of accessions, to show its perspectives for application to the increasing amounts of data available through the different ~omics technologies. In our analysis, the markers showing significant association with metabolites identified by the RF approach overlapped with markers obtained from association mapping. Those markers could potentially be used for marker-assisted selection (MAS) in breeding for these secondary metabolites in different morphotypes or sub-populations. Knowledge of genetic distance as evaluated in this chapter allowed the choice of parents to create a segregating population for QTL analyses by maximizing genetic variation between the parents.
In Chapter 4, a doubled haploid (DH) population from a cross of genetically diverse morphotypes of B. rapa, an oil-type yellow sarson (YS143) and a vegetable pak choi (PC175) (Chapter 2), was used to evaluate the genetic basis of seed germination and seedling vigour traits under both non-stress and salt stress conditions. The yellow sarson parent had larger seed size and higher thousand-seed weight than the pak choi parent, and displayed earlier onset, higher uniformity in germination, faster germination and maximum germination, and higher root- and shoot- lengths and biomass under both non-stress and salt stress conditions. Positive correlations of thousand-seed weight with earliness, speed and uniformity of germination and maximum germination percentage, supports that larger seeds germinate earlier, faster, more uniformly and to a higher maximum germination percentage than smaller seeds. Thus, we conclude that yellow sarson had higher seed quality and seedling vigour than pak choi. However yellow sarson also contributed negative alleles to seed germination, as illustrated by its allele of the QTL at A05 which decreases the uniformity of seed germination. In addition we also observed that yellow sarson seedling growth was more affected by salt stress than pak choi. All traits were scored over the DH population, and this clearly showed transgressive variation for most traits. Eight QTL hotspots were identified for seed weight, seed germination, and root and shoot lengths. A QTL hotspot for seed germination on A02 co-located with a homologue of the FLOWERING LOCUS C (BrFLC2) genes and its cis-acting expression QTL (cis-eQTL). FLC2 (BrFLC2 in B. rapa) is an important repressor of flowering time in both A. thaliana and B. rapa and recently, FLC2 was reported for its pleiotropic effect on seed germination in A. thaliana. A QTL hotspot on A05 with salt stress specific QTL co-located with the FATTY ACID DESATURASE 2 (BrFAD2) gene and its cis-eQTL. Besides the role of FAD2 in fatty acid desaturation, the up-regulation of this gene was associated with enhanced seed germination and hypocotyl elongation under salinity in B. napus (BnFAD2) and A. thaliana (FAD2). We observed epistatic interactions between the QTL hotspots at the BrFLC2 and BrFAD2 loci, and between other QTL hotspots.
Seed development is regulated by many dynamic metabolic processes controlled by complex networks of spatially and temporally expressed genes. Therefore, morphological characteristics and the transcriptional signatures of developing seeds from yellow- and brown/black-seeded genotypes were studied to get to know the timing of key metabolic processes, to explore the major transcriptional differences and to identify the optimum stage for a genetical genomics study for B. rapa seed traits (Chapter 3). This is the first study of genome-wide profiling of transcript abundance during seed development in B. rapa. Most transcriptional changes occurred between 25 and 35 days after pollination (between the bent-cotyledon stage and the stage when the embryo fully fills the seed), which is later than in the related species B. napus. A weighted gene co-expression network analysis (WGCNA) identified 47 gene modules with different co-expression patterns, of which 17 showed a genotype effect, 4 modules a time effect during seed development and 6 modules both genotype and time effects. Based on the number of genes in gene modules, the predominant variation in gene expression was according to developmental stages rather than morphotype differences. We identified 17 putative cis-regulatory elements (motifs) for four co-regulated gene clusters of genes related to lipid metabolism. The identification of key physiological events, major expression patterns, and putative cis-regulatory elements provides useful information to construct gene regulatory networks in B. rapa developing seeds and provides a starting point for a genetical genomics study of fatty acid composition and additional seed traits in Chapter 5.
Since Brassica seeds are sources of vegetable oil, genetic studies of the gene regulatory mechanisms underlying lipid metabolism is of high importance, not only in relation to seed and seedling vigour, but also for Brassica oilseed breeding. In Chapter 5, an integrative approach of QTL mapping for fatty acids composition and for transcript abundance (eQTL) of genes related to lipid metabolism, together with gene co-expression networks was used to unravel the genetic regulation of seed fatty acid composition in the DH population of B. rapa. In this study, a confounding effect of flowering time variation was observed on fatty acid QTLs (metabolite level) at linkage group A02 and of seed colour variation on eQTLs (transcript level) at linkage group A09. At A02, fatty acid QTLs from 2009 seeds co-locate with the genetic position of a gene-targeted marker for BrFLC2, its cis-QTL, and a major flowering time QTL. Flowering time variation is very obvious in this DH population and the BrFLC2 gene at A02 (16.7 cM) is the major regulator of flowering time, with a non-functional allele in the yellow sarson parent. When QTL analysis was performed on seeds from 2011, from DH lines that flowered synchronously due to staggered sowing, this fatty acid QTL hotspot disappeared. The 2011 seed lot was used for further analysis combining fatty acid QTLs with eQTLs in this study. On A09, a large trans-eQTL hotspot was co-localized with a major seed colour QTL, in the region where the causal gene, the bHLH transcription factor BrTT8, was cloned. The role of this gene in seed colour development was functionally proven in B. rapa. As the yellow sarson and pak choi parents of this population have contrasting seed coat colour (Chapter 3) the DH lines segregated for seed colour. When seed colour variation was used as a co-variate in our statistical model, we could exclude its confounding effect on eQTL mapping. We compared the fatty acid QTL and eQTL results from the analyses before and after seed colour correction and later discuss the results from the analysis after correction. The distribution of major QTLs for fatty acids showed a relationship with the types of fatty acids: linkage group A03 contained major QTLs for monounsaturated fatty acids (MUFAs), A04 for saturated fatty acids (SFAs) and A05 for polyunsaturated fatty acids (PUFAs). Using a genetical genomics approach, eQTL hotspots were found at major fatty acid QTLs on A03, A04 and A05 and on A09. Finally, an eQTL-guided gene co-expression network of lipid metabolism related genes showed major hubs at the genes BrPLA2-ALPHA, BrWD-40, a number of seed storage protein genes and a transcription factor BrMD-2, suggesting essential roles of these genes in lipid metabolism. Several genes, such as BrFAE1, BrTAG1, BrFAD2, BrFAD5, BrFAD7, which were reported as important genes for fatty acid composition in seeds in other studies of related species, had relatively lower degrees of connection in the networks. However their cis-eQTLs co-localized with specific fatty acid QTLs, making them candidate genes for the observed variation. We hypothesize that these play a role in modifying fatty acid content or composition across genotypes, rather than playing essential roles in the pathway itself. These results suggest the need of a global study of lipid metabolism rather than a strict focus on the fatty acid biosynthesis pathway per se. This study gives a starting point for understanding the genetic regulation of lipid metabolism, by identification of a number of key regulatory genes, identified as major hub genes, and candidate genes for fatty acid QTLs.
In the final chapter (Chapter 6) we summarize and critically discuss the relationships among phenotypic traits, metabolites and expression variation as well as the co-localization of QTLs from these different levels. In this thesis, we developed methodology to integrate transcriptomics and metabolomics data sets and to construct gene regulatory networks related to major fatty acids, and found a set of (possible) candidate genes involved in lipid metabolism. In the future, we recommend to integrate the genome-wide transcriptome data set with all major seed metabolites and phenotypic data on seed and seedling vigour to directly link all three components: transcriptome, metabolome and phenotypic traits, and ultimately expand the knowledge on the genetic regulation of seed metabolites, seed quality and seedling vigour in B. rapa to other Brassica species.
Supporting Local Seed Businesses : A Training Manual for ISSD Uganda
Mastenbroek, A. ; Chebet, A. ; Muwanika, C.T. ; Adong, C.J. ; Okot, F. ; Otim, G. ; Birungi, J. ; Kansiime, M. ; Oyee, P. ; Ninsiima, P. - \ 2015
Wageningen : Centre for Development Innovation, Wageningen UR - 273
seed production - seed development - seed quality - rural development - farming - markets - businesses - small businesses - regional development - training courses - training - agricultural development - uganda - west africa - africa - zaadproductie - zaadontwikkeling - zaadkwaliteit - plattelandsontwikkeling - landbouw bedrijven - markten - bedrijven - kleine bedrijven - regionale ontwikkeling - scholingscursussen - opleiding - landbouwontwikkeling - west-afrika - afrika
The training manual is developed in Uganda to train partner organisations in coaching farmer groups to become sustainable local seed businesses. It introduces the Integrated Seed Sector Development Programme in Uganda and the concept of local seed businesses (LSBs). The manual has 5 modules covering selection, monitoring and sustaining local seed businesses; technically equipping local seed businesses, professionally organising LSBs; orienting LSBs to the market and strategically linking them to service providers.
Fruit illumination stimulates cell division but has no detectable effect on fruit size in tomato (Solanum lycopersicum)
Okello, R.C. ; Heuvelink, E. ; Visser, P.H.B. de; Lammers, M. ; Maagd, R.A. de; Marcelis, L.F.M. ; Struik, P.C. - \ 2015
Physiologia Plantarum 154 (2015)1. - ISSN 0031-9317 - p. 114 - 127.
arabidopsis-thaliana - elongation growth - plant development - seed development - sink metabolism - cucumber fruits - gene-expression - hormone-levels - abscisic-acid - phytochrome-b
Light affects plant growth through assimilate availability and signals regulating development. The effects of light on growth of tomato fruit were studied using cuvettes with light-emitting diodes providing white, red or blue light to individual tomato trusses for different periods during daytime. Hypotheses tested were as follows: (1) light-grown fruits have stronger assimilate sinks than dark-grown fruits, and (2) responses depend on light treatment provided, and fruit development stage. Seven light treatments [dark, 12-h white, 24-h white, 24-h red and 24-h blue light, dark in the first 24 days after anthesis (DAA) followed by 24-h white light until breaker stage, and its reverse] were applied. Observations were made between anthesis and breaker stage at fruit, cell and gene levels. Fruit size and carbohydrate content did not respond to light treatments while cell division was strongly stimulated at the expense of cell expansion by light. The effects of light on cell number and volume were independent of the combination of light color and intensity. Increased cell division and decreased cell volume when fruits were grown in the presence of light were not clearly corroborated by the expression pattern of promoters and inhibitors of cell division and expansion analyzed in this study, implying a strong effect of posttranscriptional regulation. Results suggest the existence of a complex homeostatic regulatory system for fruit growth in which reduced cell division is compensated by enhanced cell expansion.
A roadmap to embryo identity in plants
Radoeva, T.M. ; Weijers, D. - \ 2014
Trends in Plant Science 19 (2014)11. - ISSN 1360-1385 - p. 709 - 716.
microspore-derived embryos - somatic embryogenesis - arabidopsis-thaliana - pattern-formation - transcription factor - leafy cotyledon1 - seed development - gene-expression - wuschel gene - cell
Although plant embryogenesis is usually studied in the context of seed development, there are many alternative roads to embryo initiation. These include somatic embryogenesis in tissue culture and microspore embryogenesis, both widely used in breeding and crop propagation, but also include other modes of ectopic embryo initiation. In the past decades several genes, mostly transcription factors, were identified that can induce embryogenesis in somatic cells. Because the genetic networks in which such regulators operate to promote embryogenesis are largely unknown, a key question is how their activity relates to zygotic and alternative embryo initiation. We describe here the many roads to plant embryo initiation and discuss a framework for defining the developmental roles and mechanisms of plant embryogenesis regulators.
Local Auxin Sources Orient the Apical-Basal Axis in Arabidopsis Embryos
Robert, H.S. ; Grones, P. ; Stepanova, A.N. ; Robles, L.M. ; Lokerse, A.S. ; Alonso, J.M. ; Weijers, D. ; Friml, J. - \ 2013
Current Biology 23 (2013)24. - ISSN 0960-9822 - p. 2506 - 2512.
plant development - cotyledon development - seed development - pin proteins - biosynthesis - gene - polarity - embryogenesis - expression - efflux
Establishment of the embryonic axis foreshadows the main body axis of adults both in plants and in animals, but underlying mechanisms are considered distinct. Plants utilize directional, cell-to-cell transport of the growth hormone auxin [1 and 2] to generate an asymmetric auxin response that specifies the embryonic apical-basal axis [3, 4, 5 and 6]. The auxin flow directionality depends on the polarized subcellular localization of PIN-FORMED (PIN) auxin transporters [7 and 8]. It remains unknown which mechanisms and spatial cues guide cell polarization and axis orientation in early embryos. Herein, we provide conceptually novel insights into the formation of embryonic axis in Arabidopsis by identifying a crucial role of localized tryptophan-dependent auxin biosynthesis [ 9, 10, 11 and 12]. Local auxin production at the base of young embryos and the accompanying PIN7-mediated auxin flow toward the proembryo are required for the apical auxin response maximum and the specification of apical embryonic structures. Later in embryogenesis, the precisely timed onset of localized apical auxin biosynthesis mediates PIN1 polarization, basal auxin response maximum, and specification of the root pole. Thus, the tight spatiotemporal control of distinct local auxin sources provides a necessary, non-cell-autonomous trigger for the coordinated cell polarization and subsequent apical-basal axis orientation during embryogenesis and, presumably, also for other polarization events during postembryonic plant life [ 13 and 14].
Different Auxin Response Machineries Control Distinct Cell Fates in the Early Plant Embryo
Rademacher, E.H. ; Lokerse, A.S. ; Schlereth, A. ; Llavata Peris, C.I. ; Bayer, M. ; Kientz, M. ; Freire Rios, A. ; Borst, J.W. ; Lukowitz, W. ; Juergens, G. ; Weijers, D. - \ 2012
Developmental cell 22 (2012)1. - ISSN 1534-5807 - p. 211 - 222.
arabidopsis embryo - transcription factor - seed development - expression map - root-meristem - aux/iaa genes - proteins - monopteros - family - transformation
The cell types of the plant root are first specified early during embryogenesis and are maintained throughout plant life. Auxin plays an essential role in embryonic root initiation, in part through the action of the ARF5/MP transcription factor and its auxin-labile inhibitor IAA12/BDL. MP and BDL function in embryonic cells but promote auxin transport to adjacent extraembryonic suspensor cells, including the quiescent center precursor (hypophysis). Here we show that a cell-autonomous auxin response within this cell is required for root meristem initiation. ARF9 and redundant ARFs, and their inhibitor IAA10, act in suspensor cells to mediate hypophysis specification and, surprisingly, also to prevent transformation to embryo identity. ARF misexpression, and analysis of the short suspensor mutant, demonstrates that lineage-specific expression of these ARFs is required for normal embryo development. These results imply the existence of a prepattern for a cell-type-specific auxin response that underlies the auxin-dependent specification of embryonic cell types
Multi-dimensional regulation of metabolic networks shaping plant development and performance
Kooke, R. ; Keurentjes, J.J.B. - \ 2012
Journal of Experimental Botany 63 (2012)9. - ISSN 0022-0957 - p. 3353 - 3365.
natural genetic-variation - shade-avoidance response - arabidopsis-thaliana - circadian clock - secondary metabolites - enzyme-activities - seed development - glucosinolate accumulation - carbohydrate-metabolism - laser microdissection
The metabolome is an integral part of a plant’s life cycle and determines for a large part its external phenotype. It is the final, internal product of chemical interactions, obtained through developmental, genetic, and environmental inputs, and as such, it defines the state of a plant in terms of development and performance. Understanding its regulation will provide knowledge and new insights into the biochemical pathways and genetic interactions that shape the plant and its surroundings. In this review, we will focus on four dimensions that contribute to the huge diversity of metabolomes and we will illustrate how this diversity shapes the plant in terms of development and performance: (i) temporal regulation: the metabolome is extremely dynamic and temporal changes in the environment can have an immense impact on its composition; (ii) spatial regulation: metabolites can be very specific, in both quantitative and qualitative terms, to specialized organs, tissues, and cell types; (iii) environmental regulation: the metabolic profile of plants is highly dependent on environmental signals, such as light, temperature, and nutrients, and very susceptible to biotic and abiotic stresses; and (iv) genetic regulation: the biosynthesis, structure, and accumulation of metabolites have a genetic origin, and there is quantitative and qualitative variation for metabolomes within a species. We will address the contribution of these dimensions to the wide diversity of metabolomes and highlight how the multi-dimensional regulation of metabolism defines the plant’s phenotype
Transcriptome and Metabolite Profiling Show That APETALA2a Is a Major Regulator of Tomato Fruit Ripening
Karlova, R.B. ; Rosin, F.M.A. ; Busscher-Lange, J. ; Parapunova, V.A. ; Do, P.T. ; Fernie, A.R. ; Fraser, P.D. ; Baxter, C. ; Angenent, G.C. ; Maagd, R.A. de - \ 2011
The Plant Cell 23 (2011)3. - ISSN 1040-4651 - p. 923 - 941.
homeotic gene apetala2 - ethylene biosynthesis - flower development - 1-aminocyclopropane-1-carboxylate synthase - chromoplast differentiation - lycopersicon-esculentum - expression analysis - arabidopsis flower - seed development - organ identity
Fruit ripening in tomato (Solanum lycopersicum) requires the coordination of both developmental cues as well as the plant hormone ethylene. Although the role of ethylene in mediating climacteric ripening has been established, knowledge regarding the developmental regulators that modulate the involvement of ethylene in tomato fruit ripening is still lacking. Here, we show that the tomato APETALA2a (AP2a) transcription factor regulates fruit ripening via regulation of ethylene biosynthesis and signaling. RNA interference (RNAi)-mediated repression of AP2a resulted in alterations in fruit shape, orange ripe fruits, and altered carotenoid accumulation. Microarray expression analyses of the ripe AP2 RNAi fruits showed altered expression of genes involved in various metabolic pathways, such as the phenylpropanoid and carotenoid pathways, as well as in hormone synthesis and perception. Genes involved in chromoplast differentiation and other ripening-associated processes were also differentially expressed, but softening and ethylene biosynthesis occurred in the transgenic plants. Ripening regulators RIPENING-INHIBITOR, NON-RIPENING, and COLORLESS NON-RIPENING (CNR) function upstream of AP2a and positively regulate its expression. In the pericarp of AP2 RNAi fruits, mRNA levels of CNR were elevated, indicating that AP2a and CNR are part of a negative feedback loop in the regulation of ripening. Moreover, we demonstrated that CNR binds to the promoter of AP2a in vitro
The conserved splicing factor SUA controls alternative splicing of the developmental regulator ABI3 in Arabidopsis.
Sugliani, M. ; Brambilla, V. ; Clerkx, E.J.M. ; Koornneef, M. ; Soppe, W.J.J. - \ 2010
The Plant Cell 22 (2010). - ISSN 1040-4651 - p. 1936 - 1946.
rna-binding proteins - messenger-rna - higher-plants - seed development - gene-regulation - factor u2af - thaliana - complex - domain - aba
ABSCISIC ACID INSENSITIVE3 (ABI3) is a major regulator of seed maturation in Arabidopsis thaliana. We detected two ABI3 transcripts, ABI3- and ABI3-ß, which encode full-length and truncated proteins, respectively. Alternative splicing of ABI3 is developmentally regulated, and the ABI3-ß transcript accumulates at the end of seed maturation. The two ABI3 transcripts differ by the presence of a cryptic intron in ABI3-, which is spliced out in ABI3-ß. The suppressor of abi3-5 (sua) mutant consistently restores wild-type seed features in the frameshift mutant abi3-5 but does not suppress other abi3 mutant alleles. SUA is a conserved splicing factor, homologous to the human protein RBM5, and reduces splicing of the cryptic ABI3 intron, leading to a decrease in ABI3-ß transcript. In the abi3-5 mutant, ABI3-ß codes for a functional ABI3 protein due to frameshift restoration
The MADS domain protein DIANA together with AGAMOUS-LIKE80 to specify the central cell in Arabidopsis ovules
Bemer, M. ; Wolters-Arts, M. ; Grossniklaus, U. ; Angenent, G.C. - \ 2008
The Plant Cell 20 (2008). - ISSN 1040-4651 - p. 2088 - 2101.
female gametophyte development - polycomb group gene - seed development - box gene - endosperm development - wild-type - thaliana - identification - fertilization - expression
MADS box genes in plants consist of MIKC-type and type I genes. While MIKC-type genes have been studied extensively, the functions of type I genes are still poorly understood. Evidence suggests that type I MADS box genes are involved in embryo sac and seed development. We investigated two independent T-DNA insertion alleles of the Arabidopsis thaliana type I MADS box gene AGAMOUS-LIKE61 (AGL61) and showed that in agl61 mutant ovules, the polar nuclei do not fuse and central cell morphology is aberrant. Furthermore, the central cell begins to degenerate before fertilization takes place. Although pollen tubes are attracted and perceived by the mutant ovules, neither endosperm development nor zygote formation occurs. AGL61 is expressed in the central cell during the final stages of embryo sac development. An AGL61:green fluorescent protein¿-glucoronidase fusion protein localizes exclusively to the polar nuclei and the secondary nucleus of the central cell. Yeast two-hybrid analysis showed that AGL61 can form a heterodimer with AGL80 and that the nuclear localization of AGL61 is lost in the agl80 mutant. Thus, AGL61 and AGL80 appear to function together to differentiate the central cell in Arabidopsis. We renamed AGL61 DIANA, after the virginal Roman goddess of the hunt.
Sucrose Synthase Is Associated with the Cell Wall of Tobacco Pollen Tubes
Persia, D. ; Cai, G. ; Casino, C. ; Willemse, M.T.M. ; Cresti, M. - \ 2008
Plant Physiology 147 (2008)4. - ISSN 0032-0889 - p. 1603 - 1618.
nicotiana-alata link - japanese pear fruit - membrane association - callose synthase - cellulose synthesis - brefeldin-a - protein-phosphorylation - starch biosynthesis - lilium-longiflorum - seed development
Sucrose synthase (Sus; EC 18.104.22.168) is a key enzyme of sucrose metabolism in plant cells, providing carbon for respiration and for the synthesis of cell wall polymers and starch. Since Sus is important for plant cell growth, insights into its structure, localization, and features are useful for defining the relationships between nutrients, growth, and cell morphogenesis. We used the pollen tube of tobacco (Nicotiana tabacum) as a cell model to characterize the main features of Sus with regard to cell growth and cell wall synthesis. Apart from its role during sexual reproduction, the pollen tube is a typical tip-growing cell, and the proper construction of its cell wall is essential for correct shaping and direction of growth. The outer cell wall layer of pollen tubes consists of pectins, but the inner layer is composed of cellulose and callose; both polymers require metabolic precursors in the form of UDP-glucose, which is synthesized by Sus. We identified an 88-kD polypeptide in the soluble, plasma membrane and Golgi fraction of pollen tubes. The protein was also found in association with the cell wall. After purification, the protein showed an enzyme activity similar to that of maize (Zea mays) Sus. Distribution of Sus was affected by brefeldin A and depended on the nutrition status of the pollen tube, because an absence of metabolic sugars in the growth medium caused Sus to distribute differently during tube elongation. Analysis by bidimensional electrophoresis indicated that Sus exists as two isoforms, one of which is phosphorylated and more abundant in the cytoplasm and cell wall and the other of which is not phosphorylated and is specific to the plasma membrane. Results indicate that the protein has a role in the construction of the extracellular matrix and thus in the morphogenesis of pollen tubes.
Detailed Analysis of the Expression of an Alpha-gliadin Promoter and the Deposition of Alpha-gliadin Protein During Wheat Grain Development
Herpen, T.W.J.M. van; Riley, M. ; Sparks, C. ; Jones, H.D. ; Gritsch, C. ; Dekking, E.H. ; Hamer, R.J. ; Bosch, H.J. ; Salentijn, E.M.J. ; Smulders, M.J.M. ; Shewry, P.R. ; Gilissen, L.J.W.J. - \ 2008
Annals of Botany 102 (2008)3. - ISSN 0305-7364 - p. 331 - 342.
endosperm-specific expression - celiac-disease - activates transcription - transgenic wheat - gene-expression - functional-analysis - barley endosperm - seed development - gcn4-like motif - bzip protein
Background and Aims: Alpha-gliadin proteins are important for the industrial quality of bread wheat flour, but they also contain many epitopes that can trigger celiac (c¿liac) disease (CD). The B-genome-encoded -gliadin genes, however, contain very few epitopes. Controlling -gliadin gene expression in wheat requires knowledge on the processes of expression and deposition of -gliadin protein during wheat grain development. Methods: A 592-bp fragment of the promotor of a B-genome-encoded -gliadin gene driving the expression of a GUS reporter gene was transformed into wheat. A large number of transgenic lines were used for data collection. GUS staining was used to determine GUS expression during wheat kernel development, and immunogold labelling and tissue printing followed by staining with an -gliadin-specific antibody was used to detect -gliadin protein deposited in developing wheat kernels. The promoter sequence was screened for regulatory motifs and compared to other available -gliadin promoter sequences. Key Results: GUS expression was detected primarily in the cells of the starchy endosperm, notably in the subaleurone layer but also in the aleurone layer. The -gliadin promoter was active from 11 days after anthesis (DAA) until maturity, with an expression similar to that of a 326-bp low molecular weight (LMW) subunit gene promoter reported previously. An -gliadin-specific antibody detected -gliadin protein in protein bodies in the starchy endosperm and in the subaleurone layer but, in contrast to the promoter activity, no -gliadin was detected in the aleurone cell layer. Sequence comparison showed differences in regulatory elements between the promoters of -gliadin genes originating from different genomes (A and B) of bread wheat both in the region used here and upstream. Conclusions: The results suggest that additional regulator elements upstream of the promoter region used may specifically repress expression in the aleurone cell layer. Observed differences in expression regulator motifs between the -gliadin genes on the different genomes (A and B) of bread wheat leads to a better understanding how -gliadin expression can be controlled
Embryonic factor 1 encodes an AMP deaminase and is essential for the zygote to embryo transition in Arabidopsis
Xu, J. ; Zhang, H.Y. ; Xue, H.W. ; Dijkhuis, P. ; Liu, C.M. - \ 2005
The Plant Journal (2005). - ISSN 0960-7412
polycomb group gene - seed development - early embryogenesis - egg activation - expression - protein - fertilization - thaliana - cells - transformation
Fusion of the egg and the sperm cells in plants produces a zygote that develops into an embryo. Screening of ethyl methanesulfonate-mutagenized populations of Arabidopsis led to the identification of EMBRYONIC FACTOR 1 (FAC1), a locus that gives a zygote-lethal phenotype when mutated. The FAC1 gene was identified by positional cloning and confirmed by a genetic complementation test against a T-DNA insertion allele. It encodes an AMP deaminase (AMPD) that is known in human and yeast to convert AMP to IMP to maintain the energy potential. Expression of FAC1 in a yeast AMPD mutant after removal of its N-terminal putative transmembrane domain complemented the mutant phenotype, suggesting a functional conservancy but a structural divergence through evolution. Although a low level of FAC1 expression was observed in all organs tested, using a reporter construct we observed a significantly increased FAC1 expression in the zygote, early embryo and endosperm. Furthermore, during somatic embryogenesis, a high level of FAC1 expression was observed in developing embryos including putative embryogenic cells. FAC1, therefore, represents one of the earliest expressed genes known in plants. It may act through AMP depletion to provide sufficient energy for the zygote to proceed through development.
AtGA3ox2, a key gene responsible for bioactive gibberellin biosynthesis, is regulated during embryogenesis by LEAFY COTYLEDON2 and FUSCA3 in Arabidopsis
Curaba, J. ; Moritz, T. ; Blervaque, R. ; Parcy, F. ; Raz, V. ; Herzog, M. ; Vachon, G. - \ 2004
Plant Physiology 136 (2004)3. - ISSN 0032-0889 - p. 3660 - 3669.
late embryo development - thaliana l heynh - abscisic-acid - seed development - leafy cotyledon1 - transcription factor - fus3 gene - mutants - germination - expression
Embryonic regulators LEC2 (LEAFY COTYLEDON2) and FUS3 (FUSCA3) are involved in multiple aspects of Arabidopsis (Arabidopsis thaliana) seed development, including repression of leaf traits and premature germination and activation of seed storage protein genes. In this study, we show that gibberellin (GA) hormone biosynthesis is regulated by LEC2 and FUS3 pathways. The level of bioactive GAs is increased in immature seeds of lec2 and fus3 mutants relative to wild-type level. In addition, we show that the formation of ectopic trichome cells on lec2 and fus3 embryos is a GA-dependent process as in true leaves, suggesting that the GA pathway is misactivated in embryonic mutants. We next demonstrate that the GA-biosynthesis gene AtGA3ox2, which encodes the key enzyme AtGA3ox2 that catalyzes the conversion of inactive to bioactive GAs, is ectopically activated in embryos of the two mutants. Interestingly, both B-glucuronidase reporter gene expression and in situ hybridization indicate that FUS3 represses AtGA3ox2 expression mainly in epidermal cells of embryo axis, which is distinct from AtGA3ox2 pattern at germination. Finally, we show that the FUS3 protein physically interacts with two RY elements (CATGCATG) present in the AtGA3ox2 promoter. This work suggests that GA biosynthesis is directly controlled by embryonic regulators during Arabidopsis embryonic development.
Transcript profiling of transcription factor genes during silique development in Arabidopsis
Folter, S. de; Lange, J. ; Colombo, L. ; Losa, A. ; Angenent, G.C. - \ 2004
Plant Molecular Biology 56 (2004)3. - ISSN 0167-4412 - p. 351 - 366.
mads-box genes - shoot apical meristem - fruit-development - ovule development - seed development - domain protein - zinc-finger - plant - expression - thaliana
Flower development is a key process for all angiosperms and is essential for sexual reproduction. The last phase in flower development is fertilization of the ovules and formation of the fruits, which are both biologically and economically of importance. Here, we report the expression profiles of over 1100 unique Arabidopsis genes coding for known and putative transcription factors (TFs) during silique development using high-density filter array hybridizations. Hierarchical cluster analyses revealed distinct expression profiles for the different silique developmental stages. This allowed a functional classification of these expression profiles in groups, namely pistil development, embryogenesis, seed maturation, fruit maturation, and fruit development. A further focus was made on the MADS-box family, which contains many members that are functionally well-characterized. The expression profiles of these MADS-box genes during silique development give additional clues on their functions and evolutionary relationship.
In situ FTIR assessment of desiccation-tolerant tissues
Wolkers, W.F. ; Hoekstra, F.A. - \ 2003
Spectroscopy: an international journal 17 (2003). - ISSN 0712-4813 - p. 297 - 313.
transform infrared-spectroscopy - protein secondary structure - electron-paramagnetic-resonance - typha-latifolia l - plant craterostigma-plantagineum - carrot somatic embryos - abscisic-acid - arabidopsis-thaliana - phase-transitions - seed development
This essay shows how Fourier transform infrared (FTIR) microspectroscopy can be applied to study thermodynamic parameters and conformation of endogenous biomolecules in desiccation-tolerant biological tissues. Desiccation tolerance is the remarkable ability of some organisms to survive complete dehydration. Seed and pollen of higher plants are well known examples of desiccation-tolerant tissues. FTIR studies on the overall protein secondary structure indicate that during the acquisition of desiccation tolerance, plant embryos exhibit proportional increases in alpha-helical structures and that beta-sheet structures dominate upon drying of desiccation sensitive-embryos. During ageing of pollen and seeds, the overall protein secondary structure remains stable, whereas drastic changes in the thermotropic response of membranes occur, which coincide with a complete loss of viability. Properties of the cytoplasmic glassy matrix in desiccation-tolerant plant organs can be studied by monitoring the position of the OH-stretching vibration band of endogenous carbohydrates and proteins as a function of temperature. By applying these FTIR techniques to maturation-defective mutant seeds of Arabidopsis thaliana we were able to establish a correlation between macromolecular stability and desiccation tolerance. Taken together, in situ FTIR studies can give unique information on conformation and stability of endogenous biomolecules in desiccation-tolerant tissues.
Sexual and apomictic reproduction in Hieracium sub genus Pilosella are closely interrelated developmental pathways
Tucker, M.R. ; Araujo, A.C.G. ; Paech, N.A. ; Hecht, V. ; Schmidt, E.D.L. ; Rossel, J.B. ; Vries, S.C. de; Koltunow, A.M. - \ 2003
The Plant Cell 15 (2003). - ISSN 1040-4651 - p. 1524 - 1537.
endosperm development - arabidopsis-thaliana - pattern-formation - ovule development - seed development - apomixis - expression - gene - embryo - fertilization
Seed formation in flowering plants requires meiosis of the megaspore mother cell (MMC) inside the ovule, selection of a megaspore that undergoes mitosis to form an embryo sad, and double fertilization to initiate embryo and endosperm formation. During apomixis, or asexual seed formation, in Hieracium ovules, a somatic aposporous initial (All) cell divides to form a structurally variable aposporous embryo sac and embryo. This entire process, including endosperm development, is fertilization independent. Introduction of reproductive tissue marker genes into sexual and apomictic Hieracium showed that Al cells do not express a MMC marker. Spatial and temporal gene expression patterns of other introduced genes were conserved commencing with the first nuclear division of the Al cell in apomicts and the mitotic initiation of embryo sac formation in sexual plants. Conservation in expression patterns also occurred during embryo and endosperm development, indicating that sexuality and apomixis are interrelated pathways that share regulatory components. The induction of a modified sexual reproduction program in Al cells may enable the manifestation of apomixis in Hieracium.
Toward the analysis of the Petunia MADS box gene family by reverse and forward transposon insertion mutagenesis approaches: B, C, and D Floral organ identity functions require SEPALLATA-Like MADS box genes in Petunia
Vandenbussche, M. ; Zethof, J. ; Souer, E. ; Koes, R. ; Tornielli, G.B. ; Pezzotti, M. ; Ferrario, S.I.T. ; Angenent, G.C. ; Gerats, T. - \ 2003
The Plant Cell 15 (2003). - ISSN 1040-4651 - p. 2680 - 2693.
protein-protein interactions - transcription factor family - flower development - antirrhinum-majus - meristem identity - ovule development - seed development - homeotic genes - arabidopsis - plants
We have initiated a systematic functional analysis of the MADS box, intervening region, K domain, C domain-type MADS box gene family in petunia. The starting point for this has been a reverse-genetics approach, aiming to select for transposon insertions into any MADS box gene. We have developed and applied a family signature insertion screening protocol that is highly suited for this purpose, resulting in the isolation of 32 insertion mutants in 20 different MADS box genes. In addition, we identified three more MADS box gene insertion mutants using a candidate-gene approach. The defined insertion lines provide a sound foundation for a systematic functional analysis of the MADS box gene family in petunia. Here, we focus on the analysis of Floral Binding Protein2 (FBP2) and FBP5 genes that encode the E-function, which in Arabidopsis has been shown to be required for B and C floral organ identity functions. fbp2 mutants display sepaloid petals and ectopic inflorescences originating from the third floral whorl, whereas fbp5 mutants appear as wild type. In fbp2 fbp5 double mutants, reversion of floral organs to leaf-like organs is increased further. Strikingly, ovules are replaced by leaf-like structures in the carpel, indicating that in addition to the B- and C-functions, the D-function, which specifies ovule development, requires E-function activity. Finally, we compare our data with results obtained using cosuppression approaches and conclude that the latter might be less suited for assigning functions to individual members of the MADS box gene family
Seed dormancy and germination
Bentsink, L. ; Koornneef, M. - \ 2002
In: Unknown - p. 1 - 18.
arabidopsis thaliana - kiemrust - zaadkieming - zaadontwikkeling - seed dormancy - seed germination - seed development
Arabidopsis possesses dormancy, as is the case for many other plant species, which is controlled by environmental factors such as light, temperature and time of dry storage as well as by genetic factors. The use of genetics and molecular genetics in Arabidopsis is starting to shed light on some aspects of the mechanism of dormancy and germination by the identification of mutants and genes that control these processes. This review provides an overview of current knowledge of factors and genes controlling seed dormancy and germination in Arabidopsis
Chlorophyll in tomato seeds: marker for seed performance?
Suhartanto, M.R. - \ 2002
University. Promotor(en): L.H.W. van der Plas; Henk Hilhorst; H. Jalink. - S.l. : S.n. - ISBN 9789058086679 - 149
solanum lycopersicum - tomaten - zaadontwikkeling - zaadnarijping - zaadkwaliteit - chlorofyl - merkers - fluorescentie - zaden - tomatoes - seeds - seed development - seed maturation - seed quality - chlorophyll - markers - fluorescence
<p>Using Xe-PAM, laser induced fluorometry and high performance liquid chromatography we found that chlorophyll was present in young tomato (cv. Moneymaker) seeds and was degraded during maturation. Fluorescence microscopy and imaging showed that the majority of chlorophyll is located in the seed coat but low levels of chlorophyll were also detected in the embryo, mainly in the radicle tip. Seed chlorophyll fluorescence appeared to be a sensitive indicator of physiological maturity of tomato seeds. The chlorophyll in young tomato seeds is physiologically active and functional. Quantum yield, Rubisco activity and photosynthetic oxygen evolution were observable <u>in vitro</u> until 40 days after flowering. The role of hormones (abscisic acid and gibberellins) and phytochrome was discussed in relation to the occurrence of chlorophyll in tomato seeds. Seeds with low chlorophyll content during early growth may have low quality but the presence of chlorophyll during maturation is highly undesirable since it is associated with lower quality, particularly seed longevity.