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Sample records for arabidopsis cyp51a2 mutant

  1. Arabidopsis MET1 cytosine methyltransferase mutants.

    PubMed Central

    Kankel, Mark W; Ramsey, Douglas E; Stokes, Trevor L; Flowers, Susan K; Haag, Jeremy R; Jeddeloh, Jeffrey A; Riddle, Nicole C; Verbsky, Michelle L; Richards, Eric J

    2003-01-01

    We describe the isolation and characterization of two missense mutations in the cytosine-DNA-methyltransferase gene, MET1, from the flowering plant Arabidopsis thaliana. Both missense mutations, which affect the catalytic domain of the protein, led to a global reduction of cytosine methylation throughout the genome. Surprisingly, the met1-2 allele, with the weaker DNA hypomethylation phenotype, alters a well-conserved residue in methyltransferase signature motif I. The stronger met1-1 allele caused late flowering and a heterochronic delay in the juvenile-to-adult rosette leaf transition. The distribution of late-flowering phenotypes in a mapping population segregating met1-1 indicates that the flowering-time phenotype is caused by the accumulation of inherited defects at loci unlinked to the met1 mutation. The delay in flowering time is due in part to the formation and inheritance of hypomethylated fwa epialleles, but inherited defects at other loci are likely to contribute as well. Centromeric repeat arrays hypomethylated in met1-1 mutants are partially remethylated when introduced into a wild-type background, in contrast to genomic sequences hypomethylated in ddm1 mutants. ddm1 met1 double mutants were constructed to further our understanding of the mechanism of DDM1 action and the interaction between two major genetic loci affecting global cytosine methylation levels in Arabidopsis. PMID:12663548

  2. Mutants of Arabidopsis thaliana with altered phototropism

    NASA Technical Reports Server (NTRS)

    Khurana, J. P.; Poff, K. L.

    1989-01-01

    Thirty five strains of Arabidopsis thaliana (L.) Heynh. have been identified with altered phototropic responses to 450-nm light. Four of these mutants have been more thoroughly characterized. Strain JK224 shows normal gravitropism and "second positive" phototropism. However, while the amplitude for "first positive" phototropism is the same as that in the wild-type, the threshold and fluence for the maximum response in "first positive" phototropism are shifted to higher fluence by a factor of 20-30. This mutant may represent an alteration in the photoreceptor pigment for phototropism. Strain JK218 exhibits no curvature to light at any fluence from 1 micromole m-2 to 2700 micromoles m-2, but shows normal gravitropism. Strain JK345 shows no "first positive" phototropism, and reduced gravitropism and "second positive" phototropism. Strain JK229 shows no measurable "first positive" phototropism, but normal gravitropism and "second positive" phototropism. Based on these data, it is suggested that: 1. gravitropism and phototropism contain at least one common element; 2. "first positive" and "second positive" phototropism contain at least one common element; and 3. "first positive" phototropism can be substantially altered without any apparent alteration of "second positive" phototropism.

  3. Imaging lipid droplets in Arabidopsis mutants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Confocal fluorescence microscopy was adapted for the imaging of neutral lipids in plant leaves with defects in normal lipid metabolism using two different fluorescent dyes. Disruptions in a gene locus, At4g24160, yielded Arabidopsis thaliana plants with a preponderance of oil bodies in their leaves ...

  4. An annotated database of Arabidopsis mutants of acyl lipid metabolism

    SciTech Connect

    McGlew, Kathleen; Shaw, Vincent; Zhang, Meng; Kim, Ryeo Jin; Yang, Weili; Shorrosh, Basil; Suh, Mi Chung; Ohlrogge, John

    2014-12-10

    Mutants have played a fundamental role in gene discovery and in understanding the function of genes involved in plant acyl lipid metabolism. The first mutant in Arabidopsis lipid metabolism (fad4) was described in 1985. Since that time, characterization of mutants in more than 280 genes associated with acyl lipid metabolism has been reported. This review provides a brief background and history on identification of mutants in acyl lipid metabolism, an analysis of the distribution of mutants in different areas of acyl lipid metabolism and presents an annotated database (ARALIPmutantDB) of these mutants. The database provides information on the phenotypes of mutants, pathways and enzymes/proteins associated with the mutants, and allows rapid access via hyperlinks to summaries of information about each mutant and to literature that provides information on the lipid composition of the mutants. Mutants for at least 30 % of the genes in the database have multiple names, which have been compiled here to reduce ambiguities in searches for information. Furthermore, the database should also provide a tool for exploring the relationships between mutants in acyl lipid-related genes and their lipid phenotypes and point to opportunities for further research.

  5. An annotated database of Arabidopsis mutants of acyl lipid metabolism

    DOE PAGES

    McGlew, Kathleen; Shaw, Vincent; Zhang, Meng; ...

    2014-12-10

    Mutants have played a fundamental role in gene discovery and in understanding the function of genes involved in plant acyl lipid metabolism. The first mutant in Arabidopsis lipid metabolism (fad4) was described in 1985. Since that time, characterization of mutants in more than 280 genes associated with acyl lipid metabolism has been reported. This review provides a brief background and history on identification of mutants in acyl lipid metabolism, an analysis of the distribution of mutants in different areas of acyl lipid metabolism and presents an annotated database (ARALIPmutantDB) of these mutants. The database provides information on the phenotypes ofmore » mutants, pathways and enzymes/proteins associated with the mutants, and allows rapid access via hyperlinks to summaries of information about each mutant and to literature that provides information on the lipid composition of the mutants. Mutants for at least 30 % of the genes in the database have multiple names, which have been compiled here to reduce ambiguities in searches for information. Furthermore, the database should also provide a tool for exploring the relationships between mutants in acyl lipid-related genes and their lipid phenotypes and point to opportunities for further research.« less

  6. Architectural phenotypes in the transparent testa mutants of Arabidopsis thaliana

    PubMed Central

    Buer, Charles S.; Djordjevic, Michael A.

    2009-01-01

    Flavonoids are low molecular weight secondary plant metabolites with a myriad of functions. As flavonoids affect auxin transport (an important growth-controlling hormone) and are biologically active in eukaryotes, flavonoid mutants were expected to have undescribed architectural phenotypes. The Arabidopsis thaliana transparent testa (tt) mutants are compromised in the enzymatic steps or transcriptional regulators affecting flavonoid synthesis. tt mutant seedlings were grown on hard-slanted agar (a stress condition), under varying light conditions, and in soil to examine the resulting growth patterns. These tt mutants revealed a wide variety of architectural phenotypes in root and aerial tissues. Mutants with increased inflorescences, siliques, and lateral root density or reduced stature are traits that could affect plant yield or performance under certain environmental conditions. The regulatory genes affected in architectural traits may provide useful molecular targets for examination in other plants. PMID:19129166

  7. Mutants of Arabidopsis as tools for physiology and molecular biology

    SciTech Connect

    Somerville, C.R.; Artus, N.; Browse, J.; Caspar, T.; Estelle, M.; Haughn, G.; Kunst, L.; Martinez, J.; McCourt, P.; Moffatt, B.

    1986-04-01

    The authors discuss the importance of developing a facile system for genetic analysis in higher plants which can be used to approach problems specific to plant biology in much the same way that molecular genetic approaches have been used in other classes of organisms such as yeast and Drosophila. Toward this end, they have developed methods for the isolation and analysis of mutants of Arabidopsis with specific alterations in photosynthesis, photorespiration, starch metabolism, lipid metabolism, purine metabolism, amino acid metabolism and phytohormone responses. The utility of this collection of mutants for studying problems in physiology and biochemistry is illustrated with selected examples.

  8. Mutants of Arabidopsis with altered regulation of starch degradation

    SciTech Connect

    Caspar, T.; Lin, Tsanpiao; Kakefuda, G.; Benbow, L.; Preiss, J.; Somerville, C. )

    1991-04-01

    Mutants of Arabidopsis thaliana (L.) Heynh. with altered regulation of starch degradation were identified by screening for plants that retained high levels of leaf starch after a period of extended darkness. The mutant phenotype was also expressed in seeds, flowers, and roots, indicating that the same pathway of starch degradation is used in these tissues. In many respects, the physiological consequences of the mutations were equivalent to the effects observed in previously characterized mutants of Arabidopsis that are unable to synthesize starch. One mutant line, which was characterized in detail, had normal levels of activity of the starch degradative enzymes {alpha}-amylase, {beta}-amylase, phosphorylase, D-enzyme, and debranching enzyme. Thus, it was not possible to establish a biochemical basis for the phenotype, which was due to a recessive mutant at a locus designated sex 1 at position 12.2 on chromosome 1. This raises the possibility that hitherto unidentified factors, altered by the mutation, play a key role in regulating or catalyzing starch degradation.

  9. Gravitropism in roots of intermediate-starch mutants of Arabidopsis

    NASA Technical Reports Server (NTRS)

    Kiss, J. Z.; Wright, J. B.; Caspar, T.

    1996-01-01

    Gravitropism was studied in roots of wild type (WT) Arabidopsis thaliana (L.) Heynh. (strain Wassilewskija) and three starch-deficient mutants that were generated by T-DNA insertional mutagenesis. One of these mutants was starchless while the other two were intermediate mutants, which had 51% and 60%, respectively, of the WT amount of starch as determined by light and electron microscopy. The four parameters used to assay gravitropism were: orientation during vertical growth, time course of curvature, induction, and intermittent stimulation experiments. WT roots were much more responsive to gravity than were roots of the starchless mutant, and the intermediate starch mutants exhibited an intermediate graviresponse. Our data suggest that lowered starch content in the mutants primarily affects gravitropism rather than differential growth because both phototropic curvature and growth rates were approximately equal among all four genotypes. Since responses of intermediate-starch mutants were closer to the WT response than to the starchless mutant, it appears that 51-60% of the WT level of starch is near the threshold amount needed for full gravitropic sensitivity. While other interpretations are possible, the data are consistent with the starch statolith hypothesis for gravity perception in that the degree of graviresponsiveness is proportional to the total mass of plastids per cell.

  10. Characterization of Sugar Insensitive (sis) Mutants of Arabidopsis

    SciTech Connect

    Gibson, Susan I.

    2009-06-08

    Despite the fact that soluble sugar levels have been postulated to play an important role in the control of a wide variety of plant metabolic and developmental pathways, the mechanisms by which plants respond to soluble sugar levels remain poorly understood. Plant responses to soluble sugar levels are also important in bioenergy production, as plant sugar responses are believed to help regulate both carbon fixation and carbon partitioning. For example, accumulation of soluble sugars, such as sucrose and glucose, in source tissues leads to feedback inhibition of photosynthesis, thereby decreasing rates of carbon fixation. Soluble sugar levels can also affect sink strengths, affecting the rates of accumulation of carbon-based compounds into both particular molecular forms (e.g. carbohydrates versus lipids versus proteins) and particular plant organs and tissues. Mutants of Arabidopsis that are defective in the ability to respond to soluble sugar levels were isolated and used as tools to identify some of the factors involved in plant sugar response. These sugar insensitive (sis) mutants were isolated by screening mutagenized seeds for those that were able to germinate and develop relatively normal shoot systems on media containing 0.3 M glucose or 0.3 M sucrose. At these sugar concentrations, wild-type Arabidopsis germinate and produce substantial root systems, but show little to no shoot development. Twenty-eight sis mutants were isolated during the course of four independent mutant screens. Based on a preliminary characterization of all of these mutants, sis3 and sis6 were chosen for further study. Both of these mutations appear to lie in previously uncharacterized loci. Unlike many other sugar-response mutants, sis3 mutants exhibit a wild-type or near wild-type response in all phytohormone-response assays conducted to date. The sis6-1 mutation is unusual in that it appears to be due to overexpression of a gene, rather than representing a loss of function mutation

  11. Flavonoid accumulation patterns of transparent testa mutants of arabidopsis

    NASA Technical Reports Server (NTRS)

    Peer, W. A.; Brown, D. E.; Tague, B. W.; Muday, G. K.; Taiz, L.; Murphy, A. S.

    2001-01-01

    Flavonoids have been implicated in the regulation of auxin movements in Arabidopsis. To understand when and where flavonoids may be acting to control auxin movement, the flavonoid accumulation pattern was examined in young seedlings and mature tissues of wild-type Arabidopsis. Using a variety of biochemical and visualization techniques, flavonoid accumulation in mature plants was localized in cauline leaves, pollen, stigmata, and floral primordia, and in the stems of young, actively growing inflorescences. In young Landsberg erecta seedlings, aglycone flavonols accumulated developmentally in three regions, the cotyledonary node, the hypocotyl-root transition zone, and the root tip. Aglycone flavonols accumulated at the hypocotyl-root transition zone in a developmental and tissue-specific manner with kaempferol in the epidermis and quercetin in the cortex. Quercetin localized subcellularly in the nuclear region, plasma membrane, and endomembrane system, whereas kaempferol localized in the nuclear region and plasma membrane. The flavonoid accumulation pattern was also examined in transparent testa mutants blocked at different steps in the flavonoid biosynthesis pathway. The transparent testa mutants were shown to have precursor accumulation patterns similar to those of end product flavonoids in wild-type Landsberg erecta, suggesting that synthesis and end product accumulation occur in the same cells.

  12. Arabidopsis Mutant bik1 Exhibits Strong Resistance to Plasmodiophora brassicae

    PubMed Central

    Chen, Tao; Bi, Kai; He, Zhangchao; Gao, Zhixiao; Zhao, Ying; Fu, Yanping; Cheng, Jiasen; Xie, Jiatao; Jiang, Daohong

    2016-01-01

    Botrytis-induced kinase1 (BIK1), a receptor-like cytoplasmic kinase, plays an important role in resistance against pathogens and insects in Arabidopsis thaliana. However, it remains unknown whether BIK1 functions against Plasmodiophora brassicae, an obligate biotrophic protist that attacks cruciferous plants and induces gall formation on roots. Here, we investigated the potential roles of receptors FLS2, BAK1, and BIK1 in the infection of P. brassicae cruciferous plants. Wild-type plants, fls2, and bak1 mutants showed typical symptom on roots, and the galls were filled with large quantities of resting spores, while bik1 mutant plants exhibited strong resistance to P. brassicae. Compared with that of the wild-type plants, the root hair and cortical infection rate of bik1 mutant were significantly reduced by about 40–50%. A considerable portion of bik1 roots failed to form typical galls. Even if some small galls were formed, they were filled with multinucleate secondary plasmodia. The bik1 plants accumulated less reactive oxygen species (ROS) at infected roots than other mutants and wild-type plants. Exogenous salicylic acid (SA) treatment alleviated the clubroot symptoms in wild-type plants, and the expression of the SA signaling marker gene PR1 was significantly increased in bik1. Both sid2 (salicylic acid induction-deficient 2) and npr1-1 [non-expresser of PR genes that regulate systemic acquired resistance (SAR)] mutants showed increased susceptibility to P. brassicae compared with wild-type plants. These results suggest that the resistance of bik1 to P. brassicae is possibly mediated by SA inducible mechanisms. PMID:27679580

  13. Distinct Phyllosphere Bacterial Communities on Arabidopsis Wax Mutant Leaves

    PubMed Central

    Reisberg, Eva E.; Hildebrandt, Ulrich; Riederer, Markus; Hentschel, Ute

    2013-01-01

    The phyllosphere of plants is inhabited by diverse microorganisms, however, the factors shaping their community composition are not fully elucidated. The plant cuticle represents the initial contact surface between microorganisms and the plant. We thus aimed to investigate whether mutations in the cuticular wax biosynthesis would affect the diversity of the phyllosphere microbiota. A set of four Arabidopsis thaliana eceriferum mutants (cer1, cer6, cer9, cer16) and their respective wild type (Landsberg erecta) were subjected to an outdoor growth period and analysed towards this purpose. The chemical distinctness of the mutant wax phenotypes was confirmed by gas chromatographic measurements. Next generation amplicon pyrosequencing of the bacterial communities showed distinct community patterns. This observation was supported by denaturing gradient gel electrophoresis experiments. Microbial community analyses revealed bacterial phylotypes that were ubiquitously present on all plant lines (termed “core” community) while others were positively or negatively affected by the wax mutant phenotype (termed “plant line-specific“ community). We conclude from this study that plant cuticular wax composition can affect the community composition of phyllosphere bacteria. PMID:24223831

  14. A Mutant of Arabidopsis with Increased Levels of Stearic Acid.

    PubMed Central

    Lightner, J.; Wu, J.; Browse, J.

    1994-01-01

    A mutation at the fab2 locus of Arabidopsis caused increased levels of stearate in leaves. The increase in leaf stearate in fab2 varied developmentally, and the largest increase occurred in young leaves, where stearate accounted for almost 20% of total leaf fatty acids. The fatty acid composition of leaf lipids isolated from the fab2 mutant showed increased stearate in all the major glycerolipids of both the chloroplast and extrachloroplast membranes. Although the stearate content was increased, the fab2 mutant still contained abundant amounts of 18:1, 18:2, and 18:3 fatty acids. These results are consistent with the expectations for a mutation partially affecting the action of the stromal stearoyl-acyl carrier protein desaturase. Positional analysis indicated that the extra 18:0 is excluded with high specificity from the sn-2 position of both chloroplast and extrachloroplast glycerolipids. Although stearate content was increased in all the major leaf membrane lipids, the amount of increase varied considerably among the different lipids, from a high of 25% of fatty acids in phosphatidylcholine to a low of 2.9% of fatty acids in monogalactosyldiacylglycerol. PMID:12232421

  15. Inflorescence abnormalities occur with overexpression of Arabidopsis lyrata FT in the fwa mutant of Arabidopsis thaliana.

    PubMed

    Kawanabe, Takahiro; Fujimoto, Ryo

    2011-10-01

    Arabidopsis thaliana is a quantitative long-day plant with the timing of the floral transition being regulated by both endogenous signals and multiple environmental factors. fwa is a late-flowering mutant, and this phenotype is due to ectopic FWA expression caused by hypomethylation at the FWA locus. The floral transition results in the activation of the floral development process, the key regulators being the floral meristem identity genes, AP1 (APETALA1) and LFY (LEAFY). In this study, we describe inflorescence abnormalities in plants overexpressing the Arabidopsis lyrata FT (AlFT) and A. thaliana FWA (AtFWA) genes simultaneously. The inflorescence abnormality phenotype was present in only a proportion of plants. All plants overexpressing both AlFT and AtFWA flowered earlier than fwa, suggesting that the inflorescence abnormality and earlier flowering time are caused independently. The inflorescence abnormality phenotype was similar to that of the double mutant of ap1 and lfy, and AP1 and LFY genes were down-regulated in the abnormal inflorescences. From these results, we suggest that not only does ectopic AtFWA expression inhibit AtFT/AlFT function to delay flowering but that overexpression of AtFWA and AlFT together inhibits AP1 and LFY function to produce abnormal inflorescences.

  16. Regulation of chloroplast biogenesis: the immutans mutant of Arabidopsis

    SciTech Connect

    Rodermel, Steven

    2015-11-16

    The immutans (im) variegation mutant of Arabidopsis is an ideal model to gain insight into factors that control chloroplast biogenesis. im defines the gene for PTOX, a plastoquinol terminal oxidase that participates in control of thylakoid redox. Here, we report that the im defect can be suppressed during the late stages of plant development by gigantea (gi2), which defines the gene for GIGANTEA (GI), a central component of the circadian clock that plays a poorly-understood role in diverse plant developmental processes. imgi2 mutants are late-flowering and display other well-known phenotypes associated with gi2, such as starch accumulation and resistance to oxidative stress. We show that the restoration of chloroplast biogenesis in imgi2 is caused by a developmental-specific de-repression of cytokinin signaling that involves crosstalk with signaling pathways mediated by gibberellin (GA) and SPINDLY (SPY), a GA response inhibitor. Suppression of the plastid defect in imgi2 is likely caused by a relaxation of excitation pressures in developing plastids by factors contributed by gi2, including enhanced rates of photosynthesis and increased resistance to oxidative stress. Interestingly, the suppression phenotype of imgi can be mimicked by crossing im with the starch accumulation mutant, sex1, perhaps because sex1 utilizes pathways similar to gi. We conclude that our studies provide a direct genetic linkage between GIGANTEA and chloroplast biogenesis, and we construct a model of interactions between signaling pathways mediated by gi, GA, SPY, cytokinins, and sex1 that are required for chloroplast biogenesis.

  17. Phenotypic analysis of Arabidopsis mutants: quantitative analysis of root growth.

    PubMed

    Doerner, Peter

    2008-03-01

    INTRODUCTIONThe growth of plant roots is very easy to measure and is particularly straightforward in Arabidopsis thaliana, because the increase in organ size is essentially restricted to one dimension. The precise measurement of root apical growth can be used to accurately determine growth activity (the rate of growth at a given time) during development in mutants, transgenic backgrounds, or in response to experimental treatments. Root growth is measured in a number of ways, the simplest of which is to grow the seedlings in a Petri dish and record the position of the advancing root tip at appropriate time points. The increase in root length is measured with a ruler and the data are entered into Microsoft Excel for analysis. When dealing with large numbers of seedlings, however, this procedure can be tedious, as well as inaccurate. An alternative approach, described in this protocol, uses "snapshots" of the growing plants, which are taken using gel-documentation equipment (i.e., a video camera with a frame-grabber unit, now commonly used to capture images from ethidium-bromide-stained electrophoresis gels). The images are analyzed using publicly available software (NIH-Image), which allows the user simply to cut and paste data into Microsoft Excel.

  18. Plastid distribution in columella cells of a starchless Arabidopsis mutant grown in microgravity

    NASA Technical Reports Server (NTRS)

    Hilaire, E.; Paulsen, A. Q.; Brown, C. S.; Guikema, J. A.; Spooner, B. S. (Principal Investigator)

    1997-01-01

    Wild-type and starchless Arabidopsis thaliana mutant seedlings (TC7) were grown and fixed in the microgravity environment of a U.S. Space Shuttle spaceflight. Computer image analysis of longitudinal sections from columella cells suggest a different plastid positioning mechanism for mutant and wild-type in the absence of gravity.

  19. A double-mutant collection targeting MAP kinase related genes in Arabidopsis for studying genetic interactions.

    PubMed

    Su, Shih-Heng; Krysan, Patrick J

    2016-12-01

    Mitogen-activated protein kinase cascades are conserved in all eukaryotes. In Arabidopsis thaliana there are approximately 80 genes encoding MAP kinase kinase kinases (MAP3K), 10 genes encoding MAP kinase kinases (MAP2K), and 20 genes encoding MAP kinases (MAPK). Reverse genetic analysis has failed to reveal abnormal phenotypes for a majority of these genes. One strategy for uncovering gene function when single-mutant lines do not produce an informative phenotype is to perform a systematic genetic interaction screen whereby double-mutants are created from a large library of single-mutant lines. Here we describe a new collection of 275 double-mutant lines derived from a library of single-mutants targeting genes related to MAP kinase signaling. To facilitate this study, we developed a high-throughput double-mutant generating pipeline using a system for growing Arabidopsis seedlings in 96-well plates. A quantitative root growth assay was used to screen for evidence of genetic interactions in this double-mutant collection. Our screen revealed four genetic interactions, all of which caused synthetic enhancement of the root growth defects observed in a MAP kinase 4 (MPK4) single-mutant line. Seeds for this double-mutant collection are publicly available through the Arabidopsis Biological Resource Center. Scientists interested in diverse biological processes can now screen this double-mutant collection under a wide range of growth conditions in order to search for additional genetic interactions that may provide new insights into MAP kinase signaling.

  20. Cytochemical Analysis of Pollen Development in Wild-Type Arabidopsis and a Male-Sterile Mutant.

    PubMed Central

    Regan, SM; Moffatt, BA

    1990-01-01

    Microsporogenesis has been examined in wild-type Arabidopsis thaliana and the nuclear male-sterile mutant BM3 by cytochemical staining. The mutant lacks adenine phosphoribosyltransferase, an enzyme of the purine salvage pathway that converts adenine to AMP. Pollen development in the mutant began to diverge from wild type just after meiosis, as the tetrads of microspores were released from their callose walls. The first indication of abnormal pollen development in the mutant was a darker staining of the microspore wall due to an incomplete synthesis of the intine. Vacuole formation was delayed and irregular in the mutant, and the majority of the mutant microspores failed to undergo mitotic divisions. Enzyme activities of alcohol dehydrogenase and esterases decreased in the mutant soon after meiosis and were undetectable in mature pollen grains of the mutant. RNA accumulation was also diminished. These results are discussed in relation to the possible role(s) of adenine salvage in pollen development. PMID:12354970

  1. Sinapic acid ester metabolism in wild type and a sinapoylglucose-accumulating mutant of arabidopsis.

    PubMed Central

    Lorenzen, M; Racicot, V; Strack, D; Chapple, C

    1996-01-01

    Sinapoylmalate is one of the major phenylpropanoid metabolites that is accumulated in the vegetative tissue of Arabidopsis thaliana. A thin-layer chromatography-based mutant screen identified two allelic mutant lines that accumulated sinapoylglucose in their leaves in place of sinapoylmalate. Both mutations were found to be recessive and segregated as single Mendelian genes. These mutants define a new locus called SNG1 for sinapoylglucose accumulator. Plants that are homozygous for the sng1 mutation accumulate normal levels of malate in their leaves but lack detectable levels of the final enzyme in sinapate ester biosynthesis, sinapoylglucose:malate sinapoyltransferase. A study of wild-type and sng1 seedlings found that sinapic acid ester biosynthesis in Arabidopsis is developmentally regulated and that the accumulation of sinapate esters is delayed in sng1 mutant seedlings. PMID:8972602

  2. The phenotype of Arabidopsis thaliana det1 mutants suggest a role for cytokinins in greening

    SciTech Connect

    Chory, J.; Aguilar, N.; Peto, C.A.

    1990-01-01

    When grown in the absence of light, the det1 mutants of Arabidopsis thaliana develop characteristics of light-grown plants by morphological, cellular, and molecular criteria. Further, in light-grown plants, mutations in the DET1 gene affect cell-type-specific expression of light-regulated genes and the chloroplast developmental program. Here we show that the addition of exogenously added cytokinins (either 2-isopentenyl adenine, kinetin, or benzyladenine) to the growth medium of dark-germinated wild-type seedlings results in seedlings that resemble det1 mutants, instead of having the normal etiolated morphology. Like det1 mutants, these dark-grown seedlings now contain chloroplasts and have high levels of expression of genes that are normally light''-regulated. These results suggest an important role for cytokinins during greening of Arabidopsis, and may implicate cytokinin levels or an increased sensitivity to cytokinins as explanations for some of the observed phenotypes of det1 mutants.

  3. hydra Mutants of Arabidopsis Are Defective in Sterol Profiles and Auxin and Ethylene Signaling

    PubMed Central

    Souter, Martin; Topping, Jennifer; Pullen, Margaret; Friml, Jiri; Palme, Klaus; Hackett, Rachel; Grierson, Don; Lindsey, Keith

    2002-01-01

    The hydra mutants of Arabidopsis are characterized by a pleiotropic phenotype that shows defective embryonic and seedling cell patterning, morphogenesis, and root growth. We demonstrate that the HYDRA1 gene encodes a Δ8-Δ7 sterol isomerase, whereas HYDRA2 encodes a sterol C14 reductase, previously identified as the FACKEL gene product. Seedlings mutant for each gene are similarly defective in the concentrations of the three major Arabidopsis sterols. Promoter::reporter gene analysis showed misexpression of the auxin-regulated DR5 and ACS1 promoters and of the epidermal cell file–specific GL2 promoter in the mutants. The mutants exhibit enhanced responses to auxin. The phenotypes can be rescued partially by inhibition of auxin and ethylene signaling but not by exogenous sterols or brassinosteroids. We propose a model in which correct sterol profiles are required for regulated auxin and ethylene signaling through effects on membrane function. PMID:12034894

  4. Rubisco small subunits from the unicellular green alga Chlamydomonas complement Rubisco-deficient mutants of Arabidopsis.

    PubMed

    Atkinson, Nicky; Leitão, Nuno; Orr, Douglas J; Meyer, Moritz T; Carmo-Silva, Elizabete; Griffiths, Howard; Smith, Alison M; McCormick, Alistair J

    2017-04-01

    Introducing components of algal carbon concentrating mechanisms (CCMs) into higher plant chloroplasts could increase photosynthetic productivity. A key component is the Rubisco-containing pyrenoid that is needed to minimise CO2 retro-diffusion for CCM operating efficiency. Rubisco in Arabidopsis was re-engineered to incorporate sequence elements that are thought to be essential for recruitment of Rubisco to the pyrenoid, namely the algal Rubisco small subunit (SSU, encoded by rbcS) or only the surface-exposed algal SSU α-helices. Leaves of Arabidopsis rbcs mutants expressing 'pyrenoid-competent' chimeric Arabidopsis SSUs containing the SSU α-helices from Chlamydomonas reinhardtii can form hybrid Rubisco complexes with catalytic properties similar to those of native Rubisco, suggesting that the α-helices are catalytically neutral. The growth and photosynthetic performance of complemented Arabidopsis rbcs mutants producing near wild-type levels of the hybrid Rubisco were similar to those of wild-type controls. Arabidopsis rbcs mutants expressing a Chlamydomonas SSU differed from wild-type plants with respect to Rubisco catalysis, photosynthesis and growth. This confirms a role for the SSU in influencing Rubisco catalytic properties.

  5. An Arabidopsis thaliana embryo arrest mutant exhibiting germination potential

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The ability to initiate radicle elongation, or germination potential, occurs in developing embryos before the completion of seed maturation. Green embryos after walking-stick stage in developing Arabidopsis thaliana seeds germinate when excised from seeds and incubated in MS media containing 1 % suc...

  6. Comparative proteomic study of Arabidopsis mutants mpk4 and mpk6

    PubMed Central

    Takáč, Tomáš; Vadovič, Pavol; Pechan, Tibor; Luptovčiak, Ivan; Šamajová, Olga; Šamaj, Jozef

    2016-01-01

    Arabidopsis MPK4 and MPK6 are implicated in different signalling pathways responding to diverse external stimuli. This was recently correlated with transcriptomic profiles of Arabidopsis mpk4 and mpk6 mutants, and thus it should be reflected also on the level of constitutive proteomes. Therefore, we performed a shot gun comparative proteomic analysis of Arabidopsis mpk4 and mpk6 mutant roots. We have used bioinformatic tools and propose several new proteins as putative MPK4 and MPK6 phosphorylation targets. Among these proteins in the mpk6 mutant were important modulators of development such as CDC48A and phospholipase D alpha 1. In the case of the mpk4 mutant transcriptional reprogramming might be mediated by phosphorylation and change in the abundance of mRNA decapping complex VCS. Further comparison of mpk4 and mpk6 root differential proteomes showed differences in the composition and regulation of defense related proteins. The mpk4 mutant showed altered abundances of antioxidant proteins. The examination of catalase activity in response to oxidative stress revealed that this enzyme might be preferentially regulated by MPK4. Finally, we proposed developmentally important proteins as either directly or indirectly regulated by MPK4 and MPK6. These proteins contribute to known phenotypic defects in the mpk4 and mpk6 mutants. PMID:27324189

  7. Restoration of gravitropic sensitivity in starch-deficient mutants of Arabidopsis by hypergravity

    NASA Technical Reports Server (NTRS)

    Fitzelle, K. J.; Kiss, J. Z.

    2001-01-01

    Despite the extensive study of plant gravitropism, there have been few experiments which have utilized hypergravity as a tool to investigate gravisensitivity in flowering plants. Previous studies have shown that starch-deficient mutants of Arabidopsis are less sensitive to gravity compared to the wild-type (WT). In this report, the question addressed was whether hypergravity could restore the sensitivity of starch-deficient mutants of Arabidopsis. The strains examined include a WT, a starchless mutant and a reduced-starch mutant. Vertical orientation studies with dark-grown seedlings indicate that increased centrifugal acceleration improves orientation relative to the acceleration vector for all strains, even the WT. For starchless roots, growth of seedlings under constant 5 g acceleration was required to restore orientation to the level of the WT at 1 g. In contrast, approximately 10 g was required to restore the orientation of the starchless mutant hypocotyls to a WT level at 1 g. Examination of plastid position in root cap columella cells of the starchless mutant revealed that the restoration of gravitropic sensitivity was correlated with the sedimentation of plastids toward the distal cell wall. Even in WT plants, hypergravity caused greater sedimentation of plastids and improved gravitropic capability. Collectively, these experiments support the hypothesis of a statolith-based system of gravity perception in plants. As far as is known, this is the first report to use hypergravity to study the mechanisms of gravitropism in Arabidopsis.

  8. A comprehensive dataset of genes with a loss-of-function mutant phenotype in Arabidopsis.

    PubMed

    Lloyd, Johnny; Meinke, David

    2012-03-01

    Despite the widespread use of Arabidopsis (Arabidopsis thaliana) as a model plant, a curated dataset of Arabidopsis genes with mutant phenotypes remains to be established. A preliminary list published nine years ago in Plant Physiology is outdated, and genome-wide phenotype information remains difficult to obtain. We describe here a comprehensive dataset of 2,400 genes with a loss-of-function mutant phenotype in Arabidopsis. Phenotype descriptions were gathered primarily from manual curation of the scientific literature. Genes were placed into prioritized groups (essential, morphological, cellular-biochemical, and conditional) based on the documented phenotypes of putative knockout alleles. Phenotype classes (e.g. vegetative, reproductive, and timing, for the morphological group) and subsets (e.g. flowering time, senescence, circadian rhythms, and miscellaneous, for the timing class) were also established. Gene identities were classified as confirmed (through molecular complementation or multiple alleles) or not confirmed. Relationships between mutant phenotype and protein function, genetic redundancy, protein connectivity, and subcellular protein localization were explored. A complementary dataset of 401 genes that exhibit a mutant phenotype only when disrupted in combination with a putative paralog was also compiled. The importance of these genes in confirming functional redundancy and enhancing the value of single gene datasets is discussed. With further input and curation from the Arabidopsis community, these datasets should help to address a variety of important biological questions, provide a foundation for exploring the relationship between genotype and phenotype in angiosperms, enhance the utility of Arabidopsis as a reference plant, and facilitate comparative studies with model genetic organisms.

  9. Characterization of gravitropic inflorescence bending in brassinosteroid biosynthesis and signaling Arabidopsis mutants.

    PubMed

    Arteca, Richard N; Arteca, Jeannette M

    2011-07-15

    The interaction between the plant hormones, brassinosteroids and auxins has been documented in various processes using a variety of plants and plant parts. In this study, detached inflorescences from brassinosteroid biosynthesis and signaling Arabidopsis mutants were evaluated for their gravitropic bending in response to epibrassinolide (EBR) and indole-3-acetic acid (IAA). EBR supplied to the base of detached inflorescences stimulated gravitropic bending in all BR biosynthetic mutants but there was no effect on the BR signaling mutant or wild type plants. When IAA was supplied to the base of BR mutant inflorescences both natural and EBR-induced gravitropic bending was inhibited. Treatment with the auxin inhibitors also decreased both natural and EBR-induced gravitropic bending. No gravitropic bending was observed when the apical tips of BR mutant inflorescences were removed. IAA treatment to the tips of decapitated BR mutant inflorescences restored gravitropic bending to values observed in the inflorescences with an apical tip, however, EBR applied to the tip had no effect. When decapitated inflorescences from BR mutants were treated with IAA to the base and either gel, EBR or IAA was applied to the tip; there was no gravitropic bending. These results show that brassinosteroids have a role in the gravitropic bending response in Arabidopsis and mutants serve to uncover this hidden contributor.

  10. Cadmium-sensitive, cad1 mutants of Arabidopsis thaliana are phytochelatin deficient.

    PubMed Central

    Howden, R; Goldsbrough, P B; Andersen, C R; Cobbett, C S

    1995-01-01

    An allelic series of cad1, cadmium-sensitive mutants of Arabidopsis thaliana, was isolated. These mutants were sensitive to cadmium to different extents and were deficient in their ability to form cadmium-peptide complexes as detected by gel-filtration chromatography. Each mutant was deficient in its ability to accumulate phytochelatins (PCs) as detected by high-performance liquid chromatography and the amount of PCs accumulated by each mutant correlated with its degree of sensitivity to cadmium. The mutants had wild-type levels of glutathione, the substrate for PC biosynthesis, and in vitro assays demonstrated that each of the mutants was deficient in PC synthase activity. These results demonstrate conclusively the importance of PCs for cadmium tolerance in plants. PMID:7770517

  11. Purification of a. beta. -amylase that accumulates in Arabidopsis thaliana mutants defective in starch metabolism. [Arabidopsis thaliana

    SciTech Connect

    Monroe, J.D.; Preiss, J. )

    1990-11-01

    Amylase activity is elevated 5- to 10-fold in leaves of several different Arabidopsis thaliana mutants defective in starch metabolism when they are grown under a 12-hour photoperiod. Activity is also increased when plants are grown under higher light intensity. It was previously determined that the elevated activity was an extrachloroplastic {beta}-(exo)amylase. Due to the location of this enzyme outside the chloroplast, its function is not known. The enzyme was purified to homogeneity from leaves of both a starchless mutant deficient in plastid phosphoglucomutase and from the wild type using polyethylene glycol fractionation and cyclohexaamylose affinity chromatography. The molecular mass of the {beta}-amylase from both sources was 55,000 daltons as determined by denaturing gel electrophoresis. Gel filtration studies indicated that the enzyme was a monomer. The specific activities of the purified protein from mutant and wild-type sources, their substrate specificities, and K{sub m} for amylopectin were identical. Based on these results it was concluded that the mutant contained an increased level of {beta}-amylase protein. Enzyme neutralization studies using a polyclonal antiserum raised to purified {beta}-amylase showed that in each of two starchless mutants, one starch deficient mutant and one starch overproducing mutant, the elevated amylase activity was due to elevated {beta}-amylase protein.

  12. Homologous Recombination Defective Arabidopsis Mutants Exhibit Enhanced Sensitivity to Abscisic Acid

    PubMed Central

    Roy, Sujit; Das, Kali Pada

    2017-01-01

    Abscisic acid (ABA) acts as an important plant hormone in regulating various aspects of plant growth and developmental processes particularly under abiotic stress conditions. An increased ABA level in plant cells inhibits DNA replication and cell division, causing plant growth retardation. In this study, we have investigated the effects of ABA on the growth responses of some major loss-of-function mutants of DNA double-stand break (DSB) repair genes in Arabidopsis during seed germination and early stages of seedling growth for understanding the role of ABA in the induction of genome instability in plants. A comparative analysis of ABA sensitivity of wild-type Arabidopsis and the knockout mutant lines related to DSB sensors, including atatm, atatr, the non-homologous end joining (NHEJ) pathway genes, and mutants related to homologous recombination (HR) pathway genes showed relatively enhanced sensitivity of atatr and HR-related mutants to ABA treatment. The expression levels of HR-related genes were increased in wild-type Arabidopsis (Col-0) during seed germination and early stages of seedling growth. Immunoblotting experiments detected phosphorylation of histone H2AX in wild-type (Col-0) and DSB repair gene mutants after ABA treatment, indicating the activation of DNA damage response due to ABA treatment. Analyses of DSB repair kinetics using comet assay under neutral condition have revealed comparatively slower DSB repair activity in HR mutants. Overall, our results have provided comprehensive information on the possible effect of ABA on DNA repair machinery in plants and also indicated potential functional involvement of HR pathway in repairing ABA induced DNA damage in Arabidopsis. PMID:28046013

  13. Engineering vitamin E content: from Arabidopsis mutant to soy oil.

    PubMed

    Van Eenennaam, Alison L; Lincoln, Kim; Durrett, Timothy P; Valentin, Henry E; Shewmaker, Christine K; Thorne, Greg M; Jiang, Jian; Baszis, Susan R; Levering, Charlene K; Aasen, Eric D; Hao, Ming; Stein, Joshua C; Norris, Susan R; Last, Robert L

    2003-12-01

    We report the identification and biotechnological utility of a plant gene encoding the tocopherol (vitamin E) biosynthetic enzyme 2-methyl-6-phytylbenzoquinol methyltransferase. This gene was identified by map-based cloning of the Arabidopsis mutation vitamin E pathway gene3-1 (vte3-1), which causes increased accumulation of delta-tocopherol and decreased gamma-tocopherol in the seed. Enzyme assays of recombinant protein supported the hypothesis that At-VTE3 encodes a 2-methyl-6-phytylbenzoquinol methyltransferase. Seed-specific expression of At-VTE3 in transgenic soybean reduced seed delta-tocopherol from 20 to 2%. These results confirm that At-VTE3 protein catalyzes the methylation of 2-methyl-6-phytylbenzoquinol in planta and show the utility of this gene in altering soybean tocopherol composition. When At-VTE3 was coexpressed with At-VTE4 (gamma-tocopherol methyltransferase) in soybean, the seed accumulated to >95% alpha-tocopherol, a dramatic change from the normal 10%, resulting in a greater than eightfold increase of alpha-tocopherol and an up to fivefold increase in seed vitamin E activity. These findings demonstrate the utility of a gene identified in Arabidopsis to alter the tocopherol composition of commercial seed oils, a result with both nutritional and food quality implications.

  14. Arabidopsis thaliana mutant lpsi reveals impairment in the root responses to local phosphate availability.

    PubMed

    Karthikeyan, Athikkattuvalasu S; Jain, Ajay; Nagarajan, Vinay K; Sinilal, Bhaskaran; Sahi, Shivendra V; Raghothama, Kashchandra G

    2014-04-01

    Phosphate (Pi) deficiency triggers local Pi sensing-mediated inhibition of primary root growth and development of root hairs in Arabidopsis (Arabidopsis thaliana). Generation of activation-tagged T-DNA insertion pools of Arabidopsis expressing the luciferase gene (LUC) under high-affinity Pi transporter (Pht1;4) promoter, is an efficient approach for inducing genetic variations that are amenable for visual screening of aberrations in Pi deficiency responses. Putative mutants showing altered LUC expression during Pi deficiency were identified and screened for impairment in local Pi deficiency-mediated inhibition of primary root growth. An isolated mutant was analyzed for growth response, effects of Pi deprivation on Pi content, primary root growth, root hair development, and relative expression levels of Pi starvation-responsive (PSR) genes, and those implicated in starch metabolism and Fe and Zn homeostasis. Pi deprived local phosphate sensing impaired (lpsi) mutant showed impaired primary root growth and attenuated root hair development. Although relative expression levels of PSR genes were comparable, there were significant increases in relative expression levels of IRT1, BAM3 and BAM5 in Pi deprived roots of lpsi compared to those of the wild-type. Better understanding of molecular responses of plants to Pi deficiency or excess will help to develop suitable remediation strategies for soils with excess Pi, which has become an environmental concern. Hence, lpsi mutant will serve as a valuable tool in identifying molecular mechanisms governing adaptation of plants to Pi deficiency.

  15. Plastid sedimentation kinetics in roots of wild-type and starch-deficient mutants of Arabidopsis

    NASA Technical Reports Server (NTRS)

    MacCleery, S. A.; Kiss, J. Z.

    1999-01-01

    Sedimentation and movement of plastids in columella cells of the root cap were measured in seedlings of wild-type, a reduced starch mutant, and a starchless mutant of Arabidopsis. To assay for sedimentation, we used both linear measurements and the change of angle from the cell center as indices in vertical and reoriented plants with the aid of computer-assisted image analysis. Seedlings were fixed at short periods after reorientation, and plastid sedimentation correlated with starch content in the three strains of Arabidopsis. Amyloplasts of wild-type seedlings showed the greatest sedimentation, whereas plastids of the starchless mutant showed no significant sedimentation in the vertically grown and reoriented seedlings. Because previous research has shown that a full complement of starch is needed for full gravitropic sensitivity, this study correlates increased sensitivity with plastid sedimentation. However, although plastid sedimentation contributed to gravisensitivity, it was not required, because the gravitropic starchless mutant had plastids that did not sediment. This is the first study, to our knowledge, to measure plastid sedimentation in Arabidopsis roots after reorientation of seedlings. Taken together, the results of this study are consistent with the classic plastid-based and protoplast-based models of graviperception and suggest that multiple systems of perception exist in plant cells.

  16. Light-Induced Acclimation of the Arabidopsis chlorina1 Mutant to Singlet Oxygen[C][W

    PubMed Central

    Ramel, Fanny; Ksas, Brigitte; Akkari, Elsy; Mialoundama, Alexis S.; Monnet, Fabien; Krieger-Liszkay, Anja; Ravanat, Jean-Luc; Mueller, Martin J.; Bouvier, Florence; Havaux, Michel

    2013-01-01

    Singlet oxygen (1O2) is a reactive oxygen species that can function as a stress signal in plant leaves leading to programmed cell death. In microalgae, 1O2-induced transcriptomic changes result in acclimation to 1O2. Here, using a chlorophyll b–less Arabidopsis thaliana mutant (chlorina1 [ch1]), we show that this phenomenon can also occur in vascular plants. The ch1 mutant is highly photosensitive due to a selective increase in the release of 1O2 by photosystem II. Under photooxidative stress conditions, the gene expression profile of ch1 mutant leaves very much resembled the gene responses to 1O2 reported in the Arabidopsis mutant flu. Preexposure of ch1 plants to moderately elevated light intensities eliminated photooxidative damage without suppressing 1O2 formation, indicating acclimation to 1O2. Substantial differences in gene expression were observed between acclimation and high-light stress: A number of transcription factors were selectively induced by acclimation, and contrasting effects were observed for the jasmonate pathway. Jasmonate biosynthesis was strongly induced in ch1 mutant plants under high-light stress and was noticeably repressed under acclimation conditions, suggesting the involvement of this hormone in 1O2-induced cell death. This was confirmed by the decreased tolerance to photooxidative damage of jasmonate-treated ch1 plants and by the increased tolerance of the jasmonate-deficient mutant delayed-dehiscence2. PMID:23590883

  17. Organ fusion and defective cuticle function in a lacs1 lacs2 double mutant of Arabidopsis.

    PubMed

    Weng, Hua; Molina, Isabel; Shockey, Jay; Browse, John

    2010-04-01

    As the outermost layer on aerial tissues of the primary plant body, the cuticle plays important roles in plant development and physiology. The major components of the cuticle are cutin and cuticular wax, both of which are composed primarily of fatty acid derivatives synthesized in the epidermal cells. Long-chain acyl-CoA synthetases (LACS) catalyze the formation of long-chain acyl-CoAs and the Arabidopsis genome contains a family of nine genes shown to encode LACS enzymes. LACS2 is required for cutin biosynthesis, as revealed by previous investigations on lacs2 mutants. Here, we characterize lacs1 mutants of Arabidopsis that reveals a role for LACS1 in biosynthesis of cuticular wax components. lacs1 lacs2 double-mutant plants displayed pleiotropic phenotypes including organ fusion, abnormal flower development and reduced seed set; phenotypes not found in either of the parental mutants. The leaf cuticular permeability of lacs1 lacs2 was higher than that of either lacs1 or lacs2 single mutants, as determined by measurements of chlorophyll leaching from leaves immersed in 80% ethanol, staining with toluidine blue dye and direct measurements of water loss. Furthermore, lacs1 lacs2 mutant plants are highly susceptible to drought stress. Our results indicate that a deficiency in cuticular wax synthesis and a deficiency in cutin synthesis together have compounding effects on the functional integrity of the cuticular barrier, compromising the ability of the cuticle to restrict water movement, protect against drought stress and prevent organ fusion.

  18. Generation of Arabidopsis mutants by heterologous expression of a full length cDNA library from tomato fruits

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Heterologous expression of cDNA libraries in Arabidopsis and other plants has been used for gene identifications. To identify functions of tomato genes, we expressed a tomato full-length cDNA library in Arabidopsis thaliana and generated over 7,000 mutants. We constructed a tomato cDNA library with ...

  19. A sequence-based map of Arabidopsis genes with mutant phenotypes.

    PubMed

    Meinke, David W; Meinke, Laura K; Showalter, Thomas C; Schissel, Anna M; Mueller, Lukas A; Tzafrir, Iris

    2003-02-01

    The classical genetic map of Arabidopsis contains 462 genes with mutant phenotypes. Chromosomal locations of these genes have been determined over the past 25 years based on recombination frequencies with visible and molecular markers. The most recent update of the classical map was published in a special genome issue of Science that dealt with Arabidopsis (D.W. Meinke, J.M. Cherry, C. Dean, S.D. Rounsley, M. Koornneef [1998] Science 282: 662-682). We present here a comprehensive list and sequence-based map of 620 cloned genes with mutant phenotypes. This map documents for the first time the exact locations of large numbers of Arabidopsis genes that give a phenotype when disrupted by mutation. Such a community-based physical map should have broad applications in Arabidopsis research and should serve as a replacement for the classical genetic map in the future. Assembling a comprehensive list of genes with a loss-of-function phenotype will also focus attention on essential genes that are not functionally redundant and ultimately contribute to the identification of the minimal gene set required to make a flowering plant.

  20. Cellular differentiation regulated by gibberellin in the Arabidopsis thaliana pickle mutant.

    PubMed

    Ogas, J; Cheng, J C; Sung, Z R; Somerville, C

    1997-07-04

    The plant growth regulator gibberellin (GA) has a profound effect on shoot development and promotes developmental transitions such as flowering. Little is known about any analogous effect GA might have on root development. In a screen for mutants, Arabidopsis plants carrying a mutation designated pickle (pkl) were isolated in which the primary root meristem retained characteristics of embryonic tissue. Expression of this aberrant differentiation state was suppressed by GA. Root tissue from plants carrying the pkl mutation spontaneously regenerated new embryos and plants.

  1. A direct screening procedure for gravitropism mutants in Arabidopsis thaliana (L. ) Heynh

    SciTech Connect

    Bullen, B.L.; Best, T.R.; Gregg, M.M.; Barsel, S.E.; Poff, K.L. )

    1990-06-01

    In order to isolate gravitropism mutants of Arabidopsis thaliana (L.) Heynh. var Estland for the genetic dissection of the gravitropism pathway, a direct screening procedure has been developed in which mutants are selected on the basis of their gravitropic response. Variability in hypocotyl curvature was dependent on the germination time of each seed stock, resulting in the incorrect identification of several lines as gravitropism mutants when a standard protocol for the potentiation of germination was used. When the protocol was adjusted to allow for differences in germination time, these lines were eliminated from the collection. Out of the 60,000 M2 seedlings screened, 0.3 to 0.4% exhibited altered gravitropism. In approximately 40% of these mutant lines, only gravitropism by the root or the hypocotyl was altered, while the response of the other organ was unaffected. These data support the hypothesis that root and hypocotyl gravitropism are genetically separable.

  2. A starch-accumulating mutant of Arabidopsis thaliana deficient in a chloroplastic starch-hydrolysing enzyme.

    PubMed

    Zeeman, S C; Northrop, F; Smith, A M; Rees, T

    1998-08-01

    The aim of this work was to identify enzymes that participate in the degradation of transitory starch in Arabidopsis. A mutant line was isolated by screening leaves at the end of the night for the presence of starch. The mutant had a higher starch content than the wild-type throughout the diurnal cycle. This accumulation was due to a reduction in starch breakdown, leading to an imbalance between the rates of synthesis and degradation. No reduction in the activity of endo-amylase (alpha-amylase), beta-amylase, starch phosphorylase, maltase, pullulanase or D-enzyme could be detected in crude extracts of leaves of the mutant. However, native PAGE in gels containing amylopectin revealed that a starch-hydrolysing activity, putatively identified as an endo-amylase and present in wild-type chloroplasts, was absent or appreciably reduced in the mutant. This is the first time that a specific enzyme required for starch degradation has been identified in leaves.

  3. A direct screening procedure for gravitropism mutants in Arabidopsis thaliana (L.) Heynh

    NASA Technical Reports Server (NTRS)

    Bullen, B. L.; Best, T. R.; Gregg, M. M.; Poff, K. L.; Barsel, S-E (Principal Investigator)

    1990-01-01

    In order to isolate gravitropism mutants of Arabidopsis thaliana (L.) Heynh. var Estland for the genetic dissection of the gravitropism pathway, a direct screening procedure has been developed in which mutants are selected on the basis of their gravitropic response. Variability in hypocotyl curvature was dependent on the germination time of each seed stock, resulting in the incorrect identification of several lines as gravitropism mutants when a standard protocol for the potentiation of germination was used. When the protocol was adjusted to allow for differences in germination time, these lines were eliminated from the collection. Out of the 60,000 M2 seedlings screened, 0.3 to 0.4% exhibited altered gravitropism. In approximately 40% of these mutant lines, only gravitropism by the root or the hypocotyl was altered, while the response of the other organ was unaffected. These data support the hypothesis that root and hypocotyl gravitropism are genetically separable.

  4. An Arabidopsis brassinosteroid-dependent mutant is blocked in cell elongation.

    PubMed Central

    Azpiroz, R; Wu, Y; LoCascio, J C; Feldmann, K A

    1998-01-01

    Cell elongation is a developmental process that is regulated by light and phytohormones and is of critical importance for plant growth. Mutants defective in their response to light and various hormones are often dwarfs. The dwarfed phenotype results because of a failure in normal cell elongation. Little is known, however, about the basis of dwarfism as a common element in these diverse signaling pathways and the nature of the cellular functions responsible for cell elongation. Here, we describe an Arabidopsis mutant, dwarf4 (dwf4), whose phenotype can be rescued with exogenously supplied brassinolide. dwf4 mutants display features of light-regulatory mutants, but the dwarfed phenotype is entirely and specifically brassinosteroid dependent; no other hormone can rescue dwf4 to a wild-type phenotype. Therefore, an intact brassinosteroid system is an absolute requirement for cell elongation. PMID:9490745

  5. On the origin of a slowly reversible fluorescence decay component in the Arabidopsis npq4 mutant.

    PubMed

    Dall'Osto, Luca; Cazzaniga, Stefano; Wada, Masamitsu; Bassi, Roberto

    2014-04-19

    Over-excitation of photosynthetic apparatus causing photoinhibition is counteracted by non-photochemical quenching (NPQ) of chlorophyll fluorescence, dissipating excess absorbed energy into heat. The PsbS protein plays a key role in this process, thus making the PsbS-less npq4 mutant unable to carry out qE, the major and most rapid component of NPQ. It was proposed that npq4 does perform qE-type quenching, although at lower rate than WT Arabidopsis. Here, we investigated the kinetics of NPQ in PsbS-depleted mutants of Arabidopsis. We show that red light was less effective than white light in decreasing maximal fluorescence in npq4 mutants. Also, the kinetics of fluorescence dark recovery included a decay component, qM, exhibiting the same amplitude and half-life in both WT and npq4 mutants. This component was uncoupler-sensitive and unaffected by photosystem II repair or mitochondrial ATP synthesis inhibitors. Targeted reverse genetic analysis showed that traits affecting composition of the photosynthetic apparatus, carotenoid biosynthesis and state transitions did not affect qM. This was depleted in the npq4phot2 mutant which is impaired in chloroplast photorelocation, implying that fluorescence decay, previously described as a quenching component in npq4 is, in fact, the result of decreased photon absorption caused by chloroplast relocation rather than a change in the activity of quenching reactions.

  6. A chilling sensitive mutant of Arabidopsis with altered steryl-ester metabolism

    SciTech Connect

    Hugly, S.; McCourt, P.; Somerville, C. ); Browse, J. ); Patterson, G.W. )

    1990-07-01

    A chilling-sensitive mutant of Arabidopsis thaliana was isolated and subjected to genetic, physiological, and biochemical analysis. The chilling-sensitive nature of the mutant line is due to a single recessive nuclear mutation at a locus designated chs1. In contrast to wild-type plants, which are not adversely affected by low temperatures, the chs1 mutant is killed by several days of exposure to temperatures below 18{degree}C. Following exposure to chilling temperatures, the mutant displays two common symptoms of chilling injury - leaf chlorosis and electrolyte leakage. In these respects, the physiological response of the mutant to low temperatures mimics the response observed in some naturally occurring chilling sensitive species. The biochemical basis of chilling sensitivity was explored by examining the pattern of incorporation of {sup 14}CO{sub 2} into soluble metabolites and lipids in wild-type and mutant plants. The only difference observed between the mutant and wild type was that following low temperature treatment, the mutant accumulated 10-fold more radioactivity in a specific class of neutral lipids which were identified by a variety of criteria to be steryl-esters. The accumulation of radioactivity in the steryl-ester fraction occurs 24 hours before there is any visible evidence of chilling injury.

  7. The TOC159 mutant of Arabidopsis thaliana accumulates altered levels of saturated and polyunsaturated fatty acids.

    PubMed

    Afitlhile, Meshack; Fry, Morgan; Workman, Samantha

    2015-02-01

    We evaluated whether the TOC159 mutant of Arabidopsis called plastid protein import 2-2 (ppi2-2) accumulates normal levels of fatty acids, and transcripts of fatty acid desaturases and galactolipid synthesis enzymes. The ppi2-2 mutant accumulates decreased pigments and total fatty acid content. The MGD1 gene was downregulated and the mutant accumulates decreased levels of monogalactosyldiacylglycerol (MGDG) and 16:3, which suggests that the prokaryotic pathway was impaired in the mutant. The HY5 gene, which encodes long hypocotyl5 transcription factor, was upregulated in the mutant. The DGD1 gene, an HY5 target was marginally increased and the mutant accumulates digalactosyldiacylglycerol at the control level. The mutant had increased expression of 3-ketoacyl-ACP synthase II gene, which encodes a plastid enzyme that elongates 16:0 to 18:0. Interestingly, glycerolipids in the mutant accumulate increased levels of 18:0. A gene that encodes stearoyl-ACP desaturase (SAD) was expressed at the control level and 18:1 was increased, which suggest that SAD may be strongly regulated at the posttranscriptional level. The molar ratio of MGDG to bilayer forming plastid lipids was decreased in the cold-acclimated wild type but not in the ppi2-2 mutant. This indicates that the mutant was unresponsive to cold-stress, and is consistent with increased levels of 18:0, and decreased 16:3 and 18:3 in the ppi2-2 mutant. Overall, these data indicate that a defective Toc159 receptor impaired the synthesis of MGDG, and affected desaturation of 16 and 18-carbon fatty acids. We conclude that expression of the MGD1 gene and synthesis of MGDG are tightly linked to plastid biogenesis.

  8. Use of Arabidopsis eceriferum mutants to explore plant cuticle biosynthesis.

    PubMed

    Samuels, Lacey; DeBono, Allan; Lam, Patricia; Wen, Miao; Jetter, Reinhard; Kunst, Ljerka

    2008-05-31

    The plant cuticle is a waxy outer covering on plants that has a primary role in water conservation, but is also an important barrier against the entry of pathogenic microorganisms. The cuticle is made up of a tough crosslinked polymer called "cutin" and a protective wax layer that seals the plant surface. The waxy layer of the cuticle is obvious on many plants, appearing as a shiny film on the ivy leaf or as a dusty outer covering on the surface of a grape or a cabbage leaf thanks to light scattering crystals present in the wax. Because the cuticle is an essential adaptation of plants to a terrestrial environment, understanding the genes involved in plant cuticle formation has applications in both agriculture and forestry. Today, we'll show the analysis of plant cuticle mutants identified by forward and reverse genetics approaches.

  9. Arabidopsis thaliana siRNA biogenesis mutants have the lower frequency of homologous recombination

    PubMed Central

    Yao, Youli; Bilichak, Andriy; Golubov, Andrey; Kovalchuk, Igor

    2016-01-01

    ABSTRACT Small interfering RNAs (siRNAs) are involved in the regulation of plant development and response to stress. We have previously shown that mutants impaired in Dicer-like 2 (DCL2), DCL3 and DCL4, RDR2, RDR6 and NPRD1 are partially impaired in their response to stress and dcl2 and dcl3 plants are also impaired in transgenerational response to stress, including changes in homologous recombination frequency (HRF). Here, we have analyzed genome stability of dcl2, dcl3, dcl4, dcl2 dcl3, dcl2 dcl3 dcl4 and rdr6 mutants by measuring the non-induced and the stress-induced recombination frequency. We found that all mutants had the lower spontaneous HRF. The analysis of strand breaks showed that all tested Arabidopsis mutants had a higher level of spontaneous strand breaks, suggesting that the lower HRF is not due to the unusually low level of breaks. Exposure to methyl methane sulfonate (MMS) resulted in an increase in the level of strand breaks in wild-type plants and a decrease in mutants. All mutants had the higher methylation of cytosines at CpG sites under non-induced conditions. Exposure to MMS resulted in a decrease in methylation level in wild-type plants and an increase in methylation in all dcl mutants. The expression of several DNA repair genes was altered in dcl4 plants under non-induced and induced conditions. Our data suggest that siRNA biogenesis may be essential for the maintenance of the genome stability and stress response in Arabidopsis. PMID:26901311

  10. Arabidopsis thaliana siRNA biogenesis mutants have the lower frequency of homologous recombination.

    PubMed

    Yao, Youli; Bilichak, Andriy; Golubov, Andrey; Kovalchuk, Igor

    2016-07-02

    Small interfering RNAs (siRNAs) are involved in the regulation of plant development and response to stress. We have previously shown that mutants impaired in Dicer-like 2 (DCL2), DCL3 and DCL4, RDR2, RDR6 and NPRD1 are partially impaired in their response to stress and dcl2 and dcl3 plants are also impaired in transgenerational response to stress, including changes in homologous recombination frequency (HRF). Here, we have analyzed genome stability of dcl2, dcl3, dcl4, dcl2 dcl3, dcl2 dcl3 dcl4 and rdr6 mutants by measuring the non-induced and the stress-induced recombination frequency. We found that all mutants had the lower spontaneous HRF. The analysis of strand breaks showed that all tested Arabidopsis mutants had a higher level of spontaneous strand breaks, suggesting that the lower HRF is not due to the unusually low level of breaks. Exposure to methyl methane sulfonate (MMS) resulted in an increase in the level of strand breaks in wild-type plants and a decrease in mutants. All mutants had the higher methylation of cytosines at CpG sites under non-induced conditions. Exposure to MMS resulted in a decrease in methylation level in wild-type plants and an increase in methylation in all dcl mutants. The expression of several DNA repair genes was altered in dcl4 plants under non-induced and induced conditions. Our data suggest that siRNA biogenesis may be essential for the maintenance of the genome stability and stress response in Arabidopsis.

  11. Mutation rate and novel tt mutants of Arabidopsis thaliana induced by carbon ions.

    PubMed Central

    Shikazono, Naoya; Yokota, Yukihiko; Kitamura, Satoshi; Suzuki, Chihiro; Watanabe, Hiroshi; Tano, Shigemitsu; Tanaka, Atsushi

    2003-01-01

    Irradiation of Arabidopsis thaliana by carbon ions was carried out to investigate the mutational effect of ion particles in higher plants. Frequencies of embryonic lethals and chlorophyll-deficient mutants were found to be significantly higher after carbon-ion irradiation than after electron irradiation (11-fold and 7.8-fold per unit dose, respectively). To estimate the mutation rate of carbon ions, mutants with no pigments on leaves and stems (tt) and no trichomes on leaves (gl) were isolated at the M2 generation and subjected to analysis. Averaged segregation rate of the backcrossed mutants was 0.25, which suggested that large deletions reducing the viability of the gametophytes were not transmitted, if generated, in most cases. During the isolation of mutants, two new classes of flavonoid mutants (tt18, tt19) were isolated from carbon-ion-mutagenized M2 plants. From PCR and sequence analysis, two of the three tt18 mutant alleles were found to have a small deletion within the LDOX gene and the other was revealed to contain a rearrangement. Using the segregation rates, the mutation rate of carbon ions was estimated to be 17-fold higher than that of electrons. The isolation of novel mutants and the high mutation rate suggest that ion particles can be used as a valuable mutagen for plant genetics. PMID:12702688

  12. A mutant of Arabidopsis lacking a chloroplastic isoamylase accumulates both starch and phytoglycogen.

    PubMed Central

    Zeeman, S C; Umemoto, T; Lue, W L; Au-Yeung, P; Martin, C; Smith, A M; Chen, J

    1998-01-01

    In this study, our goal was to evaluate the role of starch debranching enzymes in the determination of the structure of amylopectin. We screened mutant populations of Arabidopsis for plants with alterations in the structure of leaf starch by using iodine staining. The leaves of two mutant lines stained reddish brown, whereas wild-type leaves stained brownish black, indicating that a more highly branched polyglucan than amylopectin was present. The mutants were allelic, and the mutation mapped to position 18.8 on chromosome 1. One mutant line lacked the transcript for a gene with sequence similarity to higher plant debranching enzymes, and both mutants lacked a chloroplastic starch-hydrolyzing enzyme. This enzyme was identified as a debranching enzyme of the isoamylase type. The loss of this isoamylase resulted in a 90% reduction in the accumulation of starch in this mutant line when compared with the wild type and in the accumulation of the highly branched water-soluble polysaccharide phytoglycogen. Both normal starch and phytoglycogen accumulated simultaneously in the same chloroplasts in the mutant lines, suggesting that isoamylase has an indirect rather than a direct role in determining amylopectin structure. PMID:9761796

  13. Reduced gravitropism in hypocotyls of starch-deficient mutants of Arabidopsis.

    PubMed

    Kiss, J Z; Guisinger, M M; Miller, A J; Stackhouse, K S

    1997-05-01

    Gravitropism was examined in dark- and light-grown hypocotyls of wild-type (WT), two reduced starch mutants (ACG 20 and ACG 27), and a starchless mutant (ACG 21) of Arabidopsis. In addition, the starch content of these four strains was studied with light and electron microscopy. Based on time course of curvature and orientation studies, the graviresponse in hypocotyls is proportional to the amount of starch in a genotype. Furthermore, starch mutations seem to primarily affect gravitropism rather than differential growth since both phototropic curvature and growth rates among the four genotypes are approximately equal. Our results suggest that gravity perception may require a greater plastid mass in hypocotyls compared to roots. The kinetics of gravitropic curvature also was compared following reorientation at 45 degrees, 90 degrees, and 135 degrees. As has been reported for other plant species, the optimal angle of reorientation is 135 degrees for WT Arabidopsis and the two reduced starch mutants, but the magnitude of curvature of the starchless mutant appears to be independent of the initial angle of displacement. Taken together, the results of the present study and our previous experiments with roots of the same four genotypes [Kiss et al. (1996) Physiol. Plant. 97: 237] support a plastid-based hypothesis for gravity perception in plants.

  14. Spaceflight experiments with Arabidopsis starch-deficient mutants support a statolith-based model for graviperception

    NASA Astrophysics Data System (ADS)

    Kiss, John Z.; Edelmann, Richard E.

    1999-01-01

    In order to help resolve some of the controversy associated with ground-based research that has supported the starch-statolith theory of gravity perception in plants, we performed spaceflight experiments with Arabidopsis in Biorack during the January 1997 and May 1997 missions of the Space Shuttle. Seedlings of wild-type (WT) Arabidopsis, two reduced-starch strains, and a starchless mutant were grown in microgravity and then were given either a 30, 60, or 90 minute gravity stimulus on a centrifuge. By the 90 min 1-g stimulus, the WT exhibited the greatest magnitude of curvature and the starchless mutant exhibited the smallest curvature while the two reduced starch mutants had an intermediate magnitude of curvature. In addition, space-grown plants had two structural features that distinguished them from the controls: a greater number of root hairs and an anomalous hypocotyl hook structure. However, the morphological changes observed in the flight seedlings are likely to be due to the effects of ethylene present in the spacecraft. (Additional ground-based studies demonstrated that this level of ethylene did not significantly affect gravitropism nor did it affect the relative gravitropic sensitivity among the four strains.) Nevertheless, this experiment on gravitropism was performed the “right way” in that brief gravitational stimuli were provided, and the seedlings were allowed to express the response without further gravity stimuli. Our spaceflight results support previous ground-based studies of these and other mutants since increasing amounts of starch correlated positively with increasing sensitivity to gravity.

  15. The aba mutant of Arabidopsis thaliana is impaired in epoxy-carotenoid biosynthesis

    SciTech Connect

    Rock, C.D.; Zeevaart, J.A.D. )

    1991-09-01

    The three mutant alleles of the ABA locus of Arabidopsis thaliana result in plants that are deficient in the plant growth regulator abscisic acid (ABA). The authors have used {sup 18}O{sub 2} to label ABA in water-stressed leaves of mutant and wild-type Arabidopsis. Analysis by selected ion monitoring and tandem mass spectrometry of ({sup 18}O)ABA and its catabolites, phaseic acid and ABA-glucose ester ({beta}-D-glucopyranosyl abscisate), indicates that the aba genotypes are impaired in ABA biosynthesis and have a small ABA precursor pool of compounds that contain oxygens on the rings, presumably oxygenated carotenoids (xanthophylls). Quantitation of the carotenoids form mutant and wild-type leaves establishes that the aba alleles cause a deficiency of the epoxy-carotenoids violaxanthin and neoxanthin and an accumulation of their biosynthetic precursor, zeaxanthin. These results provide evidence that ABA is synthesized by oxidative cleavage of epoxy-carotenoids (the indirect pathway). Furthermore the carotenoid mutant they describe undergoes normal greening. Thus the aba alleles provide an opportunity to study the physiological roles of epoxy-carotenoids in photosynthesis in a higher plants.

  16. Reduced gravitropism in hypocotyls of starch-deficient mutants of Arabidopsis

    NASA Technical Reports Server (NTRS)

    Kiss, J. Z.; Guisinger, M. M.; Miller, A. J.; Stackhouse, K. S.

    1997-01-01

    Gravitropism was examined in dark- and light-grown hypocotyls of wild-type (WT), two reduced starch mutants (ACG 20 and ACG 27), and a starchless mutant (ACG 21) of Arabidopsis. In addition, the starch content of these four strains was studied with light and electron microscopy. Based on time course of curvature and orientation studies, the graviresponse in hypocotyls is proportional to the amount of starch in a genotype. Furthermore, starch mutations seem to primarily affect gravitropism rather than differential growth since both phototropic curvature and growth rates among the four genotypes are approximately equal. Our results suggest that gravity perception may require a greater plastid mass in hypocotyls compared to roots. The kinetics of gravitropic curvature also was compared following reorientation at 45 degrees, 90 degrees, and 135 degrees. As has been reported for other plant species, the optimal angle of reorientation is 135 degrees for WT Arabidopsis and the two reduced starch mutants, but the magnitude of curvature of the starchless mutant appears to be independent of the initial angle of displacement. Taken together, the results of the present study and our previous experiments with roots of the same four genotypes [Kiss et al. (1996) Physiol. Plant. 97: 237] support a plastid-based hypothesis for gravity perception in plants.

  17. Effects of ion beam irradiation on size of mutant sector and genetic damage in Arabidopsis

    NASA Astrophysics Data System (ADS)

    Hase, Yoshihiro; Nozawa, Shigeki; Narumi, Issay; Oono, Yutaka

    2017-01-01

    Size of mutant sector and genetic damage were evaluated in Arabidopsis to further our understanding of effective ion beam use in plant mutation breeding. Arabidopsis seeds, heterozygous for the GLABRA1 (GL1) gene (GL1/gl1-1), were irradiated with 15.8 MeV/u neon ions (mean linear energy transfer (LET): 352 keV/μm), 17.3 MeV/u carbon ions (113 keV/μm), or 60Co gamma rays. The frequency and size of glabrous sectors generated because of inactivation of the GL1 allele were examined. The frequency and overall size of large deletions were evaluated based on the loss of heterozygosity of DNA markers using DNA isolated from glabrous tissue. Irrespective of the radiation properties, plants with mutant sectors were obtained at similar frequencies at the same effective dosage necessary for survival reduction. Ion beams tended to induce larger mutant sectors than gamma rays. The frequency of large deletions (>several kbp) increased as the LET value increased, with chromosome regions larger than 100 kbp lost in most large deletions. The distorted segregation ratio of glabrous plants in the progenies of irradiated GL1/gl1-1 plants suggested frequent occurrence of chromosome rearrangement, especially those subjected to neon ions. Exposure to ion beams with moderate LET values (30-110 keV/μm) is thought effective for inducing mutant sectors without causing extensive genetic damage.

  18. Spaceflight experiments with Arabidopsis starch-deficient mutants support a statolith-based model for graviperception.

    PubMed

    Kiss, J Z; Edelmann, R E

    1999-01-01

    In order to help resolve some of the controversy associated with ground-based research that has supported the starch-statolith theory of gravity perception in plants, we performed spaceflight experiments with Arabidopsis in Biorack during the January 1997 and May 1997 missions of the Space Shuttle. Seedlings of wild-type (WT) Arabidopsis, two reduced-starch strains, and a starchless mutant were grown in microgravity and then were given either a 30, 60, or 90 minute gravity stimulus on a centrifuge. By the 90 min 1-g stimulus, the WT exhibited the greatest magnitude of curvature and the starchless mutant exhibited the smallest curvature while the two reduced starch mutants had an intermediate magnitude of curvature. In addition, space-grown plants had two structural features that distinguished them from the controls: a greater number of root hairs and an anomalous hypocotyl hook structure. However, the morphological changes observed in the flight seedlings are likely to be due to the effects of ethylene present in the spacecraft. (Additional ground-based studies demonstrated that this level of ethylene did not significantly affect gravitropism nor did it affect the relative gravitropic sensitivity among the four strains.) Nevertheless, this experiment on gravitropism was performed the "right way" in that brief gravitational stimuli were provided, and the seedlings were allowed to express the response without further gravity stimuli. Our spaceflight results support previous ground-based studies of these and other mutants since increasing amounts of starch correlated positively with increasing sensitivity to gravity.

  19. Selection of T-DNA-tagged male and female gametophytic mutants by segregation distortion in Arabidopsis.

    PubMed Central

    Howden, R; Park, S K; Moore, J M; Orme, J; Grossniklaus, U; Twell, D

    1998-01-01

    As a strategy for the identification of T-DNA-tagged gametophytic mutants, we have used T-DNA insertional mutagenesis based on screening for distorted segregation ratios by antibiotic selection. Screening of approximately 1000 transgenic Arabidopsis families led to the isolation of eight lines showing reproducible segregation ratios of approximately 1:1, suggesting that these lines are putative gametophytic mutants caused by T-DNA insertion at a single locus. Genetic analysis of T-DNA transmission through reciprocal backcrosses with wild type showed severe reductions in genetic transmission of the T-DNA through the male and/or female gametes. Direct evidence for mutant phenotypes in these lines was investigated by DAPI staining of mature pollen grains and by the analysis of seed set and embryo sac morphology in cleared ovules. One line, termed limpet pollen, showed a novel pollen phenotype in that the generative cell failed to migrate inward after pollen mitosis I, such that the generative or sperm cells remained against the pollen wall. Two other lines, andarta and tistrya, were defective in female transmission and showed an early arrest of embryo sac development with the viable megaspore not initiating the nuclear division cycles. These data demonstrate the efficacy of a segregation ratio distortion strategy for the identification of T-DNA-tagged gametophytic mutants in Arabidopsis. PMID:9611178

  20. Chloroplast dysfunction causes multiple defects in cell cycle progression in the Arabidopsis crumpled leaf mutant.

    PubMed

    Hudik, Elodie; Yoshioka, Yasushi; Domenichini, Séverine; Bourge, Mickaël; Soubigout-Taconnat, Ludivine; Mazubert, Christelle; Yi, Dalong; Bujaldon, Sandrine; Hayashi, Hiroyuki; De Veylder, Lieven; Bergounioux, Catherine; Benhamed, Moussa; Raynaud, Cécile

    2014-09-01

    The majority of research on cell cycle regulation is focused on the nuclear events that govern the replication and segregation of the genome between the two daughter cells. However, eukaryotic cells contain several compartmentalized organelles with specialized functions, and coordination among these organelles is required for proper cell cycle progression, as evidenced by the isolation of several mutants in which both organelle function and overall plant development were affected. To investigate how chloroplast dysfunction affects the cell cycle, we analyzed the crumpled leaf (crl) mutant of Arabidopsis (Arabidopsis thaliana), which is deficient for a chloroplastic protein and displays particularly severe developmental defects. In the crl mutant, we reveal that cell cycle regulation is altered drastically and that meristematic cells prematurely enter differentiation, leading to reduced plant stature and early endoreduplication in the leaves. This response is due to the repression of several key cell cycle regulators as well as constitutive activation of stress-response genes, among them the cell cycle inhibitor SIAMESE-RELATED5. One unique feature of the crl mutant is that it produces aplastidic cells in several organs, including the root tip. By investigating the consequence of the absence of plastids on cell cycle progression, we showed that nuclear DNA replication occurs in aplastidic cells in the root tip, which opens future research prospects regarding the dialogue between plastids and the nucleus during cell cycle regulation in higher plants.

  1. A mutant of the Arabidopsis thaliana Toc159 gene accumulates reduced levels of linolenic acid and monogalactosyldiacylglycerol

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies have shown that a null mutant of Arabidopsis that lacks Toc159 receptor is impaired in chloroplast biogenesis and incapable of importing photosynthetic proteins. The mutant is referred to as plastid protein import 2 or ppi2, and has an albino phenotype. In this study, we measured ...

  2. A mutant of the Arabidopsis thaliana TOC159 gene accumulates reduced levels of linolenic acid and monogalactosyldiacylglycerol

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Previous studies have shown that a mutant of Arabidopsis that lacks the Toc159 receptor is impaired in chloroplast biogenesis. The mutant is referred as plastid protein import 2 or ppi2 and has an albino phenotype due to its inability to import the photosynthetic proteins. In this study, we measured...

  3. Mighty Dwarfs: Arabidopsis Autoimmune Mutants and Their Usages in Genetic Dissection of Plant Immunity

    PubMed Central

    van Wersch, Rowan; Li, Xin; Zhang, Yuelin

    2016-01-01

    Plants lack the adaptive immune system possessed by mammals. Instead they rely on innate immunity to defend against pathogen attacks. Genomes of higher plants encode a large number of plant immune receptors belonging to different protein families, which are involved in the detection of pathogens and activation of downstream defense pathways. Plant immunity is tightly controlled to avoid activation of defense responses in the absence of pathogens, as failure to do so can lead to autoimmunity that compromises plant growth and development. Many autoimmune mutants have been reported, most of which are associated with dwarfism and often spontaneous cell death. In this review, we summarize previously reported Arabidopsis autoimmune mutants, categorizing them based on their functional groups. We also discuss how their obvious morphological phenotypes make them ideal tools for epistatic analysis and suppressor screens, and summarize genetic screens that have been carried out in various autoimmune mutant backgrounds. PMID:27909443

  4. Overexpression of the FAD3 desaturase gene in a mutant of Arabidopsis.

    PubMed Central

    Shah, S; Xin, Z; Browse, J

    1997-01-01

    A mutant of Arabidopsis contained increased levels of 18:3 fatty acids and correspondingly decreased levels of 18:2. The fatty acid phenotype was strongly expressed in root and seed tissues and this observation, together with other data, suggested that the mutation leads to increased activity of the endoplasmic reticulum 18:2 desaturase encoded by the FAD3 gene. Gel-blot analysis of RNA from wild-type and mutant plants established that FAD3 transcript levels were increased 80% in the mutant relative to the wild type. Genetic analysis demonstrated a linkage between the new mutation and the fad3 locus. Linkage of the mutation to fad3 raises the possibility that the lesion is an alteration to the promoter or another regulatory region of the FAD3 gene, which results in increased transcription. PMID:9276960

  5. Gravity-induced modifications to development in hypocotyls of Arabidopsis tubulin mutants.

    PubMed

    Matsumoto, Shouhei; Kumasaki, Saori; Soga, Kouichi; Wakabayashi, Kazuyuki; Hashimoto, Takashi; Hoson, Takayuki

    2010-02-01

    We investigated the roles of cortical microtubules in gravity-induced modifications to the development of stem organs by analyzing morphology and orientation of cortical microtubule arrays in hypocotyls of Arabidopsis (Arabidopsis thaliana) tubulin mutants, tua3(D205N), tua4(S178Delta), and tua6(A281T), cultivated under 1g and hypergravity (300g) conditions. Hypocotyls of tubulin mutants were shorter and thicker than the wild type even at 1g, and hypergravity further suppressed elongation and stimulated expansion. The degree of such changes was clearly smaller in tubulin mutants, in particular in tua6. Hypocotyls of tubulin mutants also showed either left-handed or right-handed helical growth at 1g, and the degree of twisting phenotype was intensified under hypergravity conditions, especially in tua6. Hypergravity induced reorientation of cortical microtubules from transverse to longitudinal directions in epidermal cells of wild-type hypocotyls. In tubulin mutants, especially in tua6, the percentage of cells with longitudinal microtubules was high even at 1g, and it was further increased by hypergravity. The twisting phenotype was most obvious at cells 10 to 12 from the top, where reorientation of cortical microtubules from transverse to longitudinal directions occurred. Moreover, the left-handed helical growth mutants (tua3 and tua4) had right-handed microtubule arrays, whereas the right-handed mutant (tua6) had left-handed arrays. There was a close correlation between the alignment angle of epidermal cell files and the alignment of cortical microtubules. Gadolinium ions, blockers of mechanosensitive ion channels (mechanoreceptors), suppressed the twisting phenotype in tubulin mutants under both 1g and 300 g conditions. Microtubule arrays in tubulin mutants were oriented more transversely by gadolinium treatment, irrespective of gravity conditions. These results support the hypothesis that cortical microtubules play an essential role in maintenance of normal

  6. Ascorbate-Deficient Mutants of Arabidopsis Grow in High Light Despite Chronic Photooxidative Stress1

    PubMed Central

    Müller-Moulé, Patricia; Golan, Talila; Niyogi, Krishna K.

    2004-01-01

    Acclimation to changing environments, such as increases in light intensity, is necessary, especially for the survival of sedentary organisms like plants. To learn more about the importance of ascorbate in the acclimation of plants to high light (HL), vtc2, an ascorbate-deficient mutant of Arabidopsis, and the double mutants vtc2npq4 and vtc2npq1 were tested for growth in low light and HL and compared with the wild type. The vtc2 mutant has only 10% to 30% of wild-type levels of ascorbate, vtc2npq4 has lower ascorbate levels and lacks non-photochemical quenching of chlorophyll fluorescence (NPQ) because of the absence of the photosystem II protein PsbS, and vtc2npq1 is NPQ deficient and also lacks zeaxanthin in HL but has PsbS. All three genotypes were able to grow in HL and had wild-type levels of Lhcb1, cytochrome f, PsaF, and 2-cysteine peroxiredoxin. However, the mutants had lower electron transport and oxygen evolution rates and lower quantum efficiency of PSII compared with the wild type, implying that they experienced chronic photooxidative stress. The mutants lacking NPQ in addition to ascorbate were only slightly more affected than vtc2. All three mutants had higher glutathione levels than the wild type in HL, suggesting a possible compensation for the lower ascorbate content. These results demonstrate the importance of ascorbate for the long-term acclimation of plants to HL. PMID:14963245

  7. Isolation and characterization of Arabidopsis mutants defective in the induction of ethylene biosynthesis by cytokinin

    NASA Technical Reports Server (NTRS)

    Vogel, J. P.; Schuerman, P.; Woeste, K.; Brandstatter, I.; Kieber, J. J.; Evans, M. L. (Principal Investigator)

    1998-01-01

    Cytokinins elevate ethylene biosynthesis in etiolated Arabidopsis seedlings via a post-transcriptional modification of one isoform of the key biosynthetic enzyme ACC synthase. In order to begin to dissect the signaling events leading from cytokinin perception to this modification, we have isolated a series of mutants that lack the ethylene-mediated triple response in the presence of cytokinin due to their failure to increase ethylene biosynthesis. Analysis of genetic complementation and mapping revealed that these Cin mutants (cytokinin-insensitive) represent four distinct complementation groups, one of which, cin4, is allelic to the constitutive photomorphogenic mutant fus9/cop10. The Cin mutants have subtle effects on the morphology of adult plants. We further characterized the Cin mutants by analyzing ethylene biosynthesis in response to various other inducers and in adult tissues, as well as by assaying additional cytokinin responses. The cin3 mutant did not disrupt ethylene biosynthesis under any other conditions, nor did it disrupt any other cytokinin responses. Only cin2 disrupted ethylene biosynthesis in multiple circumstances. cin1 and cin2 made less anthocyanin in response to cytokinin. cin1 also displayed reduced shoot initiation in tissue culture in response to cytokinin, suggesting that it affects a cytokinin signaling element.

  8. New molecular phenotypes in the dst mutants of Arabidopsis revealed by DNA microarray analysis.

    PubMed

    Pérez-Amador, M A; Lidder, P; Johnson, M A; Landgraf, J; Wisman, E; Green, P J

    2001-12-01

    In this study, DNA microarray analysis was used to expand our understanding of the dst1 mutant of Arabidopsis. The dst (downstream) mutants were isolated originally as specifically increasing the steady state level and the half-life of DST-containing transcripts. As such, txhey offer a unique opportunity to study rapid sequence-specific mRNA decay pathways in eukaryotes. These mutants show a threefold to fourfold increase in mRNA abundance for two transgenes and an endogenous gene, all containing DST elements, when examined by RNA gel blot analysis; however, they show no visible aberrant phenotype. Here, we use DNA microarrays to identify genes with altered expression levels in dst1 compared with the parental plants. In addition to verifying the increase in the transgene mRNA levels, which were used to isolate these mutants, we were able to identify new genes with altered mRNA abundance in dst1. RNA gel blot analysis confirmed the microarray data for all genes tested and also was used to catalog the first molecular differences in gene expression between the dst1 and dst2 mutants. These differences revealed previously unknown molecular phenotypes for the dst mutants that will be helpful in future analyses. Cluster analysis of genes altered in dst1 revealed new coexpression patterns that prompt new hypotheses regarding the nature of the dst1 mutation and a possible role of the DST-mediated mRNA decay pathway in plants.

  9. Analysis of crystal structure of Arabidopsis MPK6 and generation of its mutants with higher activity.

    PubMed

    Wang, Bo; Qin, Xinghua; Wu, Juan; Deng, Hongying; Li, Yuan; Yang, Hailian; Chen, Zhongzhou; Liu, Guoqin; Ren, Dongtao

    2016-05-10

    Mitogen-activated protein kinase (MAPK) cascades, which are the highly conserved signalling modules in eukaryotic organisms, have been shown to play important roles in regulating growth, development, and stress responses. The structures of various MAPKs from yeast and animal have been solved, and structure-based mutants were generated for their function analyses, however, the structures of plant MAPKs remain unsolved. Here, we report the crystal structure of Arabidopsis MPK6 at a 3.0 Å resolution. Although MPK6 is topologically similar to ERK2 and p38, the structures of the glycine-rich loop, MAPK insert, substrate binding sites, and L16 loop in MPK6 show notable differences from those of ERK2 and p38. Based on the structural comparison, we constructed MPK6 mutants and analyzed their kinase activity both in vitro and in planta. MPK6(F364L) and MPK6(F368L) mutants, in which Phe364 and Phe368 in the L16 loop were changed to Leu, respectively, acquired higher intrinsic kinase activity and retained the normal MAPKK activation property. The expression of MPK6 mutants with basal activity is sufficient to induce camalexin biosynthesis; however, to induce ethylene and leaf senescence, the expression of MPK6 mutants with higher activity is required. The results suggest that these mutants can be used to analyze the specific biological functions of MPK6.

  10. Increased sensitivity to salt stress in tocopherol-deficient Arabidopsis mutants growing in a hydroponic system.

    PubMed

    Ellouzi, Hasna; Hamed, Karim Ben; Cela, Jana; Müller, Maren; Abdelly, Chedly; Munné-Bosch, Sergi

    2013-02-01

    Recent studies suggest that tocopherols could play physiological roles in salt tolerance but the mechanisms are still unknown. In this study, we analyzed changes in growth, mineral and oxidative status in vte1 and vte4 Arabidopsis thaliana mutants exposed to salt stress. vte1 and vte4 mutants lack α-tocopherol, but only the vte1 mutant is additionally deficient in γ-tocopherol. Results showed that a deficiency in vitamin E leads to reduced growth and increased oxidative stress in hydroponically-grown plants. This effect was observed at early stages, not only in rosettes but also in roots. The vte1 mutant was more sensitive to salt-induced oxidative stress than the wild type and the vte4 mutant. Salt sensitivity was associated with (i) high contents of Na(+), (ii) reduced efficiency of PSII photochemistry (Fv/Fm ratio) and (iii) more pronounced oxidative stress as indicated by increased hydrogen peroxide and malondialdeyde levels. The vte 4 mutant, which accumulates γ- instead of α-tocopherol showed an intermediate sensitivity to salt stress between the wild type and the vte1 mutant. Contents of abscisic acid, jasmonic acid and the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid were higher in the vte1 mutant than the vte4 mutant and wild type. It is concluded that vitamin E-deficient plants show an increased sensitivity to salt stress both in rosettes and roots, therefore indicating the positive role of tocopherols in stress tolerance, not only by minimizing oxidative stress, but also controlling Na(+)/K(+) homeostasis and hormonal balance.

  11. Arabidopsis mutants lacking phenolic sunscreens exhibit enhanced ultraviolet-B injury and oxidative damage

    SciTech Connect

    Landry, L.G.; Last, R.L.; Chapple, C.C.S.

    1995-12-01

    We have assessed ultraviolet-B (UV-B)-induced injury in wild-type Arabidopsis thaliana and two mutants with altered aromatic secondary product biosynthesis. Arabidopsis mutants defective in the ability to synthesize UV-B-absorbing compounds (flavonoids in transparent testa 5 [tt5] and sinapate esters in ferulic acid hydroxylase 1 [fah 1]) are more sensitive to UV-B than is the wild-type Landsberg erecta. Despite its ability to accumulate UV-absorptive flavonoid compounds, the ferulic acid hydroxylase mutant fah1 exhibits more physiological injury (growth inhibition and foliar lesions) than either wild type or tt5. The extreme UV-B sensitivity of fah1 demonstrates the importance of hydroxycinnamate esters as UV-B protectants. Consistent with the whole-plant response, the highest levels of lipid and protein oxidation products were seen in fah1. Ascorbate peroxidase enzyme activity was also increased in the leaves of UV-B-treated plants in a dose- and genotype-dependent manner. These results demonstrate that, in A. thaliana, hydryoxycinnamates are more effective UV-B protectants than flavonoids. The data also indicate that A. thaliana responds to UV-B as an oxidative stress, and sunscreen compounds reduce the oxidative damage caused by UV-B. 36 refs., 6 figs.

  12. Arabidopsis mutants lacking phenolic sunscreens exhibit enhanced ultraviolet-B injury and oxidative damage.

    PubMed Central

    Landry, L G; Chapple, C C; Last, R L

    1995-01-01

    We have assessed ultraviolet-B (UV-B)-induced injury in wild-type Arabidopsis thaliana and two mutants with altered aromatic secondary product biosynthesis. Arabidopsis mutants defective in the ability to synthesize UV-B-absorbing compounds (flavonoids in transparent testa 5 [tt5] and sinapate esters in ferulic acid hydroxylase 1 [fah1]) are more sensitive to UV-B than is the wild-type Landsberg erecta. Despite its ability to accumulate UV-absorptive flavonoid compounds, the ferulic acid hydroxylase mutant fah1 exhibits more physiological injury (growth inhibition and foliar lesions) than either wild type or tt5. The extreme UV-B sensitivity of fah1 demonstrates the importance of hydroxycinnamate esters as UV-B protectants. Consistent with the whole-plant response, the highest levels of lipid and protein oxidation products were seen in fah1. Ascorbate peroxidase enzyme activity was also increased in the leaves of UV-B-treated plants in a dose- and genotype-dependent manner. These results demonstrate that, in A. thaliana, hydroxycinnamates are more effective UV-B protectants than flavonoids. The data also indicate that A. thaliana responds to UV-B as an oxidative stress, and sunscreen compounds reduce the oxidative damage caused by UV-B. PMID:8539286

  13. Arabidopsis mutant analysis and gene regulation define a nonredundant role for glutamate dehydrogenase in nitrogen assimilation.

    PubMed

    Melo-Oliveira, R; Oliveira, I C; Coruzzi, G M

    1996-05-14

    Glutamate dehydrogenase (GDH) is ubiquitous to all organisms, yet its role in higher plants remains enigmatic. To better understand the role of GDH in plant nitrogen metabolism, we have characterized an Arabidopsis mutant (gdh1-1) defective in one of two GDH gene products and have studied GDH1 gene expression. GDH1 mRNA accumulates to highest levels in dark-adapted or sucrose-starved plants, and light or sucrose treatment each repress GDH1 mRNA accumulation. These results suggest that the GDH1 gene product functions in the direction of glutamate catabolism under carbon-limiting conditions. Low levels of GDH1 mRNA present in leaves of light-grown plants can be induced by exogenously supplied ammonia. Under such conditions of carbon and ammonia excess, GDH1 may function in the direction of glutamate biosynthesis. The Arabidopsis gdh-deficient mutant allele gdh1-1 cosegregates with the GDH1 gene and behaves as a recessive mutation. The gdh1-1 mutant displays a conditional phenotype in that seedling growth is specifically retarded on media containing exogenously supplied inorganic nitrogen. These results suggest that GDH1 plays a nonredundant role in ammonia assimilation under conditions of inorganic nitrogen excess. This notion is further supported by the fact that the levels of mRNA for GDH1 and chloroplastic glutamine synthetase (GS2) are reciprocally regulated by light.

  14. Expression of OsCAS (Calcium-Sensing Receptor) in an Arabidopsis Mutant Increases Drought Tolerance.

    PubMed

    Zhao, Xin; Xu, Mengmeng; Wei, Rongrong; Liu, Yang

    2015-01-01

    The calcium-sensing receptor (CaS), which is localized in the chloroplasts, is a crucial regulator of extracellular calcium-induced stomatal closure in Arabidopsis. It has homologs in Oryza sativa and other plants. These sequences all have a rhodanese-like protein domain, which has been demonstrated to be associated with specific stress conditions. In this study, we cloned the Oryza sativa calcium-sensing receptor gene (OsCAS) and demonstrated that OsCAS could sense an increase of extracellular Ca2+ concentration and mediate an increase in cytosolic Ca2+ concentration. The OsCAS gene was transformed into an Arabidopsis CaS knockout mutant (Salk) and overexpressed in the transgenic plants. OsCAS promoted stomatal closure. We screened homozygous transgenic Arabidopsis plants and determined physiological indices such as the oxidative damage biomarker malondialdehyde (MDA), relative membrane permeability (RMP), proline content, and chlorophyll fluorescence parameters, after 21 days of drought treatment. Our results revealed lower RMP and MDA contents and a higher Proline content in transgenic Arabidopsis plants after drought stress, whereas the opposite was observed in Salk plants. With respect to chlorophyll fluorescence, the electron transport rate and effective PSII quantum yield decreased in all lines under drought stress; however, in the transgenic plants these two parameters changed fewer and were higher than those in wild-type and Salk plants. The quantum yield of regulated energy dissipation and nonregulated energy dissipation in PSII were higher in Salk plants, whereas these values were lower in the transgenic plants than in the wild type under drought stress. The above results suggest that the transgenic plants showed better resistance to drought stress by decreasing damage to the cell membrane, increasing the amount of osmoprotectants, and maintaining a relatively high photosynthetic capacity. In conclusion, OsCAS is an extracellular calcium-sensing receptor

  15. Characterization of N-Glycans from Arabidopsis. Application to a Fucose-Deficient Mutant1

    PubMed Central

    Rayon, Catherine; Cabanes-Macheteau, Marion; Loutelier-Bourhis, Corinne; Salliot-Maire, Isabelle; Lemoine, Jérome; Reiter, Wolf-Dieter; Lerouge, Patrice; Faye, Loïc

    1999-01-01

    The structures of glycans N-linked to Arabidopsis proteins have been fully identified. From immuno- and affinodetections on blots, chromatography, nuclear magnetic resonance, and glycosidase sequencing data, we show that Arabidopsis proteins are N-glycosylated by high-mannose-type N-glycans from Man5GlcNAc2 to Man9GlcNAc2, and by xylose- and fucose (Fuc)-containing oligosaccharides. However, complex biantenary structures containing the terminal Lewis a epitope recently reported in the literature (A.-C. Fitchette-Lainé, V. Gomord, M. Cabanes, J.-C. Michalski, M. Saint Macary, B. Foucher, B. Cavalier, C. Hawes, P. Lerouge, and L. Faye [1997] Plant J 12: 1411–1417) were not detected. A similar study was done on the Arabidopsis mur1 mutant, which is affected in the biosynthesis of l-Fuc. In this mutant, one-third of the Fuc residues of the xyloglucan has been reported to be replaced by l-galactose (Gal) (E. Zablackis, W.S. York, M. Pauly, S. Hantus, W.D. Reiter, C.C.S. Chapple, P. Albersheim, and A. Darvill [1996] Science 272: 1808–1810). N-linked glycans from the mutant were identified and their structures were compared with those isolated from the wild-type plants. In about 95% of all N-linked glycans from the mur1 plant, l-Fuc residues were absent and were not replaced by another monosaccharide. However, in the remaining 5%, l-Fuc was found to be replaced by a hexose residue. From nuclear magnetic resonance and mass spectrometry data of the mur1 N-glycans, and by analogy with data reported on mur1 xyloglucan, this subpopulation of N-linked glycans was proposed to be l-Gal-containing N-glycans resulting from the replacement of l-Fuc by l-Gal. PMID:9952469

  16. The toc132toc120 heterozygote mutant of Arabidopsis thaliana accumulates reduced levels of hexadecatrienoic acid.

    PubMed

    Afitlhile, Meshack; Duffield-Duncan, Kayla; Fry, Morgan; Workman, Samantha; Hum-Musser, Sue; Hildebrand, David

    2015-11-01

    A null and heterozygous mutant for the Arabidopsis thaliana TOC132 and TOC120 genes accumulates increased levels of 16:0 and decreased 16:3, suggesting altered homeostasis in fatty acid synthesis. The FAD5 gene encodes a plastid desaturase that catalyzes the first step in the synthesis of 16:3 in monogalactosyldiacylglycerol (MGDG). In non-acclimated toc132toc120+/- mutant plants, the FAD5 gene was repressed and this correlated with decreased levels of 16:3. In cold-acclimated mutant however, the FAD5 gene was upregulated and there was a small increase in 16:3 levels relative to the non-acclimated mutant plants. The MGD1 gene was expressed at control levels and the mutant accumulated levels of MGDG that were similar to the wild type. In the mutant however, MGDG had decreased 16:3 levels, suggesting that the activity of FAD5 desaturase was compromised. In the mutant, the FAD2 and FAD3 genes were downregulated but levels of 18:3-PC were increased, suggesting posttranscriptional regulation for the ER-localized fatty acid desaturases. The Toc120 or Toc159 receptor is likely to compensate for a defective Toc132 receptor. In the cold-acclimated mutant, the TOC159 gene was repressed ca. 300-fold, whereas the TOC120 gene was repressed 7-fold relative to the non-acclimated wild type. Thus, the TOC159 gene is more sensitive to cold-stress and might not compensate for defect in the TOC132 gene under these conditions. Overall, these data show that a mutation in the TOC132 gene results in decreased 16:3 levels, indicating the need for an intact Toc132/Toc120 receptor, presumably to facilitate the import of the FAD5 preprotein into chloroplasts.

  17. Cellular differentiation regulated by gibberellin in the Arabidopsis thaliana pickle mutant

    SciTech Connect

    Ogas, J.; Somerville, C.; Cheng, Jin-Chen; Sung, R.

    1997-07-04

    The plant growth regulator gibberellin (GA) has a profound effect on shoot development and promotes developmental transitions such as flowering. Little is known about any analogous effect GA might have on root development. In a screen for mutants, Arabi-dopsis plants carrying a mutation designated pickle (pkl) were isolated in which the primary root meristem retained characteristics of embryonic tissue. Expression of this aberrant differentiation state was suppressed by GA. Root tissue from plants carrying the pkl mutation spontaneously regenerated new embryos and plants. 19 refs., 3 figs., 1 tab.

  18. Effects of gibberellins on seed germination of phytochrome-deficient mutants of Arabidopsis thaliana.

    PubMed

    Yang, Y Y; Nagatani, A; Zhao, Y J; Kang, B J; Kendrick, R E; Kamiya, Y

    1995-10-01

    Experiments were carried out to explore the involvement of gibberellins (GAs) in the light-induced germination of Arabidopsis thaliana (L.) Heynh, using wild type (WT) and phytochrome-deficient mutants (phyA, phyB and phyAphyB deficient in phytochrome A, B and A plus B, respectively). Seed germination of WT and phytochrome-deficient mutants was inhibited by uniconazole (an inhibitor of an early step in biosynthesis of GA, the oxidation of ent-kaurene) and prohexadione (an inhibitor of late steps, namely, 2 beta- and 3 beta-hydroxylation). This inhibition was overcome by simultaneous application of 10(-5) M GA4. The relative activity of GAs for promoting germination of uniconazole-treated seeds was GA4 > GA1 = GA9 > GA20. The wild type and the phyA and phyB mutants had an increased response to a red light pulse in the presence of GA1, GA4, GA9, GA20 and GA24 but there were no significant differences in activity of each GA between the mutants. Therefore, neither phytochrome A nor hytochrome B appears to regulate GA biosynthesis from GA12 to GA4 during seed germination, since the conversion of GA12 to GA9 is regulated by one enzyme (GA 20-oxidase). However, GA responsiveness appears to be regulated by phytochromes other than phytochromes A and B, since the phyAphyB double mutant retains the photoreversible increased response to GAs after a red light pulse.

  19. Characterization of an Arabidopsis thaliana mutant lacking a cytosolic non-phosphorylating glyceraldehyde-3-phosphate dehydrogenase.

    PubMed

    Rius, Sebastián P; Casati, Paula; Iglesias, Alberto A; Gomez-Casati, Diego F

    2006-08-01

    Non-phosphorylating glyceraldehyde- 3-phosphate dehydrogenase (NP-GAPDH) is a conserved cytosolic protein found in higher plants. In photosynthetic cells, the enzyme is involved in a shuttle transfer mechanism to export NADPH from the chloroplast to the cytosol. To investigate the role of this enzyme in plant tissues, we characterized a mutant from Arabidopsis thaliana having an insertion at the NP-GAPDH gene locus. The homozygous mutant was determined to be null respect to NP-GAPDH, as it exhibited undetectable levels of both transcription of NP-GAPDH mRNA, protein expression and enzyme activity. Transcriptome analysis demonstrated that the insertion mutant plant shows altered expression of several enzymes involved in carbohydrate metabolism. Significantly, cytosolic phosphorylating (NAD-dependent) glyceraldehyde-3-phosphate dehydrogenase mRNA levels are induced in the mutant, which correlates with an increase in enzyme activity. mRNA levels and enzymatic activity of glucose-6-phosphate dehydrogenase were also elevated, correlating with an increase in NADPH concentration. Moreover, increased ROS levels were measured in the mutant plants. Down-regulation of several glycolytic and photosynthetic genes suggests that NP-GAPDH is important for the efficiency of both metabolic processes. The results presented demonstrate that NP-GAPDH has a relevant role in plant growth and development.

  20. A novel root gravitropism mutant of Arabidopsis thaliana exhibiting altered auxin physiology

    NASA Technical Reports Server (NTRS)

    Simmons, C.; Migliaccio, F.; Masson, P.; Caspar, T.; Soll, D.

    1995-01-01

    A root gravitropism mutant was isolated from the DuPont Arabidopsis thaliana T-DNA insertional mutagenesis collection. This mutant has reduced root gravitropism, hence the name rgr1. Roots of rgr1 are shorter than those of wild-type, and they have reduced lateral root formation. In addition, roots of rgr1 coil clockwise on inclined agar plates, unlike wild-type roots which grow in a wavy pattern. The rgr1 mutant has increased resistance, as measured by root elongation, to exogenously applied auxins (6-fold to indole-3-acetic acid, 3-fold to 2,4-dichlorophenoxyacetic acid, and 2-fold to napthyleneacetic acid). It is also resistant to polar auxin transport inhibitors (2-fold to triiodobenzoic acid and 3- to 5-fold to napthylphthalamic acid). The rgr1 mutant does not appear to be resistant to other plant hormone classes. When grown in the presence of 10(-7) M 2,4-dichlorophenoxyacetic acid, rgr1 roots have fewer root hairs than wild type. All these rgr1 phenotypes are Mendelian recessives. Complementation tests indicate that rgr1 is not allelic to previously characterized agravitropic or auxin-resistant mutants. The rgr1 locus was mapped using visible markers to 1.4 +/- 0.6 map units from the CH1 locus at 1-65.4. The rgr1 mutation and the T-DNA cosegregate, suggesting that rgr1 was caused by insertional gene inactivation.

  1. Developmental Defects in Mutants of the PsbP Domain Protein 5 in Arabidopsis thaliana

    PubMed Central

    Roose, Johnna L.; Frankel, Laurie K.; Bricker, Terry M.

    2011-01-01

    Plants contain an extensive family of PsbP-related proteins termed PsbP-like (PPL) and PsbP domain (PPD) proteins, which are localized to the thylakoid lumen. The founding member of this family, PsbP, is an established component of the Photosystem II (PS II) enzyme, and the PPL proteins have also been functionally linked to other photosynthetic processes. However, the functions of the remaining seven PPD proteins are unknown. To elucidate the function of the PPD5 protein (At5g11450) in Arabidopsis, we have characterized a mutant T-DNA insertion line (SALK_061118) as well as several RNAi lines designed to suppress the expression of this gene. The functions of the photosynthetic electron transfer reactions are largely unaltered in the ppd5 mutants, except for a modest though significant decrease in NADPH dehydrogenase (NDH) activity. Interestingly, these mutants show striking plant developmental and morphological defects. Relative to the wild-type Col-0 plants, the ppd5 mutants exhibit both increased lateral root branching and defects associated with axillary bud formation. These defects include the formation of additional rosettes originating from axils at the base of the plant as well as aerial rosettes formed at the axils of the first few nodes of the shoot. The root-branching phenotype is chemically complemented by treatment with the synthetic strigolactone, GR24. We propose that the developmental defects observed in the ppd5 mutants are related to a deficiency in strigolactone biosynthesis. PMID:22174848

  2. Isolation and characterization of a starchless mutant of Arabidopsis thaliana (L. ) Heynh lacking ADPglucose pyrophosphorylase activity

    SciTech Connect

    Lin, Tsanpiao; Caspar, T.; Somerville, C.; Preiss, J. )

    1988-04-01

    A mutant of Arabidopsis thaliana lacking ADPglucose pyrophosphorylase activity (EC 2.7.7.27) was isolated (from a mutagenized population of plants) by screening for the absence of leaf starch. The mutant grows as vigorously as the wild type in continuous light but more slowly than the wild type in a 12 hours light/12 hours dark photoperiod. Genetic analysis showed that the deficiency of both starch and ADPglucose pyrophosphorylase activity were attributable to a single, nuclear, recessive mutation at a locus designated adg1. The absence of starch in the mutant demonstrates that starch synthesis in the chloroplast is entirely dependent on a pathway involving ADPglucose pyrophosphorylase. Analysis of leaf extracts by two-dimensional polyacrylamide gel electrophoresis followed by Western blotting experiments using antibodies specific for spinach ADPglucose pyrophosphorylase showed that two proteins, present in the wild type, were absent from the mutant. The heterozygous F{sub 1} progeny of a cross between the mutant and wild type had a specific activity of ADP-glucose pyrophosphorylase indistinguishable from the wild type. These observations suggest that the mutation in the adg1 gene in TL25 might affect a regulatory locus.

  3. Pollen tube and root-hair tip growth is disrupted in a mutant of Arabidopsis thaliana.

    PubMed Central

    Schiefelbein, J; Galway, M; Masucci, J; Ford, S

    1993-01-01

    The expansion of both root hairs and pollen tubes occurs by a process known as tip growth. In this report, an Arabidopsis thaliana mutant (tip1) is described that displays defects in both root-hair and pollen-tube growth. The root hairs of the tip1 mutant plants are shorter than those of the wild-type plants and branched at their base. The tip1 pollen-tube growth defect was identified by the aberrant segregation ratio of phenotypically normal to mutant seeds in siliques from self-pollinated, heterozygous plants. Homozygous mutant seeds are not randomly distributed in the siliques, comprising only 14.4% of the total seeds, 5.3% of the seeds from the bottom half, and 2.2% of the seeds from the bottom quarter of the heterozygous siliques. Studies of pollen-tube growth in vivo showed that mutant pollen tubes grow more slowly than wild-type pollen through the transmitting tissue of wild-type flowers. Cosegregation studies indicate that the root-hair and pollen-tube defects are caused by the same genetic lesion. Based on these findings, the TIP1 gene is likely to encode a product involved in a fundamental aspect of tip growth in plant cells. PMID:8022944

  4. An Arabidopsis mutant hypersensitive to red and far-red light signals.

    PubMed Central

    Genoud, T; Millar, A J; Nishizawa, N; Kay, S A; Schäfer, E; Nagatani, A; Chua, N H

    1998-01-01

    A new mutant called psi2 (for phytochrome signaling) was isolated by screening for elevated activity of a chlorophyll a/b binding protein-luciferase (CAB2-LUC) transgene in Arabidopsis. This mutant exhibited hypersensitive induction of CAB1, CAB2, and the small subunit of ribulose-1,5-bisphosphate carboxylase (RBCS) promoters in the very low fluence range of red light and a hypersensitive response in hypocotyl growth in continuous red light of higher fluences. In addition, at high- but not low-light fluence rates, the mutant showed light-dependent superinduction of the pathogen-related protein gene PR-1a and developed spontaneous necrotic lesions in the absence of any pathogen. Expression of genes responding to various hormone and environmental stress pathways in the mutant was not significantly different from that of the wild type. Analysis of double mutants demonstrated that the effects of the psi2 mutation are dependent on both phytochromes phyA and phyB. The mutation is recessive and maps to the bottom of chromosome 5. Together, our results suggest that PSI2 specifically and negatively regulates both phyA and phyB phototransduction pathways. The induction of cell death by deregulated signaling pathways observed in psi2 is reminiscent of retinal degenerative diseases in animals and humans. PMID:9634578

  5. Gravitropism of inflorescence stems in starch-deficient mutants of Arabidopsis

    NASA Technical Reports Server (NTRS)

    Weise, S. E.; Kiss, J. Z.

    1999-01-01

    Previous studies have assayed the gravitropic response of roots and hypocotyls of wild type Arabidopsis thaliana, two reduced-starch strains, and a starchless strain. Because there have been few reports on inflorescence gravitropism, in this article, we use microscopic analyses and time-course studies of these mutants and their wild type to study gravitropism in these stems. Sedimentation of plastids was observed in endodermal cells of the wild type and reduced-starch mutants but not in the starchless mutant. In all of these strains, the short inflorescence stems (1.0-2.9 cm) were less responsive to the gravistimulus compared with the long stems (3.0-6.0 cm). In both long and short inflorescence stems, the wild type initially had the greatest response; the starchless mutant had the least response; and the reduced starch mutants exhibited an intermediate response. Furthermore, growth rates among all four strains were approximately equal. At about 6 h after reorientation, inflorescences of all strains returned to a position parallel to the gravity vector. Thus, in inflorescence stems, sedimentation of plastids may act as an accelerator but is not required to elicit a gravitropic response. Furthermore, the site of perception appears to be diffuse throughout the inflorescence stem. These results are consistent with both a plastid-based statolith model and the protoplast pressure hypothesis, and it is possible that multiple systems for gravity perception occur in plant cells.

  6. Isolation and characterization of low-sulphur-tolerant mutants of Arabidopsis.

    PubMed

    Wu, Yu; Zhao, Qing; Gao, Lei; Yu, Xiao-Min; Fang, Ping; Oliver, David J; Xiang, Cheng-Bin

    2010-07-01

    Sulphur is an essential element for plant growth and development as well as for defence against biotic and abiotic stresses. Increasing sulphate utilization efficiency (SUE) is an important issue for crop improvement. Little is known about the genetic determinants of sulphate utilization efficiency. No gain-of-function mutants with improved SUE have been reported to date. Here the isolation and characterization of two low-sulphur-tolerant mutants, sue3 and sue4 are reported using a high-throughput genetic screen where a 'sulphur-free' solid medium was devised to give the selection pressure necessary to suppress the growth of the wild-type seedlings. Both mutants showed improved tolerance to low sulphur conditions and well-developed root systems. The mutant phenotype of both sue3 and sue4 was specific to sulphate deficiency and the mutants displayed enhanced tolerance to heavy metal and oxidative stress. Genetic analysis revealed that sue3 was caused by a single recessive nuclear mutation while sue4 was caused by a single dominant nuclear mutation. The recessive locus in sue3 is the previously identified VirE2-interacting Protein 1. The dominant locus in sue4 is a function-unknown locus activated by the four enhancers on the T-DNA. The function of SUE3 and SUE4 in low sulphur tolerance was confirmed either by multiple mutant alleles or by recapitulation analysis. Taken together, our results demonstrate that this genetic screen is a reasonable approach to isolate Arabidopsis mutants with improved low sulphur tolerance and potentially with enhanced sulphate utilization efficiency. The two loci identified in sue3 and sue4 should assist in understanding the molecular mechanisms of low sulphur tolerance.

  7. Microarray analysis of Arabidopsis WRKY33 mutants in response to the necrotrophic fungus Botrytis cinerea

    PubMed Central

    Sham, Arjun; Moustafa, Khaled; Al-Shamisi, Shamma; Alyan, Sofyan; Iratni, Rabah

    2017-01-01

    The WRKY33 transcription factor was reported for resistance to the necrotrophic fungus Botrytis cinerea. Using microarray-based analysis, we compared Arabidopsis WRKY33 overexpressing lines and wrky33 mutant that showed altered susceptibility to B. cinerea with their corresponding wild-type plants. In the wild-type, about 1660 genes (7% of the transcriptome) were induced and 1054 genes (5% of the transcriptome) were repressed at least twofold at early stages of inoculation with B. cinerea, confirming previous data of the contribution of these genes in B. cinerea resistance. In Arabidopsis wild-type plant infected with B. cinerea, the expressions of the differentially expressed genes encoding for proteins and metabolites involved in pathogen defense and non-defense responses, seem to be dependent on a functional WRKY33 gene. The expression profile of 12-oxo-phytodienoic acid- and phytoprostane A1-treated Arabidopsis plants in response to B. cinerea revealed that cyclopentenones can also modulate WRKY33 regulation upon inoculation with B. cinerea. These results support the role of electrophilic oxylipins in mediating plant responses to B. cinerea infection through the TGA transcription factor. Future directions toward the identification of the molecular components in cyclopentenone signaling will elucidate the novel oxylipin signal transduction pathways in plant defense. PMID:28207847

  8. Characterization of a Selenate-Resistant Arabidopsis Mutant. Root Growth as a Potential Target for Selenate Toxicity1[OA

    PubMed Central

    El Kassis, Elie; Cathala, Nicole; Rouached, Hatem; Fourcroy, Pierre; Berthomieu, Pierre; Terry, Norman; Davidian, Jean-Claude

    2007-01-01

    Screening an Arabidopsis (Arabidopsis thaliana) T-DNA mutant library for selenate resistance enabled us to isolate a selenate-resistant mutant line (sel1-11). Molecular and genetic characterization showed that the mutant contained a lesion in the SULTR1;2 gene that encodes a high affinity root sulfate transporter. We showed that SULTR1;2 is the only gene among 13 mutated genes of the Arabidopsis sulfate transporter family whose mutation conferred selenate resistance to Arabidopsis. The selenate resistance phenotype of the sel1-11 mutant was mirrored by an 8-fold increase of root growth in the presence of selenate as shown by the calculated lethal concentration values. The impairment of SULTR1;2 activity in sel1-11 resulted in a reduced 35S-sulfate uptake capacity by both roots and calli and a reduced sulfate and selenate content in root, shoot, and calli. Comparing sulfate-to-selenate ratios instead of absolute sulfate and selenate contents in roots and shoots enabled us to gain better insight into the mechanism of selenate toxicity in Arabidopsis. Roots of the sel1-11 mutant line showed a higher sulfate to selenate ratio than that of wild-type roots, while there were no significant differences in sulfate to selenate ratios in shoots of wild-type and mutant lines. These results indicated that the mechanism that confers the selenate resistance phenotype to the sel1-11 line takes place rather in the roots. It might be in part the result of a lower selenate uptake and of a protective effect of sulfate against the toxic effects of selenate on root growth. These results revealed in plants a central and specific role of the transporter SULTR1;2 in selenate sensitivity; they further suggested that root growth and potentially the root tip activity might be a specific target of selenate toxicity in Arabidopsis. PMID:17208959

  9. Hydroxyproline O-arabinosyltransferase mutants oppositely alter tip growth in Arabidopsis thaliana and Physcomitrella patens.

    PubMed

    MacAlister, Cora A; Ortiz-Ramírez, Carlos; Becker, Jörg D; Feijó, José A; Lippman, Zachary B

    2016-01-01

    Hydroxyproline O-arabinosyltransferases (HPATs) are members of a small, deeply conserved family of plant-specific glycosyltransferases that add arabinose sugars to diverse proteins including cell wall-associated extensins and small signaling peptides. Recent genetic studies in flowering plants suggest that different HPAT homologs have been co-opted to function in diverse species-specific developmental contexts. However, nothing is known about the roles of HPATs in basal plants. We show that complete loss of HPAT function in Arabidopsis thaliana and the moss Physcomitrella patens results in a shared defect in gametophytic tip cell growth. Arabidopsis hpat1/2/3 triple knockout mutants suffer from a strong male sterility defect as a consequence of pollen tubes that fail to fully elongate following pollination. Knocking out the two HPAT genes of Physcomitrella results in larger multicellular filamentous networks due to increased elongation of protonemal tip cells. Physcomitrella hpat mutants lack cell-wall associated hydroxyproline arabinosides and can be rescued with exogenous cellulose, while global expression profiling shows that cell wall-associated genes are severely misexpressed, implicating a defect in cell wall formation during tip growth. Our findings point to a major role for HPATs in influencing cell elongation during tip growth in plants.

  10. Interaction of root gravitropism and phototropism in Arabidopsis wild-type and starchless mutants

    NASA Technical Reports Server (NTRS)

    Vitha, S.; Zhao, L.; Sack, F. D.

    2000-01-01

    Root gravitropism in wild-type Arabidopsis and in two starchless mutants, pgm1-1 and adg1-1, was evaluated as a function of light position to determine the relative strengths of negative phototropism and of gravitropism and how much phototropism affects gravitropic measurements. Gravitropism was stronger than phototropism in some but not all light positions in wild-type roots grown for an extended period, indicating that the relationship between the two tropisms is more complex than previously reported. Root phototropism significantly influenced the time course of gravitropic curvature and the two measures of sensitivity. Light from above during horizontal exposure overestimated all three parameters for all three genotypes except the wild-type perception time. At the irradiance used (80 micromol m(-2) s(-1)), the shortest periods of illumination found to exaggerate gravitropism were 45 min of continuous illumination and 2-min doses of intermittent illumination. By growing roots in circumlateral light or by gravistimulating in the dark, corrected values were obtained for each gravitropic parameter. Roots of both starchless mutants were determined to be about three times less sensitive than prior estimates. This study demonstrates the importance of accounting for phototropism in the design of root gravitropism experiments in Arabidopsis.

  11. Mutant analysis, protein-protein interactions and subcellular localization of the Arabidopsis B sister (ABS) protein.

    PubMed

    Kaufmann, Kerstin; Anfang, Nicole; Saedler, Heinz; Theissen, Günter

    2005-09-01

    Recently, close relatives of class B floral homeotic genes, termed B(sister) genes, have been identified in both angiosperms and gymnosperms. In contrast to the B genes themselves, B(sister) genes are exclusively expressed in female reproductive organs, especially in the envelopes or integuments surrounding the ovules. This suggests an important ancient function in ovule or seed development for B(sister) genes, which has been conserved for about 300 million years. However, investigation of the first loss-of-function mutant for a B(sister) gene (ABS/TT16 from Arabidopsis) revealed only a weak phenotype affecting endothelium formation. Here, we present an analysis of two additional mutant alleles, which corroborates this weak phenotype. Transgenic plants that ectopically express ABS show changes in the growth and identity of floral organs, suggesting that ABS can interact with floral homeotic proteins. Yeast-two-hybrid and three-hybrid analyses indicated that ABS can form dimers with SEPALLATA (SEP) floral homeotic proteins and multimeric complexes that also include the AGAMOUS-like proteins SEEDSTICK (STK) or SHATTERPROOF1/2 (SHP1, SHP2). These data suggest that the formation of multimeric transcription factor complexes might be a general phenomenon among MIKC-type MADS-domain proteins in angiosperms. Heterodimerization of ABS with SEP3 was confirmed by gel retardation assays. Fusion proteins tagged with CFP (Cyan Fluorescent Protein) and YFP (Yellow Fluorescent Protein) in Arabidopsis protoplasts showed that ABS is localized in the nucleus. Phylogenetic analysis revealed the presence of a structurally deviant, but closely related, paralogue of ABS in the Arabidopsis genome. Thus the evolutionary developmental genetics of B(sister) genes can probably only be understood as part of a complex and redundant gene network that may govern ovule formation in a conserved manner, which has yet to be fully explored.

  12. A User’s Guide to the Arabidopsis T-DNA Insertional Mutant Collections

    PubMed Central

    O'Malley, Ronan C.; Barragan, Cesar C.; Ecker, Joseph R.

    2016-01-01

    Summary The T-DNA sequence-indexed mutant collections contain insertional mutants for most Arabidopsis thaliana genes and have played an important role in plant biology research for almost two decades. By providing a large source of mutant alleles for in vivo characterization of gene function, this resource has been leveraged thousands of times to study a wide-range of problems in plant biology. Our primary goal in this chapter is to provide a general guide to strategies for the effective use of the data and materials in these collections. To do this, we provide a general introduction to the T-DNA insertional sequence-indexed mutant collections with a focus on how best to use the available data sources for good line selection. As isolation of a homozygous line is a common next step once a potential disruption line has been identified, the second half of the chapter will provide a step-by-step guide for the design and implementation of a T-DNA genotyping pipeline. Finally, we describe interpretation of genotyping results and include a troubleshooting section for common types of segregation distortions that we have observed. In this chapter we introduce both basic concepts and specific applications to new and more experienced users of the collections for the design and implementation of small- to large-scale genotyping pipelines. PMID:25757780

  13. Arabidopsis haiku mutants reveal new controls of seed size by endosperm.

    PubMed

    Garcia, Damien; Saingery, Virginie; Chambrier, Pierre; Mayer, Ulrike; Jürgens, Gerd; Berger, Frédéric

    2003-04-01

    In flowering plants, maternal seed integument encloses the embryo and the endosperm, which are both derived from double fertilization. Although the development of these three components must be coordinated, we have limited knowledge of mechanisms involved in such coordination. The endosperm may play a central role in these mechanisms as epigenetic modifications of endosperm development, via imbalance of dosage between maternal and paternal genomes, affecting both the embryo and the integument. To identify targets of such epigenetic controls, we designed a genetic screen in Arabidopsis for mutants that phenocopy the effects of dosage imbalance in the endosperm. The two mutants haiku 1 and haiku 2 produce seed of reduced size that resemble seed with maternal excess in the maternal/paternal dosage. Homozygous haiku seed develop into plants indistinguishable from wild type. Each mutation is sporophytic recessive, and double-mutant analysis suggests that both mutations affect the same genetic pathway. The endosperm of haiku mutants shows a premature arrest of increase in size that causes precocious cellularization of the syncytial endosperm. Reduction of seed size in haiku results from coordinated reduction of endosperm size, embryo proliferation, and cell elongation of the maternally derived integument. We present further evidence for a control of integument development mediated by endosperm-derived signals.

  14. Identification of plant defence regulators through transcriptional profiling of Arabidopsis thaliana cdd1 mutant.

    PubMed

    Swain, Swadhin; Singh, Nidhi; Nandi, Ashis Kumar

    2015-03-01

    A sustainable balance between defence and growth is essential for optimal fitness under pathogen stress. Plants activate immune response at the cost of normal metabolic requirements. Thus, plants that constitutively activate defence are deprived of growth. Arabidopsis thaliana mutant constitutive defence without defect in growth and development1 (cdd1) is an exception. The cdd1 mutant is constitutive for salicylic acid accumulation, signalling, and defence against biotrophic and hemibiotrophic pathogens, without having much impact on growth. Thus, cdd1 offers an ideal genetic background to identify novel regulators of plant defence. Here we report the differential gene expression profile between cdd1 and wild-type plants as obtained by microarray hybridization. Expression of several defence-related genes also supports constitutive activation of defence in cdd1. We screened T-DNA insertion mutant lines of selected genes, for resistance against virulent bacterial pathogen Pseudomonas syringae pv. tomato DC3000 (Pst DC3000). Through bacterial resistance, callose deposition and pathogenesis-associated expression analyses, we identified four novel regulators of plant defence. Resistance levels in the mutants suggest that At2g19810 and [rom] At5g05790 are positive regulators, whereas At1g61370 and At3g42790 are negative regulators of plant defence against bacterial pathogens.

  15. An In Vivo Quantitative Comparison of Photoprotection in Arabidopsis Xanthophyll Mutants

    PubMed Central

    Ware, Maxwell A.; Dall’Osto, Luca; Ruban, Alexander V.

    2016-01-01

    Contribution of different LHCII antenna carotenoids to protective NPQ (pNPQ) were tested using a range of xanthophyll biosynthesis mutants of Arabidopsis: plants were either devoid of lutein (lut2), violaxanthin (npq2), or synthesized a single xanthophyll species, namely violaxanthin (aba4npq1lut2), zeaxanthin (npq2lut2), or lutein (chy1chy2lut5). A novel pulse amplitude modulated (PAM) fluorescence analysis procedure, that used a gradually increasing actinic light intensity, allowed the efficiency of pNPQ to be tested using the photochemical quenching (qP) parameter measured in the dark (qPd). Furthermore, the yield of photosystem II (ΦPSII) was calculated, and the light intensity which induces photoinhibition in 50% of leaves for each mutant was ascertained. Photoprotective capacities of each xanthophyll were quantified, taking into account chlorophyll a/b ratios and excitation pressure. Here, light tolerance, pNPQ capacity, and ΦPSII were highest in wild type plants. Of the carotenoid mutants, lut2 (lutein-deficient) plants had the highest light tolerance, and the joint the highest ΦPSII with violaxanthin only plants. We conclude that all studied mutants possess pNPQ and a more complete composition of xanthophylls in their natural binding sites is the most important factor governing photoprotection, rather than any one specific xanthophyll suggesting a strong structural effect of the molecules upon the LHCII antenna organization and discuss the results significance for future crop development. PMID:27446097

  16. 5-Fluoroindole Resistance Identifies Tryptophan Synthase Beta Subunit Mutants in Arabidopsis Thaliana

    PubMed Central

    Barczak, A. J.; Zhao, J.; Pruitt, K. D.; Last, R. L.

    1995-01-01

    A study of the biochemical genetics of the Arabidopsis thaliana tryptophan synthase beta subunit was initiated by characterization of mutants resistant to the inhibitor 5-fluoroindole. Thirteen recessive mutations were recovered that are allelic to trp2-1, a mutation in the more highly expressed of duplicate tryptophan synthase beta subunit genes (TSB1). Ten of these mutations (trp2-2 through trp2-11) cause a tryptophan requirement (auxotrophs), whereas three (trp2-100 through trp2-102) remain tryptophan prototrophs. The mutations cause a variety of changes in tryptophan synthase beta expression. For example, two mutations (trp2-5 and trp2-8) cause dramatically reduced accumulation of TSB mRNA and immunologically detectable protein, whereas trp2-10 is associated with increased mRNA and protein. A correlation exists between the quantity of mutant beta and wild-type alpha subunit levels in the trp2 mutant plants, suggesting that the synthesis of these proteins is coordinated or that the quantity or structure of the beta subunit influences the stability of the alpha protein. The level of immunologically detectable anthranilate synthase alpha subunit protein is increased in the trp2 mutants, suggesting the possibility of regulation of anthranilate synthase levels in response to tryptophan limitation. PMID:7635295

  17. Root graviresponsiveness and cellular differentiation in wild-type and a starchless mutant of Arabidopsis thaliana

    NASA Technical Reports Server (NTRS)

    Moore, R.

    1989-01-01

    Primary roots of a starchless mutant of Arabidopsis thaliana L. are strongly graviresponsive despite lacking amyloplasts in their columella cells. The ultrastructures of calyptrogen and peripheral cells in wild-type as compared to mutant seedlings are not significantly different. The largest difference in cellular differentiation in caps of mutant and wild-type roots is the relative volume of plastids in columella cells. Plastids occupy 12.3% of the volume of columella cells in wild-type seedlings, but only 3.69% of columella cells in mutant seedlings. These results indicate that: (1) amyloplasts and starch are not necessary for root graviresponsiveness; (2) the increase in relative volume of plastids that usually accompanies differentiation of columella cells is not necessary for root graviresponsiveness; and (3) the absence of starch and amyloplasts does not affect the structure of calyptrogen (i.e. meristematic) and secretory (i.e. peripheral) cells in root caps. These results are discussed relative to proposed models for root gravitropism.

  18. A user's guide to the Arabidopsis T-DNA insertion mutant collections.

    PubMed

    O'Malley, Ronan C; Barragan, Cesar C; Ecker, Joseph R

    2015-01-01

    The T-DNA sequence-indexed mutant collections contain insertional mutants for most Arabidopsis thaliana genes and have played an important role in plant biology research for almost two decades. By providing a large source of mutant alleles for in vivo characterization of gene function, this resource has been leveraged thousands of times to study a wide range of problems in plant biology. Our primary goal in this chapter is to provide a general guide to strategies for the effective use of the data and materials in these collections. To do this, we provide a general introduction to the T-DNA insertional sequence-indexed mutant collections with a focus on how best to use the available data sources for good line selection. As isolation of a homozygous line is a common next step once a potential disruption line has been identified, the second half of the chapter provides a step-by-step guide for the design and implementation of a T-DNA genotyping pipeline. Finally, we describe interpretation of genotyping results and include a troubleshooting section for common types of segregation distortions that we have observed. In this chapter we introduce both basic concepts and specific applications to both new and more experienced users of the collections for the design and implementation of small- to large-scale genotyping pipelines.

  19. A large-scale genetic screen for mutants with altered salicylic acid accumulation in Arabidopsis.

    PubMed

    Ding, Yezhang; Shaholli, Danjela; Mou, Zhonglin

    2014-01-01

    Salicylic acid (SA) is a key defense signal molecule against biotrophic and hemibiotrophic pathogens in plants, but how SA is synthesized in plant cells still remains elusive. Identification of new components involved in pathogen-induced SA accumulation would help address this question. To this end, we performed a large-scale genetic screen for mutants with altered SA accumulation during pathogen infection in Arabidopsis using a bacterial biosensor Acinetobacter sp. ADPWH_lux-based SA quantification method. A total of 35,000 M2 plants in the npr1-3 mutant background have been individually analyzed for the bacterial pathogen Pseudomonas syringae pv. maculicola (Psm) ES4326-induced SA accumulation. Among the mutants isolated, 19 had SA levels lower than npr1 (sln) and two exhibited increased SA accumulation in npr1 (isn). Complementation tests revealed that seven of the sln mutants are new alleles of eds5/sid1, two are sid2/eds16 alleles, one is allelic to pad4, and the remaining seven sln and two isn mutants are new non-allelic SA accumulation mutants. Interestingly, a large group of mutants (in the npr1-3 background), in which Psm ES4326-induced SA levels were similar to those in the wild-type Columbia plants, were identified, suggesting that the signaling network fine-tuning pathogen-induced SA accumulation is complex. We further characterized the sln1 single mutant and found that Psm ES4326-induced defense responses were compromised in this mutant. These defense response defects could be rescued by exogenous SA, suggesting that SLN1 functions upstream of SA. The sln1 mutation was mapped to a region on the north arm of chromosome I, which contains no known genes regulating pathogen-induced SA accumulation, indicating that SLN1 likely encodes a new regulator of SA biosynthesis. Thus, the new sln and isn mutants identified in this genetic screen are valuable for dissecting the molecular mechanisms underlying pathogen-induced SA accumulation in plants.

  20. Alterations of the degree of xylan acetylation in Arabidopsis xylan mutants

    PubMed Central

    Lee, Chanhui; Teng, Quincy; Zhong, Ruiqin; Ye, Zheng-Hua

    2014-01-01

    Xylan is the second most abundant polysaccharide in secondary walls of dicot plants and one of its structural features is the high degree of acetylation of xylosyl residues. In Arabidopsis, about 60% of xylosyl residues in xylan are acetylated and the biochemical mechanisms controlling xylan acetylation are largely unknown. A recent report by Yuan et al. (2013) revealed the essential role of a DUF231 domain-containing protein, ESKIMO1 (ESK1), in xylan acetylation in Arabidopsis as the esk1 mutation caused specific reductions in the degree of xylan 2-O or 3-O-monoacetylation and in the activity of xylan acetyltransferase. Interestingly, the esk1 mutation also resulted in an elevation of glucuronic acid (GlcA) substitutions in xylan. Since GlcA substitutions in xylan occur at the O-2 position of xylosyl residues, it is plausible that the increase in GlcA substitutions in the esk1 mutant is attributed to the reduction in acetylation at O-2 of xylosyl residues, which renders more O-2 positions available for GlcA substitutions. Here, we investigated the effect of removal of GlcA substitutions on the degree of xylan acetylation. We found that a complete loss of GlcA substitutions in the xylan of the gux1/2/3 triple mutant led to a significant increase in the degree of xylan acetylation, indicating that xylan acetyltransferases and glucuronyltransferases compete with each other for xylosyl residues for their acetylation or GlcA substitutions in planta. In addition, detailed structure analysis of xylan from the rwa1/2/3/4 quadruple mutant revealed that it had a uniform reduction of acetyl substitutions at different positions of the xylosyl residues, which is consistent with the proposed role of RWAs as acetyl coenzyme A transporters. The significance of these findings is discussed. PMID:24518588

  1. Arabidopsis carotenoid mutants demonstrate that lutein is not essential for photosynthesis in higher plants.

    PubMed Central

    Pogson, B; McDonald, K A; Truong, M; Britton, G; DellaPenna, D

    1996-01-01

    Lutein, a dihydroxy beta, epsilon-carotenoid, is the predominant carotenoid in photosynthetic plant tissue and plays a critical role in light-harvesting complex assembly and function. To further understand lutein synthesis and function, we isolated four lutein-deficient mutants of Arabidopsis that define two loci, lut1 and lut2 (for lutein deficient). These loci are required for lutein biosynthesis but not for the biosynthesis of beta, beta-carotenoids. The lut1 mutations are recessive, accumulate high levels of zeinoxanthin, which is the immediate precursor of lutein, and define lut1 as a disruption in epsilon ring hydroxylation. The lut2 mutations are semidominant, and their biochemical phenotype is consistent with a disruption of epsilon ring cyclization. The lut2 locus cosegregates with the recently isolated epsilon cyclase gene, thus, providing additional evidence that the lut2 alleles are mutations in the epsilon cyclase gene. It appears likely that the epsilon cyclase is a key step in regulating lutein levels and the ratio of lutein to beta,beta-carotenoids. Surprisingly, despite the absence of lutein, neither the lut1 nor lut2 mutation causes a visible deleterious phenotype or altered chlorophyll content, but both mutants have significantly higher levels of beta, beta-carotenoids. In particular, there is a stable increase in the xanthophyll cycle pigments (violaxanthin, antheraxanthin, and zeaxanthin) in both lut1 and lut2 mutants as well as an increase in zeinoxanthin in lut1 and beta-carotene in lut2. The accumulation of specific carotenoids is discussed as it pertains to the regulation of carotenoid biosynthesis and incorporation into the photosynthetic apparatus. Presumably, particular beta, beta-carotenoids are able to compensate functionally and structurally for lutein in the photosystems of Arabidopsis. PMID:8837513

  2. The phenotype of Arabidopsis thaliana det1 mutants suggest a role for cytokinins in greening. Progress report

    SciTech Connect

    Chory, J.; Aguilar, N.; Peto, C.A.

    1990-12-31

    When grown in the absence of light, the det1 mutants of Arabidopsis thaliana develop characteristics of light-grown plants by morphological, cellular, and molecular criteria. Further, in light-grown plants, mutations in the DET1 gene affect cell-type-specific expression of light-regulated genes and the chloroplast developmental program. Here we show that the addition of exogenously added cytokinins (either 2-isopentenyl adenine, kinetin, or benzyladenine) to the growth medium of dark-germinated wild-type seedlings results in seedlings that resemble det1 mutants, instead of having the normal etiolated morphology. Like det1 mutants, these dark-grown seedlings now contain chloroplasts and have high levels of expression of genes that are normally ``light``-regulated. These results suggest an important role for cytokinins during greening of Arabidopsis, and may implicate cytokinin levels or an increased sensitivity to cytokinins as explanations for some of the observed phenotypes of det1 mutants.

  3. Mechanical touch responses of Arabidopsis TCH1-3 mutant roots on inclined hard-agar surface

    NASA Astrophysics Data System (ADS)

    Zha, Guodong; Wang, Bochu; Liu, Junyu; Yan, Jie; Zhu, Liqing; Yang, Xingyan

    2016-01-01

    The gravity-induced mechanical touch stimulus can affect plant root architecture. Mechanical touch responses of plant roots are an important aspect of plant root growth and development. Previous studies have reported that Arabidopsis TCH1-3 genes are involved in mechano-related events, how-ever, the physiological functions of TCH1-3 genes in Arabidopsis root mechanoresponses remain unclear. In the present study, we applied an inclined hard agar plate method to produce mechanical touch stimulus, and provided evidence that altered mechanical environment could influence root growth. Furthermore, tch1-3 Arabidopsis mutants were investigated on inclined agar surfaces to explore the functions of TCH1-3 genes on Arabidopsis root mechanoresponses. The results showed that two tch2 mutants, cml24-2 and cml24-4, exhibited significantly reduced root length, biased skewing, and decreased density of lateral root. In addition, primary root length and density of lateral root of tch3 (cml12-2) was significantly decreased on inclined agar surfaces. This study indicates that the tch2 and tch3 mutants are hypersensitive to mechanical touch stimulus, and TCH2 (CML24-2 and CML24-4) and TCH3 (CML12-2) genes may participate in the mechanical touch response of Arabidopsis roots.

  4. The gravity persistent signal (gps) Mutants of Arabidopsis: Insights into Gravitropic Signal Transduction

    NASA Astrophysics Data System (ADS)

    Wyatt, S.

    The gravitropic response of Arabidopsis stems is rapid with a visible within 30 min and vertical reorientation within 2 h. However, horizontal gravistimulation for 3 h at 4°C does not cause curvature. When the stems are subsequently placed in the vertical position at RT, they bend in response to the previous, horizontal gravistimulation. These results indicate that the gravity perception step can occur at 4°C, but that part of the response is sensitive to cold. At 4°C, starch-containing amyloplasts in the endodermis of the inflorescence stems sedimented normally but auxin transport was abolished indicating that the cold treatment affected early events of the signal transduction pathway that occur after amyloplast sedimentation but prior to auxin transport. The gps mutants of Arabidopsis are a unique group of mutants that respond abnormally after gravistimulation at 4°C. gps1 shows no response to the cold gravistimulation, gps2 bends the wrong way as compared to wild type and gps3 over responds, bending past the anticipated curvature. The mutants were selected from a T-DNA tagged population. Cloning strategies based on the tag have been employed to identify the genes disrupted. GPS1 was cloned using TAIL PCR and is At3g20130, a cytochrome P450, CYP705A22, of unknown function. GPS1p::GFP fusions are being used to determine temporal and spatial expression of GPS1. The mutation in gps3 appears to disrupt a non-coding region downstream of At1g43950 No function has yet been determined for this region, but it appears that the mutation disrupts transcription of a transcription factor homologous to the DNA binding domain of an auxin response factor (ARF) 9-like protein. The identity of GPS2 is as yet unknown. The gps mutants represent potentially three independent aspects of signal transduction in the gravitropic response: perception or retention of the gravity signal (gps1), determination of the polarity of the response (gps2), and the tissue specificity of the

  5. Life without complex I: proteome analyses of an Arabidopsis mutant lacking the mitochondrial NADH dehydrogenase complex.

    PubMed

    Fromm, Steffanie; Senkler, Jennifer; Eubel, Holger; Peterhänsel, Christoph; Braun, Hans-Peter

    2016-05-01

    The mitochondrial NADH dehydrogenase complex (complex I) is of particular importance for the respiratory chain in mitochondria. It is the major electron entry site for the mitochondrial electron transport chain (mETC) and therefore of great significance for mitochondrial ATP generation. We recently described an Arabidopsis thaliana double-mutant lacking the genes encoding the carbonic anhydrases CA1 and CA2, which both form part of a plant-specific 'carbonic anhydrase domain' of mitochondrial complex I. The mutant lacks complex I completely. Here we report extended analyses for systematically characterizing the proteome of the ca1ca2 mutant. Using various proteomic tools, we show that lack of complex I causes reorganization of the cellular respiration system. Reduced electron entry into the respiratory chain at the first segment of the mETC leads to induction of complexes II and IV as well as alternative oxidase. Increased electron entry at later segments of the mETC requires an increase in oxidation of organic substrates. This is reflected by higher abundance of proteins involved in glycolysis, the tricarboxylic acid cycle and branched-chain amino acid catabolism. Proteins involved in the light reaction of photosynthesis, the Calvin cycle, tetrapyrrole biosynthesis, and photorespiration are clearly reduced, contributing to the significant delay in growth and development of the double-mutant. Finally, enzymes involved in defense against reactive oxygen species and stress symptoms are much induced. These together with previously reported insights into the function of plant complex I, which were obtained by analysing other complex I mutants, are integrated in order to comprehensively describe 'life without complex I'.

  6. Life without complex I: proteome analyses of an Arabidopsis mutant lacking the mitochondrial NADH dehydrogenase complex

    PubMed Central

    Fromm, Steffanie; Senkler, Jennifer; Eubel, Holger; Peterhänsel, Christoph; Braun, Hans-Peter

    2016-01-01

    The mitochondrial NADH dehydrogenase complex (complex I) is of particular importance for the respiratory chain in mitochondria. It is the major electron entry site for the mitochondrial electron transport chain (mETC) and therefore of great significance for mitochondrial ATP generation. We recently described an Arabidopsis thaliana double-mutant lacking the genes encoding the carbonic anhydrases CA1 and CA2, which both form part of a plant-specific ‘carbonic anhydrase domain’ of mitochondrial complex I. The mutant lacks complex I completely. Here we report extended analyses for systematically characterizing the proteome of the ca1ca2 mutant. Using various proteomic tools, we show that lack of complex I causes reorganization of the cellular respiration system. Reduced electron entry into the respiratory chain at the first segment of the mETC leads to induction of complexes II and IV as well as alternative oxidase. Increased electron entry at later segments of the mETC requires an increase in oxidation of organic substrates. This is reflected by higher abundance of proteins involved in glycolysis, the tricarboxylic acid cycle and branched-chain amino acid catabolism. Proteins involved in the light reaction of photosynthesis, the Calvin cycle, tetrapyrrole biosynthesis, and photorespiration are clearly reduced, contributing to the significant delay in growth and development of the double-mutant. Finally, enzymes involved in defense against reactive oxygen species and stress symptoms are much induced. These together with previously reported insights into the function of plant complex I, which were obtained by analysing other complex I mutants, are integrated in order to comprehensively describe ‘life without complex I’. PMID:27122571

  7. Leaf biomechanical properties in Arabidopsis thaliana polysaccharide mutants affect drought survival.

    PubMed

    Balsamo, Ronald; Boak, Merewyn; Nagle, Kayla; Peethambaran, Bela; Layton, Bradley

    2015-11-26

    Individual sugars are the building blocks of cell wall polysaccharides, which in turn comprise a plant׳s overall architectural structure. But which sugars play the most prominent role in maintaining a plant׳s mechanical stability during large cellular deformations induced by drought? We investigated the individual contributions of several genes that are involved in the synthesis of monosaccharides which are important for cell wall structure. We then measured drought tolerance and mechanical integrity during simulated drought in Arabidopsis thaliana. To assess mechanical properties, we designed a small-scale tensile tester for measuring failure strain, ultimate tensile stress, work to failure, toughness, and elastic modulus of 6-week-old leaves in both hydrated and drought-simulated states. Col-0 mutants used in this study include those deficient in lignin, cellulose, components of hemicellulose such as xylose and fucose, the pectic components arabinose and rhamnose, as well as mutants with enhanced arabinose and total pectin content. We found that drought tolerance is correlated to the mechanical and architectural stability of leaves as they experience dehydration. Of the mutants, S096418 with mutations for reduced xylose and galactose was the least drought tolerant, while the arabinose-altered CS8578 mutants were the least affected by water loss. There were also notable correlations between drought tolerance and mechanical properties in the diminished rhamnose mutant, CS8575 and the dehydrogenase-disrupted S120106. Our findings suggest that components of hemicellulose and pectins affect leaf biomechanical properties and may play an important role in the ability of this model system to survive drought.

  8. Photosynthetic properties of an Arabidopsis thaliana mutant possessing a defective PsbS gene.

    PubMed

    Peterson, R B; Havir, E A

    2001-11-01

    We describe the properties of npq4-9, a new mutant of Arabidopsis thaliana (L.) Heynh. with reduced nonphotochemical quenching (NPQ) capacity that possesses a single amino acid substitution in the PsbS gene encoding PSII-S, a ubiquitous pigment-binding protein associated with photosystem II (PSII) of higher plants. Growth, photosynthetic pigment contents, and levels of the major PSII antenna proteins were not affected by npq4-9. Although the extent of de-epoxidatin of violaxanthin to antheraxanthin plus zeaxanthin for leaves displaying the mutant phenotype equaled or exceeded that observed for the wild type, the relative effectiveness of de-epoxidized xanthophylls in promoting NPQ was consistently lower for the mutant. Energy partitioning in PSII was analyzed in terms of the competition for singlet chlorophyll a among the processes of fluorescence, thermal dissipation, and photochemistry. The key processes of NPQ and photochemistry in open PSII centers are represented by the relative in vivo rate constants kN and kP0, respectively. The magnitude of kP0 in normal leaves declined only slightly with increasing kN, consistent with localization of NPQ primarily in the antenna complex. Conversely, a highly significant linear decline in kP0 with increasing kN was observed for the mutant, consistent with a role for the PSII reaction center in the NPQ mechanism. Although the PSII absorption cross-section for white light was not significantly different relative to that of the wild type, PSII quantum yield was significantly lower in the mutant. The resulting lower capacity for linear electron transport in the mutant primarily affected reduction of terminal acceptors other than CO2. Parallel measurements of fluorescence and in vivo absorbance at 820 nm indicated a consistently higher steady-state level of reduction of PSII acceptors and accumulation of P700+ for the mutant. This suggests that inter-photosystem electron transport in the mutant is restricted either by a higher

  9. Physiological and genetic analysis of Arabidopsis thaliana anthocyanin biosynthesis mutants under chronic adverse environmental conditions.

    PubMed

    Misyura, Maksym; Colasanti, Joseph; Rothstein, Steven J

    2013-01-01

    Anthocyanin production is a characteristic response of flowering plants to unfavourable environmental conditions. The potential roles of flavonoids and anthocyanins in plant growth were investigated by growing Arabidopsis thaliana anthocyanin production mutants (transparent testa) under limiting nitrogen and high light conditions. Inability to produce kaempferol or subsequent intermediate compounds by some transparent testa lines was correlated with less biomass accumulation in mature plants compared with wild-type control plants under all growth conditions tested. However, under both limiting nitrogen and high light chronic stress conditions, mutant lines defective in later steps of the anthocyanin production pathway produced the same or more biomass than wild-type plants. No difference in senescence between transparent testa and wild-type plants was found using chlorophyll catabolism and SAG12 expression measurements, and no mutants were impaired in the ability to remobilize nutrients from the vegetative to reproductive tissues. Moreover, the absence of anthocyanin and/or upstream flavonoids does not affect the ability of plants to respond to limiting nitrogen by reducing photosynthetic capacity. These results support a role for kaempferol and quercetin accumulation in normal plant growth and development. Further, the absence of anthocyanins has no effect on plant growth under the chronic stress conditions tested.

  10. Ascorbate-Deficient vtc2 Mutants in Arabidopsis Do Not Exhibit Decreased Growth.

    PubMed

    Lim, Benson; Smirnoff, Nicholas; Cobbett, Christopher S; Golz, John F

    2016-01-01

    In higher plants the L-galactose pathway represents the major route for ascorbate biosynthesis. The first committed step of this pathway is catalyzed by the enzyme GDP-L-galactose phosphorylase and is encoded by two paralogs in Arabidopsis - VITAMIN C2 (VTC2) and VTC5. The first mutant of this enzyme, vtc2-1, isolated via an EMS mutagenesis screen, has approximately 20-30% of wildtype ascorbate levels and has been reported to have decreased growth under standard laboratory conditions. Here, we show that a T-DNA insertion into the VTC2 causes a similar reduction in ascorbate levels, but does not greatly affect plant growth. Subsequent segregation analysis revealed the growth defects of vtc2-1 mutants segregate independently of the vtc2-1 mutation. These observations suggest that it is the presence of an independent cryptic mutation that affects growth of vtc2-1 mutants, and not the 70-80% decrease in ascorbate levels that has been assumed in past studies.

  11. Physiological and genetic analysis of Arabidopsis thaliana anthocyanin biosynthesis mutants under chronic adverse environmental conditions

    PubMed Central

    Rothstein, Steven J.

    2013-01-01

    Anthocyanin production is a characteristic response of flowering plants to unfavourable environmental conditions. The potential roles of flavonoids and anthocyanins in plant growth were investigated by growing Arabidopsis thaliana anthocyanin production mutants (transparent testa) under limiting nitrogen and high light conditions. Inability to produce kaempferol or subsequent intermediate compounds by some transparent testa lines was correlated with less biomass accumulation in mature plants compared with wild-type control plants under all growth conditions tested. However, under both limiting nitrogen and high light chronic stress conditions, mutant lines defective in later steps of the anthocyanin production pathway produced the same or more biomass than wild-type plants. No difference in senescence between transparent testa and wild-type plants was found using chlorophyll catabolism and SAG12 expression measurements, and no mutants were impaired in the ability to remobilize nutrients from the vegetative to reproductive tissues. Moreover, the absence of anthocyanin and/or upstream flavonoids does not affect the ability of plants to respond to limiting nitrogen by reducing photosynthetic capacity. These results support a role for kaempferol and quercetin accumulation in normal plant growth and development. Further, the absence of anthocyanins has no effect on plant growth under the chronic stress conditions tested. PMID:23162120

  12. An Arabidopsis photolyase mutant is hypersensitive to ultraviolet-B radiation

    PubMed Central

    Landry, Laurie G.; Stapleton, Ann E.; Lim, Jackie; Hoffman, Peter; Hays, John B.; Walbot, Virginia; Last, Robert L.

    1997-01-01

    Photolyases are DNA repair enzymes that use energy from blue light to repair pyrimidine dimers. We report the isolation of an Arabidopsis thaliana mutant (uvr2-1) that is defective in photorepair of cyclobutylpyrimidine dimers (CPDs). Whereas uvr2-1 is indistinguishable from wild type in the absence of UV light, low UV-B levels inhibit growth and cause leaf necrosis. uvr2-1 is more sensitive to UV-B than wild type when placed under white light after UV-B treatment. In contrast, recovery in darkness or in light lacking photoreactivating blue light results in equal injury in uvr2-1 and wild type. The uvr2-1 mutant is unable to remove CPDs in vivo, and plant extracts lack detectable photolyase activity. This recessive mutation segregates as a single gene located near the top of chromosome 1, and is a structural gene mutation in the type II CPD photolyase PHR1. This mutant provides evidence that CPD photolyase is required for plant survival in the presence of UV-B light. PMID:8990208

  13. Analysis of Vascular Development in the hydra Sterol Biosynthetic Mutants of Arabidopsis

    PubMed Central

    Pullen, Margaret; Clark, Nick; Zarinkamar, Fatemeh; Topping, Jennifer; Lindsey, Keith

    2010-01-01

    Background The control of vascular tissue development in plants is influenced by diverse hormonal signals, but their interactions during this process are not well understood. Wild-type sterol profiles are essential for growth, tissue patterning and signalling processes in plant development, and are required for regulated vascular patterning. Methodology/Principal Findings Here we investigate the roles of sterols in vascular tissue development, through an analysis of the Arabidopsis mutants hydra1 and fackel/hydra2, which are defective in the enzymes sterol isomerase and sterol C-14 reductase respectively. We show that defective vascular patterning in the shoot is associated with ectopic cell divisions. Expression of the auxin-regulated AtHB8 homeobox gene is disrupted in mutant embryos and seedlings, associated with variably incomplete vascular strand formation and duplication of the longitudinal axis. Misexpression of the auxin reporter proIAA2∶GUS and mislocalization of PIN proteins occurs in the mutants. Introduction of the ethylene-insensitive ein2 mutation partially rescues defective cell division, localization of PIN proteins, and vascular strand development. Conclusions The results support a model in which sterols are required for correct auxin and ethylene crosstalk to regulate PIN localization, auxin distribution and AtHB8 expression, necessary for correct vascular development. PMID:20808926

  14. Mutants of Arabidopsis thaliana with decreased amplitude in their phototropic response

    NASA Technical Reports Server (NTRS)

    Khurana, J. P.; Ren, Z.; Steinitz, B.; Parks, B.; Best, T. R.; Poff, K. L.

    1989-01-01

    Two mutants of Arabidopsis thaliana have been identified with decreased phototropism to 450-nanometer light. Fluence-response relationships for these strains (ZR8 and ZR19) to single and multiple flashes of light show thresholds, curve shapes, and fluence for maximum curvature in first positive' phototropism which are the same as those of the wild type. Similarly, there is no alteration from the wild type in the kinetics of curvature or in the optimum dark period separating sequential flashes in a multiple flash regimen. In addition, in both strains, gravitropism is decreased compared to the wild type by an amount which is comparable to the decrease in phototropism. Based on reciprocal backcrosses, it appears that the alteration is due to a recessive nuclear mutation. It is suggested that ZR8 and ZR19 represent alterations in some step analogous to an amplifier, downstream of the photoreceptor pigment, and common to both phototropism and gravitropism.

  15. Catalase function in plants: a focus on Arabidopsis mutants as stress-mimic models.

    PubMed

    Mhamdi, Amna; Queval, Guillaume; Chaouch, Sejir; Vanderauwera, Sandy; Van Breusegem, Frank; Noctor, Graham

    2010-10-01

    Hydrogen peroxide (H(2)O(2)) is an important signal molecule involved in plant development and environmental responses. Changes in H(2)O(2) availability can result from increased production or decreased metabolism. While plants contain several types of H(2)O(2)-metabolizing proteins, catalases are highly active enzymes that do not require cellular reductants as they primarily catalyse a dismutase reaction. This review provides an update on plant catalase genes, function, and subcellular localization, with a focus on recent information generated from studies on Arabidopsis. Original data are presented on Arabidopsis catalase single and double mutants, and the use of some of these lines as model systems to investigate the outcome of increases in intracellular H(2)O(2) are discussed. Particular attention is paid to interactions with cell thiol-disulphide status; the use of catalase-deficient plants to probe the apparent redundancy of reductive H(2)O(2)-metabolizing pathways; the importance of irradiance and growth daylength in determining the outcomes of catalase deficiency; and the induction of pathogenesis-related responses in catalase-deficient lines. Within the context of strategies aimed at understanding and engineering plant stress responses, the review also considers whether changes in catalase activities in wild-type plants are likely to be a significant part of plant responses to changes in environmental conditions or biotic challenge.

  16. Arabidopsis DNA polymerase lambda mutant is mildly sensitive to DNA double strand breaks but defective in integration of a transgene.

    PubMed

    Furukawa, Tomoyuki; Angelis, Karel J; Britt, Anne B

    2015-01-01

    The DNA double-strand break (DSB) is a critical type of damage, and can be induced by both endogenous sources (e.g., errors of oxidative metabolism, transposable elements, programmed meiotic breaks, or perturbation of the DNA replication fork) and exogenous sources (e.g., ionizing radiation or radiomimetic chemicals). Although higher plants, like mammals, are thought to preferentially repair DSBs via nonhomologous end joining (NHEJ), much remains unclear about plant DSB repair pathways. Our reverse genetic approach suggests that DNA polymerase λ is involved in DSB repair in Arabidopsis. The Arabidopsis T-DNA insertion mutant (atpolλ-1) displayed sensitivity to both gamma-irradiation and treatment with radiomimetic reagents, but not to other DNA damaging treatments. The atpolλ-1 mutant showed a moderate sensitivity to DSBs, while Arabidopsis Ku70 and DNA ligase 4 mutants (atku70-3 and atlig4-2), both of which play critical roles in NHEJ, exhibited a hypersensitivity to these treatments. The atpolλ-1/atlig4-2 double mutant exhibited a higher sensitivity to DSBs than each single mutant, but the atku70/atpolλ-1 showed similar sensitivity to the atku70-3 mutant. We showed that transcription of the DNA ligase 1, DNA ligase 6, and Wee1 genes was quickly induced by BLM in several NHEJ deficient mutants in contrast to wild-type. Finally, the T-DNA transformation efficiency dropped in NHEJ deficient mutants and the lowest transformation efficiency was scored in the atpolλ-1/atlig4-2 double mutant. These results imply that AtPolλ is involved in both DSB repair and DNA damage response pathway.

  17. Arabidopsis DNA polymerase lambda mutant is mildly sensitive to DNA double strand breaks but defective in integration of a transgene

    PubMed Central

    Furukawa, Tomoyuki; Angelis, Karel J.; Britt, Anne B.

    2015-01-01

    The DNA double-strand break (DSB) is a critical type of damage, and can be induced by both endogenous sources (e.g., errors of oxidative metabolism, transposable elements, programmed meiotic breaks, or perturbation of the DNA replication fork) and exogenous sources (e.g., ionizing radiation or radiomimetic chemicals). Although higher plants, like mammals, are thought to preferentially repair DSBs via nonhomologous end joining (NHEJ), much remains unclear about plant DSB repair pathways. Our reverse genetic approach suggests that DNA polymerase λ is involved in DSB repair in Arabidopsis. The Arabidopsis T-DNA insertion mutant (atpolλ-1) displayed sensitivity to both gamma-irradiation and treatment with radiomimetic reagents, but not to other DNA damaging treatments. The atpolλ-1 mutant showed a moderate sensitivity to DSBs, while Arabidopsis Ku70 and DNA ligase 4 mutants (atku70-3 and atlig4-2), both of which play critical roles in NHEJ, exhibited a hypersensitivity to these treatments. The atpolλ-1/atlig4-2 double mutant exhibited a higher sensitivity to DSBs than each single mutant, but the atku70/atpolλ-1 showed similar sensitivity to the atku70-3 mutant. We showed that transcription of the DNA ligase 1, DNA ligase 6, and Wee1 genes was quickly induced by BLM in several NHEJ deficient mutants in contrast to wild-type. Finally, the T-DNA transformation efficiency dropped in NHEJ deficient mutants and the lowest transformation efficiency was scored in the atpolλ-1/atlig4-2 double mutant. These results imply that AtPolλ is involved in both DSB repair and DNA damage response pathway. PMID:26074930

  18. Lipid utilization, gluconeogenesis, and seedling growth in Arabidopsis mutants lacking the glyoxylate cycle enzyme malate synthase.

    PubMed

    Cornah, Johanna E; Germain, Véronique; Ward, Jane L; Beale, Michael H; Smith, Steven M

    2004-10-08

    The aim of this research was to test the role of the glyoxylate cycle enzyme malate synthase (MLS) in lipid utilization, gluconeogenesis, and seedling growth in Arabidopsis. We hypothesized that in the absence of MLS, succinate produced by isocitrate lyase (ICL) could still feed into the tricarboxylic acid cycle, whereas glyoxylate could be converted to sugars using enzymes of the photorespiratory pathway. To test this hypothesis we isolated knock-out mls mutants and studied their growth and metabolism in comparison to wild type and icl mutant seedlings. In contrast to icl seedlings, which grow slowly and are unable to convert lipid into sugars (Eastmond, P. J., Germain, V., Lange, P. R., Bryce, J. H., Smith, S. M. & Graham, I. A. (2000) Proc. Natl. Acad. Sci. U. S. A. 97, 5669-5674), mls seedlings grow faster, use their lipid more rapidly, and are better able to establish as plantlets. Transcriptome and metabolome analyses show that icl seedlings exhibit many features characteristic of carbohydrate starvation, whereas mls seedlings differ relatively little from wild type. In the light mls seedlings generate more sugars than icl seedlings, and when fed with [14C]acetate, 14C-labeling of sugars is three times greater than in icl seedlings and more than half that in wild type seedlings. The mls seedlings also accumulate more glycine and serine than icl or wild type seedlings, consistent with a diversion of glyoxylate into these intermediates of the photorespiratory pathway. We conclude that, in contrast to bacteria and fungi in which MLS is essential for gluconeogenesis from acetate or fatty acids, MLS is partially dispensable for lipid utilization and gluconeogenesis in Arabidopsis seedlings.

  19. Knockout mutants as a tool to identify the subunit composition of Arabidopsis glutamine synthetase isoforms.

    PubMed

    Dragićević, Milan; Todorović, Slađana; Bogdanović, Milica; Filipović, Biljana; Mišić, Danijela; Simonović, Ana

    2014-06-01

    Glutamine synthetase (GS) is a key enzyme in nitrogen assimilation, which catalyzes the formation of glutamine from ammonia and glutamate. Plant GS isoforms are multimeric enzymes, recently shown to be decamers. The Arabidopsis genome encodes five cytosolic (GS1) proteins labeled as GLN1;1 through GLN1;5 and one chloroplastic (GS2) isoform, GLN2;0. However, as many as 11 GS activity bands were resolved from different Arabidopsis tissues by Native PAGE and activity staining. Western analysis showed that all 11 isoforms are composed exclusively of 40 kDa GS1 subunits. Of five GS1 genes, only GLN1;1, GLN1;2 and GLN1;3 transcripts accumulated to significant levels in vegetative tissues, indicating that only subunits encoded by these three genes produce the 11-band zymogram. Even though the GS2 gene also had significant expression, the corresponding activity was not detected, probably due to inactivation. To resolve the subunit composition of 11 active GS1 isoforms, homozygous knockout mutants deficient in the expression of different GS1 genes were selected from the progeny of T-DNA insertional SALK and SAIL lines. Comparison of GS isoenzyme patterns of the selected GS1 knockout mutants indicated that all of the detected isoforms consist of varying proportions of GLN1;1, GLN1;2 and GLN1;3 subunits, and that GLN1;1 and GLN1;3, as well as GLN1;2 and GLN1;3 and possibly GLN1;1 and GLN1;2 proteins combine in all proportions to form active homo- and heterodecamers.

  20. On the role of a Lipid-Transfer Protein. Arabidopsis ltp3 mutant is compromised in germination and seedling growth.

    PubMed Central

    Pagnussat, Luciana A; Oyarburo, Natalia; Cimmino, Carlos; Pinedo, Marcela L; de la Canal, Laura

    2015-01-01

    Plant Lipid-Transfer Proteins (LTPs) exhibit the ability to reversibly bind/transport lipids in vitro. LTPs have been involved in diverse physiological processes but conclusive evidence on their role has only been presented for a few members, none of them related to seed physiology. Arabidopsis seeds rely on storage oil breakdown to supply carbon skeletons and energy for seedling growth. Here, Arabidopsis ltp3 mutant was analyzed for its ability to germinate and for seedling establishment. Ltp3 showed delayed germination and reduced germination frequency. Seedling growth appeared reduced in the mutant but this growth restriction was rescued by the addition of an exogenous carbon supply, suggesting a defective oil mobilization. Lipid breakdown analysis during seedling growth revealed a differential profile in the mutant compared to the wild type. The involvement of LTP3 in germination and seedling growth and its relationship with the lipid transfer ability of this protein is discussed. PMID:26479260

  1. On the role of a Lipid-Transfer Protein. Arabidopsis ltp3 mutant is compromised in germination and seedling growth.

    PubMed

    Pagnussat, Luciana A; Oyarburo, Natalia; Cimmino, Carlos; Pinedo, Marcela L; de la Canal, Laura

    2015-01-01

    Plant Lipid-Transfer Proteins (LTPs) exhibit the ability to reversibly bind/transport lipids in vitro. LTPs have been involved in diverse physiological processes but conclusive evidence on their role has only been presented for a few members, none of them related to seed physiology. Arabidopsis seeds rely on storage oil breakdown to supply carbon skeletons and energy for seedling growth. Here, Arabidopsis ltp3 mutant was analyzed for its ability to germinate and for seedling establishment. Ltp3 showed delayed germination and reduced germination frequency. Seedling growth appeared reduced in the mutant but this growth restriction was rescued by the addition of an exogenous carbon supply, suggesting a defective oil mobilization. Lipid breakdown analysis during seedling growth revealed a differential profile in the mutant compared to the wild type. The involvement of LTP3 in germination and seedling growth and its relationship with the lipid transfer ability of this protein is discussed.

  2. The Arabidopsis sickle Mutant Exhibits Altered Circadian Clock Responses to Cool Temperatures and Temperature-Dependent Alternative Splicing.

    PubMed

    Marshall, Carine M; Tartaglio, Virginia; Duarte, Maritza; Harmon, Frank G

    2016-10-01

    The circadian clock allows plants to anticipate and respond to daily changes in ambient temperature. Mechanisms establishing the timing of circadian rhythms in Arabidopsis thaliana through temperature entrainment remain unclear. Also incompletely understood is the temperature compensation mechanism that maintains consistent period length within a range of ambient temperatures. A genetic screen for Arabidopsis mutants affecting temperature regulation of the PSEUDO-RESPONSE REGULATOR7 promoter yielded a novel allele of the SICKLE (SIC) gene. This mutant, sic-3, and the existing sic-1 mutant both exhibit low-amplitude or arrhythmic expression of core circadian clock genes under cool ambient temperature cycles, but not under light-dark entrainment. sic mutants also lengthen free running period in a manner consistent with impaired temperature compensation. sic mutant alleles accumulate LATE ELONGATED HYPOCOTYL (LHY) and CIRCADIAN CLOCK ASSOCIATED1 (CCA1) splice variants, among other alternatively spliced transcripts, which is exacerbated by cool temperatures. The cca1-1 lhy-20 double mutant is epistatic to sic-3, indicating the LHY and CCA1 splice variants are needed for sic-3 circadian clock phenotypes. It is not expected that SIC is directly involved in the circadian clock mechanism; instead, SIC likely contributes to pre-mRNA metabolism, and the splice variants that accumulate in sic mutants likely affect the circadian clock response to cool ambient temperature.

  3. Functional Conservation of Plant Secondary Metabolic Enzymes Revealed by Complementation of Arabidopsis Flavonoid Mutants with Maize Genes1

    PubMed Central

    Dong, Xiaoyun; Braun, Edward L.; Grotewold, Erich

    2001-01-01

    Mutations in the transparent testa (tt) loci abolish pigment production in Arabidopsis seed coats. The TT4, TT5, and TT3 loci encode chalcone synthase, chalcone isomerase, and dihydroflavonol 4-reductase, respectively, which are essential for anthocyanin accumulation and may form a macromolecular complex. Here, we show that the products of the maize (Zea mays) C2, CHI1, and A1 genes complement Arabidopsis tt4, tt5, and tt3 mutants, restoring the ability of these mutants to accumulate pigments in seed coats and seedlings. Overexpression of the maize genes in wild-type Arabidopsis seedlings does not result in increased anthocyanin accumulation, suggesting that the steps catalyzed by these enzymes are not rate limiting in the conditions assayed. The expression of the maize A1 gene in the flavonoid 3′ hydroxylase Arabidopsis tt7 mutant resulted in an increased accumulation of pelargonidin. We conclude that enzymes involved in secondary metabolism can be functionally exchangeable between plants separated by large evolutionary distances. This is in sharp contrast to the notion that the more relaxed selective constrains to which secondary metabolic pathways are subjected is responsible for the rapid divergence of the corresponding enzymes. PMID:11553733

  4. Isolation and characterization of a novel ammonium overly sensitive mutant, amos2, in Arabidopsis thaliana.

    PubMed

    Li, Guangjie; Dong, Gangqiang; Li, Baohai; Li, Qing; Kronzucker, Herbert J; Shi, Weiming

    2012-02-01

    Ammonium (NH(4)(+)) toxicity is a significant agricultural problem globally, compromising crop growth and productivity in many areas. However, the molecular mechanisms of NH(4)(+) toxicity are still poorly understood, in part due to a lack of valuable genetic resources. Here, a novel Arabidopsis mutant, amos2 (ammonium overly sensitive 2), displaying hypersensitivity to NH(4) (+) in both shoots and roots, was isolated. The mutant exhibits the hallmarks of NH(4)(+) toxicity at significantly elevated levels: severely suppressed shoot biomass, increased leaf chlorosis, and inhibition of lateral root formation. Amos2 hypersensitivity is associated with excessive NH(4)(+) accumulation in shoots and a reduction in tissue potassium (K(+)), calcium (Ca(2+)), and magnesium (Mg(2+)). We show that the lesion is specific to the NH(4)(+) ion, is independent of NH(4)(+) metabolism, and can be partially rescued by elevated external K(+). The amos2 lesion was mapped to a 16-cM interval on top of chromosome 1, where no similar mutation has been previously mapped. Our study identifies a novel locus controlling cation homeostasis under NH(4)(+) stress and provides a tool for the future identification of critical genes involved in the development of NH(4)(+) toxicity.

  5. In Vitro Root Development in Arabidopsis Thaliana Wild-Type and scr Mutants under Clinorotation

    NASA Astrophysics Data System (ADS)

    Kordyum, E. L.; Sarnatska, V. V.; Talalaiev, A. S.; Ovcharenko, Y. V.

    2008-06-01

    A task of our experiments was to study in vitro rhizogenesis in Arabidopsis thaliana wild type and scr mutants under slow horizontal clinorotation as a convenient model to clear up a question, whether root morphogenesis de novo will occur normally in simulated microgravity. Two methods for obtaining A. thaliana roots in vitro were used: 1) from the primary callus of leaf origin and 2) directly from leaf explants. Light and electron microscopy and RT-PCR were used for an analysis of the experimental materials. Graviperceptive cells differentiated in roots formed de novo from callus and leaf explants of wild type and scr mutants but did not function under clinorotation. Tissue and cell type patterning in a root proper as well as gene expression in all variants in the control and under clinorotation were similar that gives new evidence on normal morphogenesis in altered gravity. We proposed such model for performing the experiments on board the ISS to study morphogenesis in vitro, including differentiation of graviperceptive cells.

  6. Isolation of uvh1, an Arabidopsis mutant hypersensitive to ultraviolet light and ionizing radiation.

    PubMed Central

    Harlow, G R; Jenkins, M E; Pittalwala, T S; Mount, D W

    1994-01-01

    A genetic screen for mutants of Arabidopsis that are hypersensitive to UV light was developed and used to isolate a new mutant designated uvh1. UV hypersensitivity in uvh1 was due to a single recessive trait that is probably located on chromosome 3. Although isolated as hypersensitive to an acute exposure to UV-C light, uvh1 was also hypersensitive to UV-B wavelengths, which are present in sunlight that reaches the earth's surface. UV-B damage to both wild-type and uvh1 plants could be significantly reduced by subsequent exposure of UV-irradiated plants to photoreactivating light, showing that photoreactivation of UV-B damage is important for plant viability and that uvh1 plants are not defective in photoreactivation. A new assay for DNA damage, the Dral assay, was developed and used to show that exposure of wild-type and uvh1 plants to a given dose of UV light induces the same amount of damage in chloroplast and nuclear DNA. Thus, uvh1 is not defective in a UV protective mechanism. uvh1 plants were also found to be hypersensitive to ionizing radiation. These results suggest that uvh1 is defective in a repair or tolerance mechanism that normally provides plants with resistance to several types of DNA damage. PMID:8148646

  7. Identification of new adventitious rooting mutants amongst suppressors of the Arabidopsis thaliana superroot2 mutation

    PubMed Central

    Pacurar, Daniel Ioan; Pacurar, Monica Lacramioara; Schwambach, Joseli; Bellini, Catherine

    2014-01-01

    The plant hormone auxin plays a central role in adventitious rooting and is routinely used with many economically important, vegetatively propagated plant species to promote adventitious root initiation and development on cuttings. Nevertheless the molecular mechanisms through which it acts are only starting to emerge. The Arabidopsis superroot2-1 (sur2-1) mutant overproduces auxin and, as a consequence, develops excessive adventitious roots in the hypocotyl. In order to increase the knowledge of adventitious rooting and of auxin signalling pathways and crosstalk, this study performed a screen for suppressors of superroot2-1 phenotype. These suppressors provide a new resource for discovery of genetic players involved in auxin signalling pathways or at the crosstalk of auxin and other hormones or environmental signals. This study reports the identification and characterization of 26 sur2-1 suppressor mutants, several of which were identified as mutations in candidate genes involved in either auxin biosynthesis or signalling. In addition to confirming the role of auxin as a central regulator of adventitious rooting, superroot2 suppressors indicated possible crosstalk with ethylene signalling in this process. PMID:24596172

  8. Effects of microgravity and clinorotation on stress ethylene production in two starchless mutants of Arabidopsis thaliana

    NASA Technical Reports Server (NTRS)

    Gallegos, Gregory L.; Hilaire, Emmanuel M.; Peterson, Barbara V.; Brown, Christopher S.; Guikema, James A.

    1995-01-01

    Starch filled plastids termed amyloplasts, contained within columella cells of the root caps of higher plant roots, are believed to play a statolith-like role in the gravitropic response of roots. Plants having amyloplasts containing less starch exhibit a corresponding reduction in gravitropic response. We have observed enhanced ethylene production by sweet clover (Melilotus alba L.) seedlings grown in the altered gravity condition of a slow rotating clinostat, and have suggested that this is a stress response resulting from continuous gravistimulation rather than as a result of the simulation of a microgravity condition. If so, we expect that plants deficient in starch accumulation in amyloplasts may produce less stress ethylene when grown on a clinostat. Therefore, we have grown Arabidopsis thaliana in the small, closed environment of the Fluid Processing Apparatus (FPA). In this preliminary report we compare stationary plants with clinorotated and those grown in microgravity aboard Discovery during the STS-63 flight in February 1995. In addition to wildtype, two mutants deficient in starch biosynthesis, mutants TC7 and TL25, which are, respectively, deficient in the activity of amyloplast phosphoglucomutase and ADP-glucose pyrophosphorylase, were grown for three days before being fixed within the FPA. Gas samples were aspirated from the growth chambers and carbon dioxide and ethylene concentations were measured using a gas chromatograph. The fixed tissue is currently undergoing further morphologic and microscopic characterization.

  9. Exaggerated root respiration accounts for growth retardation in a starchless mutant of Arabidopsis thaliana.

    PubMed

    Brauner, Katrin; Hörmiller, Imke; Nägele, Thomas; Heyer, Arnd G

    2014-07-01

    The knock-out mutation of plastidial phosphoglucomutase (pgm) causes a starchless phenotype in Arabidopsis thaliana, and results in a severe growth reduction of plants cultivated under diurnal conditions. It has been speculated that high soluble sugar levels accumulating during the light phase in leaf mesophyll might cause a reduction of photosynthetic activity or that shortage of reduced carbon during the night is the reason for the slow biomass gain of pgm. Separate simultaneous measurements of leaf net photosynthesis and root respiration demonstrate that photosynthetic activity per unit fresh weight is not reduced in pgm, whereas root respiration is strongly elevated. Comparison with a mutant defective in the dominating vacuolar invertase (AtβFruct4) revealed that high sucrose concentration in the cytosol, but not in the vacuole, of leaf cells is responsible for elevated assimilate transport to the root. Increased sugar supply to the root, as observed in pgm mutants, forces substantial respiratory losses. Because root respiration accounts for 80% of total plant respiration under long-day conditions, this gives rise to retarded biomass formation. In contrast, reduced vacuolar invertase activity leads to reduced net photosynthesis in the shoot and lowered root respiration, and affords an increased root/shoot ratio. The results demonstrate that roots have very limited capacity for carbon storage but exert rigid control of supply for their maintenance metabolism.

  10. Hyperspectral Imaging Techniques for Rapid Identification of Arabidopsis Mutants with Altered Leaf Pigment Status

    PubMed Central

    Matsuda, Osamu; Tanaka, Ayako; Fujita, Takao; Iba, Koh

    2012-01-01

    The spectral reflectance signature of living organisms provides information that closely reflects their physiological status. Because of its high potential for the estimation of geomorphic biological parameters, particularly of gross photosynthesis of plants, two-dimensional spectroscopy, via the use of hyperspectral instruments, has been widely used in remote sensing applications. In genetics research, in contrast, the reflectance phenotype has rarely been the subject of quantitative analysis; its potential for illuminating the pathway leading from the gene to phenotype remains largely unexplored. In this study, we employed hyperspectral imaging techniques to identify Arabidopsis mutants with altered leaf pigment status. The techniques are comprised of two modes; the first is referred to as the ‘targeted mode’ and the second as the ‘non-targeted mode’. The ‘targeted’ mode is aimed at visualizing individual concentrations and compositional parameters of leaf pigments based on reflectance indices (RIs) developed for Chls a and b, carotenoids and anthocyanins. The ‘non-targeted’ mode highlights differences in reflectance spectra of leaf samples relative to reference spectra from the wild-type leaves. Through the latter approach, three mutant lines with weak irregular reflectance phenotypes, that are hardly identifiable by simple observation, were isolated. Analysis of these and other mutants revealed that the RI-based targeted pigment estimation was robust at least against changes in trichome density, but was confounded by genetic defects in chloroplast photorelocation movement. Notwithstanding such a limitation, the techniques presented here provide rapid and high-sensitive means to identify genetic mechanisms that coordinate leaf pigment status with developmental stages and/or environmental stress conditions. PMID:22470059

  11. Metabolism of Benzyladenine is Impaired in a Mutant of Arabidopsis thaliana Lacking Adenine Phosphoribosyltransferase Activity 1

    PubMed Central

    Moffatt, Barbara; Pethe, Claude; Laloue, Michel

    1991-01-01

    Formation of the riboside-5′-monophosphate is a general feature of the metabolism of cytokinins in plants. As part of a study of the biological significance of the nucleotide form of cytokinins, we analyzed a mutant of Arabidopsis thaliana deficient in adenine phosphoribosyltransferase (APRT) activity for its ability to metabolize N6-benzyladenine (BA). Formation of N6-benzyladenosine-5′-monophosphate (BAMP) was assayed in vivo, by feeding tritiated BA to wild-type and mutant plantlets, and in crude plantlet extracts. Metabolites were separated by high performance liquid chromatography and quantitated by on-line liquid scintillation spectrometry. BA was rapidly absorbed by A. thaliana plantlets and primarily converted to BAMP and to BA 7- and 9-glucosides. BA was also rapidly absorbed by APRT-deficient plantlets, but its conversion to BAMP was strongly reduced. Formation of BAMP from N6-benzyladenosine was not affected in the mutant plantlets. In vitro conversion of BA to its nucleoside-5′-monophosphate was detected in crude extracts of wild-type plantlets, but not in extracts of APRT-deficient plantlets. Therefore, results of both assays indicate that APRT-deficient tissue does not convert BA to BAMP to a significant extent. Further, nondenaturing isoelectric focusing analysis of APRT activity in leaf extracts indicated that the enzyme activities which metabolize adenine and BA into their corresponding riboside-5′-monophosphate in extracts of wild-type plantlets have the same apparent isoelectric point. These activities were not detected in extracts prepared from APRT-deficient plantlets. Thus, these results demonstrate that APRT is the main enzyme which converts BA to its nucleotide form in young A. thaliana plants and that the ribophosphorylation of BA is not a prerequisite of its absorption by the plantlets. Images Figure 4 PMID:16668070

  12. Turnover of Fatty Acids during Natural Senescence of Arabidopsis, Brachypodium, and Switchgrass and in Arabidopsis β-Oxidation Mutants1[C][W][OA

    PubMed Central

    Yang, Zhenle; Ohlrogge, John B.

    2009-01-01

    During leaf senescence, macromolecule breakdown occurs and nutrients are translocated to support growth of new vegetative tissues, seeds, or other storage organs. In this study, we determined the fatty acid levels and profiles in Arabidopsis (Arabidopsis thaliana), Brachypodium distachyon, and switchgrass (Panicum virgatum) leaves during natural senescence. In young leaves, fatty acids represent 4% to 5% of dry weight and approximately 10% of the chemical energy content of the leaf tissues. In all three species, fatty acid levels in leaves began to decline at the onset of leaf senescence and progressively decreased as senescence advanced, resulting in a greater than 80% decline in fatty acids on a dry weight basis. During senescence, Arabidopsis leaves lost 1.6% of fatty acids per day at a rate of 2.1 μg per leaf (0.6 μg mg−1 dry weight). Triacylglycerol levels remained less than 1% of total lipids at all stages. In contrast to glycerolipids, aliphatic surface waxes of Arabidopsis leaves were much more stable, showing only minor reduction during senescence. We also examined three Arabidopsis mutants, acx1acx2, lacs6lacs7, and kat2, which are blocked in enzyme activities of β-oxidation and are defective in lipid mobilization during seed germination. In each case, no major differences in the fatty acid contents of leaves were observed between these mutants and the wild type, indicating that several mutations in β-oxidation that cause reduced breakdown of reserve oil in seeds do not substantially reduce the degradation of fatty acids during leaf senescence. PMID:19561121

  13. Complementation of the pha2 yeast mutant suggests functional differences for arogenate dehydratases from Arabidopsis thaliana.

    PubMed

    Bross, Crystal D; Corea, Oliver R A; Kaldis, Angelo; Menassa, Rima; Bernards, Mark A; Kohalmi, Susanne E

    2011-08-01

    The final steps of phenylalanine (Phe) biosynthesis in bacteria, fungi and plants can occur via phenylpyruvate or arogenate intermediates. These routes are determined by the presence of prephenate dehydratase (PDT, EC4.2.1.51), which forms phenylpyruvate from prephenate, or arogenate dehydratase (ADT, EC4.2.1.91), which forms phenylalanine directly from arogenate. We compared sequences from select yeast species to those of Arabidopsis thaliana. The in silico analysis showed that plant ADTs and yeast PDTs share many common features allowing them to act as dehydratase/decarboxylases. However, plant and yeast sequences clearly group independently conferring distinct substrate specificities. Complementation of the Saccharomyces cerevisiae pha2 mutant, which lacks PDT activity and cannot grow in the absence of exogenous Phe, was used to test the PDT activity of A. thaliana ADTs in vivo. Previous biochemical characterization showed that all six AtADTs had high catalytic activity with arogenate as a substrate, while AtADT1, AtADT2 and AtADT6 also had limited activity with prephenate. Consistent with these results, the complementation test showed AtADT2 readily recovered the pha2 phenotype after ∼6 days growth at 30 °C, while AtADT1 required ∼13 days to show visible growth. By contrast, AtADT6 (lowest PDT activity) and AtADT3-5 (no PDT activity) were unable to recover the phenotype. These results suggest that only AtADT1 and AtADT2, but not the other four ADTs from Arabidopsis, have functional PDT activity in vivo, showing that there are two functional distinct groups. We hypothesize that plant ADTs have evolved to use the arogenate route for Phe synthesis while keeping some residual PDT activity.

  14. Rapid analysis of seed size in Arabidopsis for mutant and QTL discovery

    PubMed Central

    2011-01-01

    Background Arabidopsis thaliana is a useful model organism for deciphering the genetic determinants of seed size; however the small size of its seeds makes measurements difficult. Bulk seed weights are often used as an indicator of average seed size, but details of individual seed is obscured. Analysis of seed images is possible but issues arise from variations in seed pigmentation and shadowing making analysis laborious. We therefore investigated the use of a consumer level scanner to facilitate seed size measurements in conjunction with open source image-processing software. Results By using the transmitted light from the slide scanning function of a flatbed scanner and particle analysis of the resulting images, we have developed a method for the rapid and high throughput analysis of seed size and seed size distribution. The technical variation due to the approach was negligible enabling us to identify aspects of maternal plant growth that contribute to biological variation in seed size. By controlling for these factors, differences in seed size caused by altered parental genome dosage and mutation were easily detected. The method has high reproducibility and sensitivity, such that a mutant with a 10% reduction in seed size was identified in a screen of endosperm-expressed genes. Our study also generated average seed size data for 91 Arabidopsis accessions and identified a number of quantitative trait loci from two recombinant inbred line populations, generated from Cape Verde Islands and Burren accessions crossed with Columbia. Conclusions This study describes a sensitive, high-throughput approach for measuring seed size and seed size distribution. The method provides a low cost and robust solution that can be easily implemented into the workflow of studies relating to various aspects of seed development. PMID:21303553

  15. Effects of gravity on growth phenotype in MAPs mutants of Arabidopsis

    NASA Astrophysics Data System (ADS)

    Higuchi, Sayoko; Kumasaki, Saori; Matsumoto, Shouhei; Soga, Kouichi; Wakabayashi, Kazuyuki; Hashimoto, Takashi; Hoson, Takayuki

    Hypergravity suppresses elongation growth and promotes lateral expansion of stem organs in various plants. It has been shown that cortical microtubules are involved in gravity-induced modifications of growth and development. Because microtubule-associated proteins (MAPs) are important in dynamics of microtubules, they may also play a role in the gravity response. In the present study, the roles of MAPs (MOR1, SPR1, SPR2, MAP65, and KTN1) in hypergravityinduced changes in growth and development were examined in Arabidopsis hypocotyls. The expression of MOR1, SPR1, SPR2 , and MAP65 genes was down-regulated, whereas that of KTN1 gene was increased transiently by hypergravity. We analyzed the growth behavior of MAPs mutants (mor1/rid5, spr1-2 , spr2-2, and katanin mutants) under hypergravity conditions. Hypergravity inhibited elongation growth of hypocotyls in spr1-2 as in wild-type. On the other hand, elongation growth of hypocotyls in mor1/rid5, spr2-2, and katanin mutants was suppressed as compared with wild-type under 1 g conditions, and was not affected further by hypergravity stimuli. Hypocotyls of mor1/rid5, spr1-2 , and spr2-2 also showed helical growth even under 1 g conditions, and in mor1/rid5 such a phenotype was intensified under hypergravity conditions. The alignment of cell line was abnormal in hypocotyls of katanin mutants under both 1 g and hypergravity conditions. The orientation of cortical microtubules in wildtype hypocotyls was changed from transverse direction to longitudinal or random directions by hypergravity stimuli. In mor1/rid5 hypocotyls, the orientation of microtubules was random even under 1 g condition, which was not affected by hypergravity. Furthermore, partial disruption of cortical microtubules was observed in mor1/rid5 hypocotyls. These results suggest that MAPs, especially MOR1, play an important role in maintenance of normal growth phenotype against gravity in plants probably via stabilization of microtubule structure.

  16. Characterization of an activation-tagged mutant uncovers a role of GLABRA2 in anthocyanin biosynthesis in Arabidopsis

    DOE PAGES

    Wang, Xiaoyu; Wang, Xianling; Hu, Qingnan; ...

    2015-06-17

    In Arabidopsis, anthocyanin biosynthesis is controlled by a MYB-bHLH-WD40 (MBW) transcriptional activator complex. The MBW complex activates the transcription of late biosynthesis genes in the flavonoid pathway, leading to the production of anthocyanins. A similar MBW complex regulates epidermal cell fate by activating the transcription of GLABRA2 (GL2), a homeodomain transcription factor required for trichome formation in shoots and non-hair cell formation in roots. Here we provide experimental evidence to show that GL2 also plays a role in regulating anthocyanin biosynthesis in Arabidopsis. From an activation-tagged mutagenized population of Arabidopsis plants, we isolated a dominant, gain-of-function mutant with reduced anthocyanins.more » Molecular cloning revealed that this phenotype is caused by an elevated expression of GL2, thus the mutant was named gl2-1D. Consistent with the view that GL2 acts as a negative regulator of anthocyanin biosynthesis, gl2-1D seedlings accumulated less whereas gl2-3 seedlings accumulated more anthocyanins in response to sucrose. Gene expression analysis indicated that expression of late, but not early, biosynthesis genes in the flavonoid pathway was dramatically reduced in gl2-1D but elevated in gl2-3 mutants. Further analysis showed that expression of some MBW component genes involved in the regulation of late biosynthesis genes was reduced in gl2-1D but elevated in gl2-3 mutants, and chromatin immunoprecipitation results indicated that some MBW component genes are targets of GL2. We also showed that GL2 functions as a transcriptional repressor. Altogether, these results indicate that GL2 negatively regulates anthocyanin biosynthesis in Arabidopsis by directly repressing the expression of some MBW component genes.« less

  17. Characterization of an activation-tagged mutant uncovers a role of GLABRA2 in anthocyanin biosynthesis in Arabidopsis

    SciTech Connect

    Wang, Xiaoyu; Wang, Xianling; Hu, Qingnan; Dai, Xuemei; Tian, Hainan; Zheng, Kaijie; Wang, Xiaoping; Mao, Tonglin; Chen, Jin -Gui; Wang, Shucai

    2015-06-17

    In Arabidopsis, anthocyanin biosynthesis is controlled by a MYB-bHLH-WD40 (MBW) transcriptional activator complex. The MBW complex activates the transcription of late biosynthesis genes in the flavonoid pathway, leading to the production of anthocyanins. A similar MBW complex regulates epidermal cell fate by activating the transcription of GLABRA2 (GL2), a homeodomain transcription factor required for trichome formation in shoots and non-hair cell formation in roots. Here we provide experimental evidence to show that GL2 also plays a role in regulating anthocyanin biosynthesis in Arabidopsis. From an activation-tagged mutagenized population of Arabidopsis plants, we isolated a dominant, gain-of-function mutant with reduced anthocyanins. Molecular cloning revealed that this phenotype is caused by an elevated expression of GL2, thus the mutant was named gl2-1D. Consistent with the view that GL2 acts as a negative regulator of anthocyanin biosynthesis, gl2-1D seedlings accumulated less whereas gl2-3 seedlings accumulated more anthocyanins in response to sucrose. Gene expression analysis indicated that expression of late, but not early, biosynthesis genes in the flavonoid pathway was dramatically reduced in gl2-1D but elevated in gl2-3 mutants. Further analysis showed that expression of some MBW component genes involved in the regulation of late biosynthesis genes was reduced in gl2-1D but elevated in gl2-3 mutants, and chromatin immunoprecipitation results indicated that some MBW component genes are targets of GL2. We also showed that GL2 functions as a transcriptional repressor. Altogether, these results indicate that GL2 negatively regulates anthocyanin biosynthesis in Arabidopsis by directly repressing the expression of some MBW component genes.

  18. Tryptophan-Derived Metabolites Are Required for Antifungal Defense in the Arabidopsis mlo2 Mutant1[C][W][OA

    PubMed Central

    Consonni, Chiara; Bednarek, Paweł; Humphry, Matt; Francocci, Fedra; Ferrari, Simone; Harzen, Anne; Ver Loren van Themaat, Emiel; Panstruga, Ralph

    2010-01-01

    Arabidopsis (Arabidopsis thaliana) genes MILDEW RESISTANCE LOCUS O2 (MLO2), MLO6, and MLO12 exhibit unequal genetic redundancy with respect to the modulation of defense responses against powdery mildew fungi and the control of developmental phenotypes such as premature leaf decay. We show that early chlorosis and necrosis of rosette leaves in mlo2 mlo6 mlo12 mutants reflects an authentic but untimely leaf senescence program. Comparative transcriptional profiling revealed that transcripts of several genes encoding tryptophan biosynthetic and metabolic enzymes hyperaccumulate during vegetative development in the mlo2 mlo6 mlo12 mutant. Elevated expression levels of these genes correlate with altered steady-state levels of several indolic metabolites, including the phytoalexin camalexin and indolic glucosinolates, during development in the mlo2 single mutant and the mlo2 mlo6 mlo12 triple mutant. Results of genetic epistasis analysis suggest a decisive role for indolic metabolites in mlo2-conditioned antifungal defense against both biotrophic powdery mildews and a camalexin-sensitive strain of the necrotrophic fungus Botrytis cinerea. The wound- and pathogen-responsive callose synthase POWDERY MILDEW RESISTANCE4/GLUCAN SYNTHASE-LIKE5 was found to be responsible for the spontaneous callose deposits in mlo2 mutant plants but dispensable for mlo2-conditioned penetration resistance. Our data strengthen the notion that powdery mildew resistance of mlo2 genotypes is based on the same defense execution machinery as innate antifungal immune responses that restrict the invasion of nonadapted fungal pathogens. PMID:20023151

  19. An altered hydrotropic response (ahr1) mutant of Arabidopsis recovers root hydrotropism with cytokinin

    PubMed Central

    Saucedo, Manuel; Ponce, Georgina; Campos, María Eugenia; Eapen, Delfeena; García, Edith; Luján, Rosario; Sánchez, Yoloxóchitl; Cassab, Gladys I.

    2012-01-01

    Roots are highly plastic and can acclimate to heterogeneous and stressful conditions. However, there is little knowledge of the effect of moisture gradients on the mechanisms controlling root growth orientation and branching, and how this mechanism may help plants to avoid drought responses. The aim of this study was to isolate mutants of Arabidopsis thaliana with altered hydrotropic responses. Here, altered hydrotropic response 1 (ahr1), a semi-dominant allele segregating as a single gene mutation, was characterized. ahr1 directed the growth of its primary root towards the source of higher water availability and developed an extensive root system over time. This phenotype was intensified in the presence of abscisic acid and was not observed if ahr1 seedlings were grown in a water stress medium without a water potential gradient. In normal growth conditions, primary root growth and root branching of ahr1 were indistinguishable from those of the wild type (wt). The altered hydrotropic growth of ahr1 roots was confirmed when the water-rich source was placed at an angle of 45° from the gravity vector. In this system, roots of ahr1 seedlings grew downward and did not display hydrotropism; however, in the presence of cytokinins, they exhibited hydrotropism like those of the wt, indicating that cytokinins play a critical role in root hydrotropism. The ahr1 mutant represents a valuable genetic resource for the study of the effects of cytokinins in the differential growth of hydrotropism and control of lateral root formation during the hydrotropic response. PMID:22442413

  20. Altered gravitropic response, amyloplast sedimentation and circumnutation in the Arabidopsis shoot gravitropism 5 mutant are associated with reduced starch levels.

    PubMed

    Tanimoto, Mimi; Tremblay, Reynald; Colasanti, Joseph

    2008-05-01

    Plants have developed sophisticated gravity sensing mechanisms to interpret environmental signals that are vital for optimum plant growth. Loss of SHOOT GRAVITROPISM 5 (SGR5) gene function has been shown to affect the gravitropic response of Arabidopsis inflorescence stems. SGR5 is a member of the INDETERMINATE DOMAIN (IDD) zinc finger protein family of putative transcription factors. As part of an ongoing functional analysis of Arabidopsis IDD genes (AtIDD) we have extended the characterisation of SGR5, and show that gravity sensing amyloplasts in the shoot endodermis of sgr5 mutants sediment more slowly than wild type, suggesting a defect in gravity perception. This is correlated with lower amyloplast starch levels, which may account for the reduced gravitropic sensitivity in sgr5. Further, we find that sgr5 mutants have a severely attenuated stem circumnutation movement typified by a reduced amplitude and an decreased periodicity. adg1-1 and sex1-1 mutants, which contain no starch or increased starch, respectively, also show alterations in the amplitude and period of circumnutation. Together these results suggest that plant growth movement may depend on starch levels and/or gravity sensing. Overall, we propose that loss of SGR5 regulatory activity affects starch accumulation in Arabidopsis shoot tissues and causes decreased sensitivity to gravity and diminished circumnutational movements.

  1. The Arabidopsis thiamin deficient mutant pale green1 lacks thiamin monophosphate phosphatase of the vitamin B1 biosynthesis pathway.

    PubMed

    Hsieh, Wei-Yu; Liao, Jo-Chien; Wang, Hsin-Tzu; Hung, Tzu-Huan; Tseng, Ching-Chih; Chung, Tsui-Yun; Hsieh, Ming-Hsiun

    2017-03-27

    Thiamin diphosphate (TPP, vitamin B1) is an essential coenzyme present in all organisms. Animals obtain TPP from their diets, but plants synthesize TPP de novo. We isolated and characterized an Arabidopsis pale green1 (pale1) mutant that contained higher concentrations of thiamin monophosphate (TMP) and less thiamin and TPP than the wild type. Supplementation with thiamin, but not the thiazole and pyrimidine precursors, rescued the mutant phenotype, indicating that the pale1 mutant is a thiamin-deficient mutant. Map-based cloning and whole-genome sequencing revealed that the pale1 mutant has a mutation in At5g32470 encoding a TMP phosphatase of the TPP biosynthesis pathway. We further confirmed that the mutation of At5g32470 is responsible for the mutant phenotypes by complementing the pale1 mutant with constructs overexpressing full-length At5g32470. Most plant TPP biosynthetic enzymes are located in the chloroplasts and cytosol, but At5g32470-GFP localized to the mitochondrion of the root, hypocotyl, mesophyll, and guard cells of the 35S:At5g32470-GFP complemented plants. The subcellular localization of a functional TMP phosphatase suggests that the complete vitamin B1 biosynthesis pathway may involve the chloroplasts, mitochondria and cytosol in plants. Analysis of PALE1 promoter-GUS activity revealed that PALE1 is mainly expressed in vascular tissues of Arabidopsis seedlings. Quantitative RT-PCR analysis of TPP biosynthesis genes and genes encoding the TPP-dependent enzymes pyruvate dehydrogenase, α-ketoglutarate dehydrogenase and transketolase revealed that the transcript levels of these genes were up-regulated in the pale1 mutant. These results suggest that endogenous levels of TPP may affect the expression of genes involved in TPP biosynthesis and TPP-dependent enzymes. This article is protected by copyright. All rights reserved.

  2. Gravitropism and development of wild-type and starch-deficient mutants of Arabidopsis during spaceflight.

    PubMed

    Kiss, J Z; Katembe, W J; Edelmann, R E

    1998-04-01

    The "starch-statolith" hypothesis has been used by plant physiologists to explain the gravity perception mechanism in higher plants. In order to help resolve some of the controversy associated with ground-based research that has supported this theory, we performed a spaceflight experiment during the January 1997 mission of the Space Shuttle STS-81. Seedlings of wild-type (WT) Arabidopsis, two reduced-starch strains, and a starchless mutant were grown in microgravity and then given a gravity stimulus on a centrifuge. In terms of development in space, germination was greater than 90% for seeds in microgravity, and flight seedlings were smaller (60% in total length) compared to control plants grown on the ground and to control plants on a rotating clinostat. Seedlings grown in space had two structural features that distinguished them from the controls: a greater density of root hairs and an anomalous hypocotyl hook structure. However, the slower growth and morphological changes observed in the flight seedlings may be due to the effects of ethylene present in the spacecraft. Nevertheless, during the flight hypocotyls of WT seedlings responded to a unilateral 60 min stimulus provided by a 1-g centrifuge while those of the starch-deficient strains did not. Thus the strain with the greatest amount of starch responded to the stimulus given in flight and therefore, these data support the starch-statolith model for gravity sensing.

  3. The Arabidopsis mutant alh1 illustrates a cross talk between ethylene and auxin.

    PubMed

    Vandenbussche, Filip; Smalle, Jan; Le, Jie; Saibo, Nelson José Madeira; De Paepe, Annelies; Chaerle, Laury; Tietz, Olaf; Smets, Raphael; Laarhoven, Lucas J J; Harren, Frans J M; Van Onckelen, Harry; Palme, Klaus; Verbelen, Jean-Pierre; Van Der Straeten, Dominique

    2003-03-01

    Ethylene or its precursor 1-aminocyclopropane-1-carboxylic acid (ACC) can stimulate hypocotyl elongation in light-grown Arabidopsis seedlings. A mutant, designated ACC-related long hypocotyl 1 (alh1), that displayed a long hypocotyl in the light in the absence of the hormone was characterized. Etiolated alh1 seedlings overproduced ethylene and had an exaggerated apical hook and a thicker hypocotyl, although no difference in hypocotyl length was observed when compared with wild type. Alh1 plants were less sensitive to ethylene, as reflected by reduction of ACC-mediated inhibition of hypocotyl growth in the dark and delay in flowering and leaf senescence. Alh1 also had an altered response to auxin, whereas auxin levels in whole alh1 seedlings remained unaffected. In contrast to wild type, alh1 seedlings showed a limited hypocotyl elongation when treated with indole-3-acetic acid. Alh1 roots had a faster response to gravity. Furthermore, the hypocotyl elongation of alh1 and of ACC-treated wild type was reverted by auxin transport inhibitors. In addition, auxin up-regulated genes were ectopically expressed in hypocotyls upon ACC treatment, suggesting that the ethylene response is mediated by auxins. Together, these data indicate that alh1 is altered in the cross talk between ethylene and auxins, probably at the level of auxin transport.

  4. Gravitropism and development of wild-type and starch-deficient mutants of Arabidopsis during spaceflight

    NASA Technical Reports Server (NTRS)

    Kiss, J. Z.; Katembe, W. J.; Edelmann, R. E.

    1998-01-01

    The "starch-statolith" hypothesis has been used by plant physiologists to explain the gravity perception mechanism in higher plants. In order to help resolve some of the controversy associated with ground-based research that has supported this theory, we performed a spaceflight experiment during the January 1997 mission of the Space Shuttle STS-81. Seedlings of wild-type (WT) Arabidopsis, two reduced-starch strains, and a starchless mutant were grown in microgravity and then given a gravity stimulus on a centrifuge. In terms of development in space, germination was greater than 90% for seeds in microgravity, and flight seedlings were smaller (60% in total length) compared to control plants grown on the ground and to control plants on a rotating clinostat. Seedlings grown in space had two structural features that distinguished them from the controls: a greater density of root hairs and an anomalous hypocotyl hook structure. However, the slower growth and morphological changes observed in the flight seedlings may be due to the effects of ethylene present in the spacecraft. Nevertheless, during the flight hypocotyls of WT seedlings responded to a unilateral 60 min stimulus provided by a 1-g centrifuge while those of the starch-deficient strains did not. Thus the strain with the greatest amount of starch responded to the stimulus given in flight and therefore, these data support the starch-statolith model for gravity sensing.

  5. Arabidopsis thaliana sku mutant seedlings show exaggerated surface-dependent alteration in root growth vector

    NASA Technical Reports Server (NTRS)

    Rutherford, R.; Masson, P. H.

    1996-01-01

    Roots of wild-type Arabidopsis thaliana seedlings in the Wassilewskija (WS) and Landsberg erecta (Ler) ecotypes often grow aslant on vertical agar surfaces. Slanted root growth always occurs to the right of the gravity vector when the root is viewed through the agar surface, and is not observed in the Columbia ecotype. Right-slanted root growth is surface-dependent and does not result directly from directional environmental stimuli or gradients in the plane of skewing. We have isolated two partially dominant mutations in WS (sku1 and sku2) that show an exaggerated right-slanting root-growth phenotype on agar surfaces. The right-slanting root-growth phenotype of wild-type and mutant roots is not the result of diagravitropism or of an alteration in root gravitropism. It is accompanied by a left-handed rotation of the root about its axis within the elongation zone, the rate of which positively correlates with the degree of right-slanted curvature. Our data suggest that the right-slanting root growth phenotype results from an endogenous structural asymmetry that expresses itself by a directional root-tip rotation.

  6. Rootcap structure in wild type and in a starchless mutant of Arabidopsis

    NASA Technical Reports Server (NTRS)

    Sack, F. D.; Kiss, J. Z.

    1989-01-01

    Rootcaps of the wild type (WT) and of a starchless, gravitropic mutant (TC7) of Arabidopsis thaliana L. were examined by electron microscopy to identify cellular polarities with respect to gravity. In columella cells, nuclei are located proximally, and the nuclear envelope is continuous with endoplasmic reticulum (ER) that is in turn connected to nearby plasmodesmata. Impregnation of ER with osmium ferricyanide revealed numerous contacts between columella plastids and ER in both genotypes. ER is present mostly in the outer regions of the columella protoplast except in older columella cells that are developing into peripheral cells. In vertical roots, only columella cells that are intermediate in development (story 2 cells) have a higher surface density (S) of ER in the distal compared to proximal regions of the cell. The distal but not the proximal S of the ER is constant throughout columella development. Plastids are less sedimented in TC7 columella cells compared to those of the WT. It is hypothesized that plastid contact with the ER plays a role in gravity perception in both genotypes.

  7. Metabolic profiling of the Arabidopsis pkl mutant reveals selective derepression of embryonic traits.

    PubMed

    Rider, Stanley Dean; Hemm, Matthew R; Hostetler, Heather A; Li, Hui-Chun; Chapple, Clint; Ogas, Joe

    2004-07-01

    Embryos express several unique differentiation characteristics, including the accumulation of a number of metabolites that are generally considered to be unique to seeds. PICKLE (PKL) codes for a CHD3-chromatin remodeling factor that is necessary for repression of embryonic traits in seedlings of Arabidopsis thaliana (L.) Heynh. In pkl mutants, primary roots are capable of expressing many embryonic traits after germination and are referred to as "pickle roots". In an attempt to examine the breadth of PKL-dependent repression of embryo-specific differentiation pathways, we determined the extent to which a variety of embryo-specific compounds accumulate in pickle roots. We found that pickle roots accumulate triacylglycerol with a fatty acid composition that is similar to that found in seeds. The major seed storage proteins are also present in pickle roots. In addition to these two well-characterized seed storage compounds, we observed that pickle roots accumulate phytate, a form of stored phosphate that is preferentially accumulated in seeds. Seeds of members of the Brassicaceae also accumulate a variety of unique secondary metabolites, including sinapate esters and glucosinolates. Surprisingly, the levels of secondary metabolites in pickle roots were not suggestive of an embryonic differentiation state, but did reveal that a mutation in PKL results in substantial changes in root secondary metabolism. Taken together, these data suggest that PKL is responsible for regulating some but not all aspects of the embryonic program as it relates to the accumulation of embryo-specific metabolites.

  8. Wax and cutin mutants of Arabidopsis: Quantitative characterization of the cuticular transport barrier in relation to chemical composition.

    PubMed

    Sadler, Christina; Schroll, Bettina; Zeisler, Viktoria; Waßmann, Friedrich; Franke, Rochus; Schreiber, Lukas

    2016-09-01

    Using (14)C-labeled epoxiconazole as a tracer, cuticular permeability of Arabidopsis thaliana leaves was quantitatively measured in order to compare different wax and cutin mutants (wax2, cut1, cer5, att1, bdg, shn3 and shn1) to the corresponding wild types (Col-0 and Ws). Mutants were characterized by decreases or increases in wax and/or cutin amounts. Permeances [ms(-1)] of Arabidopsis cuticles either increased in the mutants compared to wild type or were not affected. Thus, genetic changes in wax and cutin biosynthesis in some of the investigated Arabidopsis mutants obviously impaired the coordinated cutin and wax deposition at the outer leaf epidermal cell wall. As a consequence, barrier properties of cuticles were significantly decreased. However, increasing cutin and wax amounts by genetic modifications, did not automatically lead to improved cuticular barrier properties. As an alternative approach to the radioactive transport assay, changes in chlorophyll fluorescence were monitored after foliar application of metribuzine, an herbicide inhibiting electron transport in chloroplasts. Since both, half-times of photosynthesis inhibition as well as times of complete inhibition, in fact correlated with (14)C-epoxiconazole permeances, different rates of decline of photosynthetic yield between mutants and wild type must be a function of foliar uptake of the herbicide across the cuticle. Thus, monitoring changes in chlorophyll fluorescence, instead of conducting radioactive transport assays, represents an easy-to-handle and fast alternative evaluating cuticular barrier properties of different genotypes. This article is part of a Special Issue entitled: Plant Lipid Biology edited by Kent D. Chapman and Ivo Feussner.

  9. Gravitropism in a starchless mutant of Arabidopsis: implications for the starch-statolith theory of gravity sensing

    NASA Technical Reports Server (NTRS)

    Caspar, T.; Pickard, B. G.

    1989-01-01

    The starch-statolith theory of gravity reception has been tested with a mutant of Arabidopsis thaliana (L.) Heynh. which, lacking plastid phosphoglucomutase (EC 2.7.5.1) activity, does not synthesize starch. The hypocotyls and seedling roots of the mutant were examined by light and electron microscopy to confirm that they did not contain starch. In upright wild-type (WT) seedlings, starch-filled plastids in the starch sheath of the hypocotyl and in three of the five columellar layers of the root cap were piled on the cell floors, and sedimented to the ceilings when the plants were inverted. However, starchless plastids of the mutant were not significantly sedimented in these cells in either upright or inverted seedlings. Gravitropism of light-grown seedling roots was vigorous: e.g., 10 degrees curvature developed in mutants rotated on a clinostat following a 5 min induction at 1 g, compared with 14 degrees in the WT. Curvatures induced during intervals from 2.5 to 30 min were 70% as great in the mutant as the WT. Thus under these conditions the presence of starch and the sedimentation of plastids are unnecessary for reception of gravity by Arabidopsis roots. Gravitropism by hypocotyls of light-grown seedlings was less vigorous than that by roots, but the mutant hypocotyls exhibited an average of 70-80% as much curvature as the WT. Roots and hypocotyls of etiolated seedlings and flower stalks of mature plants were also gravitropic, although in these cases the mutant was generally less closely comparable to the WT. Thus, starch is also unnecessary for gravity reception in these tissues.

  10. Transcriptome analysis of Arabidopsis mutants suggests a crosstalk between ABA, ethylene and GSH against combined cold and osmotic stress

    PubMed Central

    Kumar, Deepak; Hazra, Saptarshi; Datta, Riddhi; Chattopadhyay, Sharmila

    2016-01-01

    The involvement of ethylene and abscisic acid in providing stress tolerance and defence response to plants is widely recognized. However, little is known about the cross-talk between glutathione with ethylene and abscisic acid to combat stress in planta. Here, transcriptome analysis of combined cold and osmotic stress treated Arabidopsis mutants were carried out to elucidate the crosstalk between the abscisic acid, ethylene and glutathione. Microarray experiment revealed the differential regulation of about 2313 and 4131 transcripts in ein2 (ethylene insensitive mutant) and aba1.6 (abscisic acid mutant) respectively. Functional analysis exposed common down-regulated stress and defence, secondary metabolite biosynthesis viz. phenylpropanoid, lignin and flavonols, redox and transcription factors related genes in ein2, aba1.6 and pad2.1 (glutathione mutant) in response to combined stress treatment. The reduced glutathione content was less in stress treated mutants in comparison to Col-0. Again, selective down-regulated transcripts in stress treated mutants were noted up-regulated after glutathione feeding. Some of the important differentially expressed genes were also validated by comparative proteomics analysis of stress treated mutants. In summary, our results suggested the role of ethylene and abscisic acid in inducing stress-responsive genes and proteins by activating glutathione biosynthesis to combat abiotic stress conditions in plant system. PMID:27845361

  11. Chloroplast Dysfunction Causes Multiple Defects in Cell Cycle Progression in the Arabidopsis crumpled leaf Mutant1[C][W

    PubMed Central

    Hudik, Elodie; Yoshioka, Yasushi; Domenichini, Séverine; Bourge, Mickaël; Soubigout-Taconnat, Ludivine; Mazubert, Christelle; Yi, Dalong; Bujaldon, Sandrine; Hayashi, Hiroyuki; De Veylder, Lieven; Bergounioux, Catherine; Benhamed, Moussa; Raynaud, Cécile

    2014-01-01

    The majority of research on cell cycle regulation is focused on the nuclear events that govern the replication and segregation of the genome between the two daughter cells. However, eukaryotic cells contain several compartmentalized organelles with specialized functions, and coordination among these organelles is required for proper cell cycle progression, as evidenced by the isolation of several mutants in which both organelle function and overall plant development were affected. To investigate how chloroplast dysfunction affects the cell cycle, we analyzed the crumpled leaf (crl) mutant of Arabidopsis (Arabidopsis thaliana), which is deficient for a chloroplastic protein and displays particularly severe developmental defects. In the crl mutant, we reveal that cell cycle regulation is altered drastically and that meristematic cells prematurely enter differentiation, leading to reduced plant stature and early endoreduplication in the leaves. This response is due to the repression of several key cell cycle regulators as well as constitutive activation of stress-response genes, among them the cell cycle inhibitor SIAMESE-RELATED5. One unique feature of the crl mutant is that it produces aplastidic cells in several organs, including the root tip. By investigating the consequence of the absence of plastids on cell cycle progression, we showed that nuclear DNA replication occurs in aplastidic cells in the root tip, which opens future research prospects regarding the dialogue between plastids and the nucleus during cell cycle regulation in higher plants. PMID:25037213

  12. The xipotl Mutant of Arabidopsis Reveals a Critical Role for Phospholipid Metabolism in Root System Development and Epidermal Cell Integrity

    PubMed Central

    Cruz-Ramírez, Alfredo; López-Bucio, José; Ramírez-Pimentel, Gabriel; Zurita-Silva, Andrés; Sánchez-Calderon, Lenin; Ramírez-Chávez, Enrique; González-Ortega, Emmanuel; Herrera-Estrella, Luis

    2004-01-01

    Phosphocholine (PCho) is an essential metabolite for plant development because it is the precursor for the biosynthesis of phosphatidylcholine, which is the major lipid component in plant cell membranes. The main step in PCho biosynthesis in Arabidopsis thaliana is the triple, sequential N-methylation of phosphoethanolamine, catalyzed by S-adenosyl-l-methionine:phosphoethanolamine N-methyltransferase (PEAMT). In screenings performed to isolate Arabidopsis mutants with altered root system architecture, a T-DNA mutagenized line showing remarkable alterations in root development was isolated. At the seedling stage, the mutant phenotype is characterized by a short primary root, a high number of lateral roots, and short epidermal cells with aberrant morphology. Genetic and biochemical characterization of this mutant showed that the T-DNA was inserted at the At3g18000 locus (XIPOTL1), which encodes PEAMT (XIPOTL1). Further analyses revealed that inhibition of PCho biosynthesis in xpl1 mutants not only alters several root developmental traits but also induces cell death in root epidermal cells. Epidermal cell death could be reversed by phosphatidic acid treatment. Taken together, our results suggest that molecules produced downstream of the PCho biosynthesis pathway play key roles in root development and act as signals for cell integrity. PMID:15295103

  13. Integrated analysis of transcriptome and metabolome of Arabidopsis albino or pale green mutants with disrupted nuclear-encoded chloroplast proteins.

    PubMed

    Satou, Masakazu; Enoki, Harumi; Oikawa, Akira; Ohta, Daisaku; Saito, Kazunori; Hachiya, Takushi; Sakakibara, Hitoshi; Kusano, Miyako; Fukushima, Atsushi; Saito, Kazuki; Kobayashi, Masatomo; Nagata, Noriko; Myouga, Fumiyoshi; Shinozaki, Kazuo; Motohashi, Reiko

    2014-07-01

    We used four mutants having albino or pale green phenotypes with disrupted nuclear-encoded chloroplast proteins to analyze the regulatory system of metabolites in chloroplast. We performed an integrated analyses of transcriptomes and metabolomes of the four mutants. Transcriptome analysis was carried out using the Agilent Arabidopsis 2 Oligo Microarray, and metabolome analysis with two mass spectrometers; a direct-infusion Fourier transform ion cyclotron resonance mass spectrometer (FT-ICR/MS) and a gas chromatograph-time of flight mass spectrometer. Among approximately 200 known metabolites detected by the FT-ICR/MS, 71 metabolites showed significant changes in the mutants when compared with controls (Ds donor plants). Significant accumulation of several amino acids (glutamine, glutamate and asparagine) was observed in the albino and pale green mutants. Transcriptome analysis revealed altered expressions of genes in several metabolic pathways. For example, genes involved in the tricarboxylic acid cycle, the oxidative pentose phosphate pathway, and the de novo purine nucleotide biosynthetic pathway were up-regulated. These results suggest that nitrogen assimilation is constitutively promoted in the albino and pale green mutants. The accumulation of ammonium ions in the albino and pale green mutants was consistently higher than in Ds donor lines. Furthermore, genes related to pyridoxin accumulation and the de novo purine nucleotide biosynthetic pathway were up-regulated, which may have occurred as a result of the accumulation of glutamine in the albino and pale green mutants. The difference in metabolic profiles seems to be correlated with the disruption of chloroplast internal membrane structures in the mutants. In albino mutants, the alteration of metabolites accumulation and genes expression is stronger than pale green mutants.

  14. A Cytosolic Acyltransferase Contributes to Triacylglycerol Synthesis in Sucrose-Rescued Arabidopsis Seed Oil Catabolism Mutants1[W][OA

    PubMed Central

    Hernández, M. Luisa; Whitehead, Lynne; He, Zhesi; Gazda, Valeria; Gilday, Alison; Kozhevnikova, Ekaterina; Vaistij, Fabián E.; Larson, Tony R.; Graham, Ian A.

    2012-01-01

    Triacylglycerol (TAG) levels and oil bodies persist in sucrose (Suc)-rescued Arabidopsis (Arabidopsis thaliana) seedlings disrupted in seed oil catabolism. This study set out to establish if TAG levels persist as a metabolically inert pool when downstream catabolism is disrupted, or if other mechanisms, such as fatty acid (FA) recycling into TAG are operating. We show that TAG composition changes significantly in Suc-rescued seedlings compared with that found in dry seeds, with 18:2 and 18:3 accumulating. However, 20:1 FA is not efficiently recycled back into TAG in young seedlings, instead partitioning into the membrane lipid fraction and diacylglycerol. In the lipolysis mutant sugar dependent1and the β-oxidation double mutant acx1acx2 (for acyl-Coenzyme A oxidase), levels of TAG actually increased in seedlings growing on Suc. We performed a transcriptomic study and identified up-regulation of an acyltransferase gene, DIACYLGLYCEROL ACYLTRANSFERASE3 (DGAT3), with homology to a peanut (Arachis hypogaea) cytosolic acyltransferase. The acyl-Coenzyme A substrate for this acyltransferase accumulates in mutants that are blocked in oil breakdown postlipolysis. Transient expression in Nicotiana benthamiana confirmed involvement in TAG synthesis and specificity toward 18:3 and 18:2 FAs. Double-mutant analysis with the peroxisomal ATP-binding cassette transporter mutant peroxisomal ABC transporter1 indicated involvement of DGAT3 in the partitioning of 18:3 into TAG in mutant seedlings growing on Suc. Fusion of the DGAT3 protein with green fluorescent protein confirmed localization to the cytosol of N. benthamiana. This work has demonstrated active recycling of 18:2 and 18:3 FAs into TAG when seed oil breakdown is blocked in a process involving a soluble cytosolic acyltransferase. PMID:22760209

  15. Reconstitution of a Secondary Cell Wall in a Secondary Cell Wall-Deficient Arabidopsis Mutant

    PubMed Central

    Sakamoto, Shingo; Mitsuda, Nobutaka

    2015-01-01

    The secondary cell wall constitutes a rigid frame of cells in plant tissues where rigidity is required. Deposition of the secondary cell wall in fiber cells contributes to the production of wood in woody plants. The secondary cell wall is assembled through co-operative activities of many enzymes, and their gene expression is precisely regulated by a pyramidal cascade of transcription factors. Deposition of a transmuted secondary cell wall in empty fiber cells by expressing selected gene(s) in this cascade has not been attempted previously. In this proof-of-concept study, we expressed chimeric activators of 24 transcription factors that are preferentially expressed in the stem, in empty fiber cells of the Arabidopsis nst1-1 nst3-1 double mutant, which lacks a secondary cell wall in fiber cells, under the control of the NST3 promoter. The chimeric activators of MYB46, SND2 and ANAC075, as well as NST3, reconstituted a secondary cell wall with different characteristics from those of the wild type in terms of its composition. The transgenic lines expressing the SND2 or ANAC075 chimeric activator showed increased glucose and xylose, and lower lignin content, whereas the transgenic line expressing the MYB46 chimeric activator showed increased mannose content. The expression profile of downstream genes in each transgenic line was also different from that of the wild type. This study proposed a new screening strategy to identify factors of secondary wall formation and also suggested the potential of the artificially reconstituted secondary cell walls as a novel raw material for production of bioethanol and other chemicals. PMID:25535195

  16. Alterations in growth, photosynthesis, and respiration in a starchless mutant of Arabidopsis thaliana (L. ) deficient in chloroplast phosphoglucomutase activity

    SciTech Connect

    Caspar, T.; Huber, S.C.; Somerville, C.

    1985-09-01

    A mutant of Arabidopsis thaliana (L.) Heynh. which lacks leaf starch was isolated by screening for plants which did not stain with iodine. When grown in a 12-h photoperiod, leaves of the wild-type accumulated substantial amounts of starch but lower levels of soluble sugars. Under these conditions, the mutant accumulated relatively high levels of soluble sugars. Rates of growth and net photosynthesis of the mutant and wild-type were indistinguishable when the plants were grown in constant illumination. However, in a short photoperiod, the growth of the mutant was severely impaired, the rate of photosynthesis was depressed relative to the wild-type, and the rate of dark respiration, which was high following the onset of darkness, exhibited an uncharacteristic decay throughout the dark period. The depressed photosynthetic capacity of the mutant may also reflect a metabolic adaptation to the accumulation of high levels of soluble carbohydrate which mimics the effects of alterations in source/sink ratio. The activities of sucrose phosphate synthase and acid invertase are significantly higher in the mutant than in the wild-type whereas ADP-glucose pyrophosphorylase activity is lower. This suggests that the activities of these enzymes may be modulated in response to metabolite concentrations or flux through the pathways.

  17. Genetic Analysis of Arabidopsis Mutants Impaired in Plastid Lipid Import Reveals a Role of Membrane Lipids in Chloroplast Division

    SciTech Connect

    Fan, J.; Xu, C.

    2011-03-01

    The biogenesis of photosynthetic membranes in plants relies largely on lipid import from the endoplasmic reticulum (ER) and this lipid transport process is mediated by TGD proteins in Arabidopsis. Such a dependency of chloroplast biogenesis on ER-to-plastid lipid transport was recently exemplified by analyzing double mutants between tgd1-1 or tgd4-3 and fad6 mutants. The fad6 mutants are defective in the desaturation of membrane lipids in chloroplasts and therefore dependent on import of polyunsaturated lipid precursors from the ER for constructing a competent thylakoid membrane system. In support of a critical role of TGD proteins in ER-to-plastid lipid trafficking, we showed that the introduction of the tgd mutations into fad6 mutant backgrounds led to drastic reductions in relative amounts of thylakoid lipids. Moreover, the tgd1-1 fad6 and tgd4-3 fad6 double mutants were deficient in polyunsaturated fatty acids in chloroplast membrane lipids, and severely compromised in the biogenesis of photosynthetic membrane systems. Here we report that these double mutants are severely impaired in chloroplast division. The possible role of membrane lipids in chloroplast division is discussed.

  18. The Cell Wall Arabinose-Deficient Arabidopsis thaliana Mutant murus5 Encodes a Defective Allele of REVERSIBLY GLYCOSYLATED POLYPEPTIDE2.

    PubMed

    Dugard, Christopher K; Mertz, Rachel A; Rayon, Catherine; Mercadante, Davide; Hart, Christopher; Benatti, Matheus R; Olek, Anna T; SanMiguel, Phillip J; Cooper, Bruce R; Reiter, Wolf-Dieter; McCann, Maureen C; Carpita, Nicholas C

    2016-07-01

    Traditional marker-based mapping and next-generation sequencing was used to determine that the Arabidopsis (Arabidopsis thaliana) low cell wall arabinose mutant murus5 (mur5) encodes a defective allele of REVERSIBLY GLYCOSYLATED POLYPEPTIDE2 (RGP2). Marker analysis of 13 F2 confirmed mutant progeny from a recombinant mapping population gave a rough map position on the upper arm of chromosome 5, and deep sequencing of DNA from these 13 lines gave five candidate genes with G→A (C→T) transitions predicted to result in amino acid changes. Of these five, only insertional mutant alleles of RGP2, a gene that encodes a UDP-arabinose mutase that interconverts UDP-arabinopyranose and UDP-arabinofuranose, exhibited the low cell wall arabinose phenotype. The identities of mur5 and two SALK insertional alleles were confirmed by allelism tests and overexpression of wild-type RGP2 complementary DNA placed under the control of the 35S promoter in the three alleles. The mur5 mutation results in the conversion of cysteine-257 to tyrosine-257 within a conserved hydrophobic cluster predicted to be distal to the active site and essential for protein stability and possible heterodimerization with other isoforms of RGP.

  19. Arabidopsis mutant sk156 reveals complex regulation of SPL15 in a miR156-controlled gene network

    PubMed Central

    2012-01-01

    Background The Arabidopsis microRNA156 (miR156) regulates 11 members of the SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) family by base pairing to complementary target mRNAs. Each SPL gene further regulates a set of other genes; thus, miR156 controls numerous genes through a complex gene regulation network. Increased axillary branching occurs in transgenic Arabidopsis overexpressing miR156b, similar to that observed in loss-of-function max3 and max4 mutants with lesions in carotenoid cleavage dioxygenases. Arabidopsis miR156b was found to enhance carotenoid levels and reproductive shoot branching when expressed in Brassica napus, suggesting a link between miR156b expression and carotenoid metabolism. However, details of the miR156 regulatory network of SPL genes related to carotenoid metabolism are not known. Results In this study, an Arabidopsis T-DNA enhancer mutant, sk156, was identified due to its altered branching and trichome morphology and increased seed carotenoid levels compared to wild type (WT) ecovar Columbia. Enhanced miR156b expression due to the 35S enhancers present on the T-DNA insert was responsible for these phenotypes. Constitutive and leaf primodium-specific expression of a miR156-insensitive (mutated) SPL15 (SPL15m) largely restored WT seed carotenoid levels and plant morphology when expressed in sk156. The Arabidopsis native miR156-sensitive SPL15 (SPL15n) and SPL15m driven by a native SPL15 promoter did not restore the WT phenotype in sk156. Our findings suggest that SPL15 function is somewhat redundant with other SPL family members, which collectively affect plant phenotypes. Moreover, substantially decreased miR156b transcript levels in sk156 expressing SPL15m, together with the presence of multiple repeats of SPL-binding GTAC core sequence close to the miR156b transcription start site, suggested feedback regulation of miR156b expression by SPL15. This was supported by the demonstration of specific in vitro interaction between DNA

  20. Arabidopsis SUC1 loads the phloem in suc2 mutants when expressed from the SUC2 promoter

    PubMed Central

    Wippel, Kathrin; Sauer, Norbert

    2012-01-01

    Active loading of sucrose into phloem companion cells (CCs) is an essential process in apoplastic loaders, such as Arabidopsis or tobacco (Nicotiana sp.), and is even used by symplastic loaders such as melon (Cucumis melo) under certain stress conditions. Reduction of the amount or complete removal of the transporters catalysing this transport step results in severe developmental defects. Here we present analyses of two Arabidopsis lines, suc2-4 and suc2-5, that carry a null allele of the SUC2 gene which encodes the Arabidopsis phloem loader. These lines were complemented with constructs expressing either the Arabidopsis SUC1 or the Ustilago maydis srt1 cDNA from the SUC2 promoter. Both SUC1 and Srt1 are energy-dependent sucrose/H+ symporters and differ in specific kinetic properties from the SUC2 protein. Transgene expression was confirmed by RT-PCRs, the subcellular localization of Srt1 in planta with an Srt1-RFP fusion, and the correct CC-specific localization of the recombinant proteins by immunolocalization with anti-Srt1 and anti-SUC1 antisera. The transport capacity of Srt1 was studied in Srt1-GFP expressing Arabidopsis protoplasts. Although both proteins were found exclusively in CCs, only SUC1 complemented the developmental defects of suc2-4 and suc2-5 mutants. As SUC1 and Srt1 are well characterized, this result provides an insight into the properties that are essential for sucrose transporters to load the phloem successfully. PMID:22021573

  1. Expression Pattern of Two Paralogs Encoding Cinnamyl Alcohol Dehydrogenases in Arabidopsis. Isolation and Characterization of the Corresponding Mutants1

    PubMed Central

    Sibout, Richard; Eudes, Aymerick; Pollet, Brigitte; Goujon, Thomas; Mila, Isabelle; Granier, Fabienne; Séguin, Armand; Lapierre, Catherine; Jouanin, Lise

    2003-01-01

    Studying Arabidopsis mutants of the phenylpropanoid pathway has unraveled several biosynthetic steps of monolignol synthesis. Most of the genes leading to monolignol synthesis have been characterized recently in this herbaceous plant, except those encoding cinnamyl alcohol dehydrogenase (CAD). We have used the complete sequencing of the Arabidopsis genome to highlight a new view of the complete CAD gene family. Among nine AtCAD genes, we have identified the two distinct paralogs AtCAD-C and AtCAD-D, which share 75% identity and are likely to be involved in lignin biosynthesis in other plants. Northern, semiquantitative restriction fragment-length polymorphism-reverse transcriptase-polymerase chain reaction and western analysis revealed that AtCAD-C and AtCAD-D mRNA and protein ratios were organ dependent. Promoter activities of both genes are high in fibers and in xylem bundles. However, AtCAD-C displayed a larger range of sites of expression than AtCAD-D. Arabidopsis null mutants (Atcad-D and Atcad-C) corresponding to both genes were isolated. CAD activities were drastically reduced in both mutants, with a higher impact on sinapyl alcohol dehydrogenase activity (6% and 38% of residual sinapyl alcohol dehydrogenase activities for Atcad-D and Atcad-C, respectively). Only Atcad-D showed a slight reduction in Klason lignin content and displayed modifications of lignin structure with a significant reduced proportion of conventional S lignin units in both stems and roots, together with the incorporation of sinapaldehyde structures ether linked at Cβ. These results argue for a substantial role of AtCAD-D in lignification, and more specifically in the biosynthesis of sinapyl alcohol, the precursor of S lignin units. PMID:12805615

  2. The gravitropism defective 2 mutants of Arabidopsis are deficient in a protein implicated in endocytosis in Caenorhabditis elegans.

    PubMed

    Silady, Rebecca A; Kato, Takehide; Lukowitz, Wolfgang; Sieber, Patrick; Tasaka, Masao; Somerville, Chris R

    2004-10-01

    The gravitropism defective 2 (grv2) mutants of Arabidopsis show reduced shoot phototropism and gravitropism. Amyloplasts in the shoot endodermal cells of grv2 do not sediment to the same degree as in wild type. The GRV2 gene encodes a 277-kD polypeptide that is 42% similar to the Caenorhabditis elegans RME-8 protein, which is required for endocytosis. We hypothesize that a defect in endocytosis may affect both the initial gravity sensing via amyloplasts sedimentation and the subsequent more general tropic growth response.

  3. Delayed degradation of chlorophylls and photosynthetic proteins in Arabidopsis autophagy mutants during stress-induced leaf yellowing.

    PubMed

    Sakuraba, Yasuhito; Lee, Sang-Hwa; Kim, Ye-Sol; Park, Ohkmae K; Hörtensteiner, Stefan; Paek, Nam-Chon

    2014-07-01

    Plant autophagy, one of the essential proteolysis systems, balances proteome and nutrient levels in cells of the whole plant. Autophagy has been studied by analysing Arabidopsis thaliana autophagy-defective atg mutants, but the relationship between autophagy and chlorophyll (Chl) breakdown during stress-induced leaf yellowing remains unclear. During natural senescence or under abiotic-stress conditions, extensive cell death and early yellowing occurs in the leaves of atg mutants. A new finding is revealed that atg5 and atg7 mutants exhibit a functional stay-green phenotype under mild abiotic-stress conditions, but leaf yellowing proceeds normally in wild-type leaves under these conditions. Under mild salt stress, atg5 leaves retained high levels of Chls and all photosystem proteins and maintained a normal chloroplast structure. Furthermore, a double mutant of atg5 and non-functional stay-green nonyellowing1-1 (atg5 nye1-1) showed a much stronger stay-green phenotype than either single mutant. Taking these results together, it is proposed that autophagy functions in the non-selective catabolism of Chls and photosynthetic proteins during stress-induced leaf yellowing, in addition to the selective degradation of Chl-apoprotein complexes in the chloroplasts through the senescence-induced STAY-GREEN1/NYE1 and Chl catabolic enzymes.

  4. Photosynthetic performance of an Arabidopsis mutant with elevated stomatal density (sdd1-1) under different light regimes.

    PubMed

    Schlüter, Urte; Muschak, Michael; Berger, Dieter; Altmann, Thomas

    2003-02-01

    In the Arabidopsis mutant sdd1-1, a point mutation in a single gene (SDD1) causes specific alterations in stomatal density and distribution. In comparison to the wild type (C24), abaxial surfaces of sdd1-1 rosette leaves have about 2.5-fold higher stomatal densities. This mutant was used to study the consequence of stomatal density on photosynthesis under various light regimes. The increased stomatal density in the mutant had no significant influence on the leaf CO(2) assimilation rate (A) under constant light conditions. Mutant and wild-type plants contained similar amounts of carbohydrates under these conditions. However, exposure of plants to increasing photon flux densities resulted in differences in gas exchange and the carbohydrate metabolism of the wild type and mutant. Increased stomatal densities in sdd1-1 enabled low-light-adapted plants to have 30% higher CO(2) assimilation rates compared to the wild type when exposed to high light intensities. After 2 d under high light conditions leaves of sdd1-1 accumulated 30% higher levels of starch and hexoses than wild-type plants.

  5. Arabidopsis serotonin N-acetyltransferase knockout mutant plants exhibit decreased melatonin and salicylic acid levels resulting in susceptibility to an avirulent pathogen.

    PubMed

    Lee, Hyoung Yool; Byeon, Yeong; Tan, Dun-Xian; Reiter, Russel J; Back, Kyoungwhan

    2015-04-01

    Serotonin N-acetyltransferase (SNAT) is the penultimate enzyme in the melatonin biosynthesis pathway in plants. We examined the effects of SNAT gene inactivation in two Arabidopsis T-DNA insertion mutant lines. After inoculation with the avirulent pathogen Pseudomonas syringe pv. tomato DC3000 harboring the elicitor avrRpt2 (Pst-avrRpt2), melatonin levels in the snat knockout mutant lines were 50% less than in wild-type Arabidopsis Col-0 plants. The snat knockout mutant lines exhibited susceptibility to pathogen infection that coincided with decreased induction of defense genes including PR1, ICS1, and PDF1.2. Because melatonin acts upstream of salicylic acid (SA) synthesis, the reduced melatonin levels in the snat mutant lines led to decreased SA levels compared to wild-type, suggesting that the increased pathogen susceptibility of the snat mutant lines could be attributed to decreased SA levels and subsequent attenuation of defense gene induction. Exogenous melatonin treatment failed to induce defense gene expression in nahG Arabidopsis plants, but restored the induction of defense gene expression in the snat mutant lines. In addition, melatonin caused translocation of NPR1 (nonexpressor of PR1) protein from the cytoplasm into the nucleus indicating that melatonin-elicited pathogen resistance in response to avirulent pathogen attack is SA-dependent in Arabidopsis.

  6. Metabolite Profiling of adh1 Mutant Response to Cold Stress in Arabidopsis

    PubMed Central

    Song, Yuan; Liu, Lijun; Wei, Yunzhu; Li, Gaopeng; Yue, Xiule; An, Lizhe

    2017-01-01

    As a result of global warming, vegetation suffers from repeated freeze-thaw cycles caused by more frequent short-term low temperatures induced by hail, snow, or night frost. Therefore, short-term freezing stress of plants should be investigated particularly in light of the current climatic conditions. Alcohol dehydrogenase (ADH) plays a central role in the metabolism of alcohols and aldehydes and it is a key enzyme in anaerobic fermentation. ADH1 responds to plant growth and environmental stress; however, the function of ADH1 in the response to short-term freezing stress remains unknown. Using real-time quantitative fluorescence PCR, the expression level of ADH1 was analyzed at low temperature (4°C). The lethal temperature was calculated based on the electrolyte leakage tests for both ADH1 deletion mutants (adh1) and wild type (WT) plants. To further investigate the relationship between ADH1 and cold tolerance in plants, low-Mr polar metabolite analyses of Arabidopsis adh1 and WT were performed at cold temperatures using gas chromatography-mass spectrometry. This investigation focused on freezing treatments (cold acclimation group: −6°C for 2 h with prior 4°C for 7 d, cold shock group: −6°C for 2 h without cold acclimation) and recovery (23°C for 24 h) with respect to seedling growth at optimum temperature. The experimental results revealed a significant increase in ADH1 expression during low temperature treatment (4°C) and at a higher lethal temperature in adh1 compared to that in the WT. Retention time indices and specific mass fragments were used to monitor 263 variables and annotate 78 identified metabolites. From these analyses, differences in the degree of metabolite accumulation between adh1 and WT were detected, including soluble sugars (e.g., sucrose) and amino acids (e.g., asparagine). In addition, the correlation-based network analysis highlighted some metabolites, e.g., melibiose, fumaric acid, succinic acid, glycolic acid, and xylose, which

  7. Metabolite Profiling of adh1 Mutant Response to Cold Stress in Arabidopsis.

    PubMed

    Song, Yuan; Liu, Lijun; Wei, Yunzhu; Li, Gaopeng; Yue, Xiule; An, Lizhe

    2016-01-01

    As a result of global warming, vegetation suffers from repeated freeze-thaw cycles caused by more frequent short-term low temperatures induced by hail, snow, or night frost. Therefore, short-term freezing stress of plants should be investigated particularly in light of the current climatic conditions. Alcohol dehydrogenase (ADH) plays a central role in the metabolism of alcohols and aldehydes and it is a key enzyme in anaerobic fermentation. ADH1 responds to plant growth and environmental stress; however, the function of ADH1 in the response to short-term freezing stress remains unknown. Using real-time quantitative fluorescence PCR, the expression level of ADH1 was analyzed at low temperature (4°C). The lethal temperature was calculated based on the electrolyte leakage tests for both ADH1 deletion mutants (adh1) and wild type (WT) plants. To further investigate the relationship between ADH1 and cold tolerance in plants, low-Mr polar metabolite analyses of Arabidopsis adh1 and WT were performed at cold temperatures using gas chromatography-mass spectrometry. This investigation focused on freezing treatments (cold acclimation group: -6°C for 2 h with prior 4°C for 7 d, cold shock group: -6°C for 2 h without cold acclimation) and recovery (23°C for 24 h) with respect to seedling growth at optimum temperature. The experimental results revealed a significant increase in ADH1 expression during low temperature treatment (4°C) and at a higher lethal temperature in adh1 compared to that in the WT. Retention time indices and specific mass fragments were used to monitor 263 variables and annotate 78 identified metabolites. From these analyses, differences in the degree of metabolite accumulation between adh1 and WT were detected, including soluble sugars (e.g., sucrose) and amino acids (e.g., asparagine). In addition, the correlation-based network analysis highlighted some metabolites, e.g., melibiose, fumaric acid, succinic acid, glycolic acid, and xylose, which

  8. Transcriptional profiles of Arabidopsis stomataless mutants reveal developmental and physiological features of life in the absence of stomata.

    PubMed

    de Marcos, Alberto; Triviño, Magdalena; Pérez-Bueno, María Luisa; Ballesteros, Isabel; Barón, Matilde; Mena, Montaña; Fenoll, Carmen

    2015-01-01

    Loss of function of the positive stomata development regulators SPCH or MUTE in Arabidopsis thaliana renders stomataless plants; spch-3 and mute-3 mutants are extreme dwarfs, but produce cotyledons and tiny leaves, providing a system to interrogate plant life in the absence of stomata. To this end, we compared their cotyledon transcriptomes with that of wild-type plants. K-means clustering of differentially expressed genes generated four clusters: clusters 1 and 2 grouped genes commonly regulated in the mutants, while clusters 3 and 4 contained genes distinctively regulated in mute-3. Classification in functional categories and metabolic pathways of genes in clusters 1 and 2 suggested that both mutants had depressed secondary, nitrogen and sulfur metabolisms, while only a few photosynthesis-related genes were down-regulated. In situ quenching analysis of chlorophyll fluorescence revealed limited inhibition of photosynthesis. This and other fluorescence measurements matched the mutant transcriptomic features. Differential transcriptomes of both mutants were enriched in growth-related genes, including known stomata development regulators, which paralleled their epidermal phenotypes. Analysis of cluster 3 was not informative for developmental aspects of mute-3. Cluster 4 comprised genes differentially up-regulated in mute-3, 35% of which were direct targets for SPCH and may relate to the unique cell types of mute-3. A screen of T-DNA insertion lines in genes differentially expressed in the mutants identified a gene putatively involved in stomata development. A collection of lines for conditional overexpression of transcription factors differentially expressed in the mutants rendered distinct epidermal phenotypes, suggesting that these proteins may be novel stomatal development regulators. Thus, our transcriptome analysis represents a useful source of new genes for the study of stomata development and for characterizing physiology and growth in the absence of stomata.

  9. Transcriptional profiles of Arabidopsis stomataless mutants reveal developmental and physiological features of life in the absence of stomata

    PubMed Central

    de Marcos, Alberto; Triviño, Magdalena; Pérez-Bueno, María Luisa; Ballesteros, Isabel; Barón, Matilde; Mena, Montaña; Fenoll, Carmen

    2015-01-01

    Loss of function of the positive stomata development regulators SPCH or MUTE in Arabidopsis thaliana renders stomataless plants; spch-3 and mute-3 mutants are extreme dwarfs, but produce cotyledons and tiny leaves, providing a system to interrogate plant life in the absence of stomata. To this end, we compared their cotyledon transcriptomes with that of wild-type plants. K-means clustering of differentially expressed genes generated four clusters: clusters 1 and 2 grouped genes commonly regulated in the mutants, while clusters 3 and 4 contained genes distinctively regulated in mute-3. Classification in functional categories and metabolic pathways of genes in clusters 1 and 2 suggested that both mutants had depressed secondary, nitrogen and sulfur metabolisms, while only a few photosynthesis-related genes were down-regulated. In situ quenching analysis of chlorophyll fluorescence revealed limited inhibition of photosynthesis. This and other fluorescence measurements matched the mutant transcriptomic features. Differential transcriptomes of both mutants were enriched in growth-related genes, including known stomata development regulators, which paralleled their epidermal phenotypes. Analysis of cluster 3 was not informative for developmental aspects of mute-3. Cluster 4 comprised genes differentially up−regulated in mute−3, 35% of which were direct targets for SPCH and may relate to the unique cell types of mute−3. A screen of T-DNA insertion lines in genes differentially expressed in the mutants identified a gene putatively involved in stomata development. A collection of lines for conditional overexpression of transcription factors differentially expressed in the mutants rendered distinct epidermal phenotypes, suggesting that these proteins may be novel stomatal development regulators. Thus, our transcriptome analysis represents a useful source of new genes for the study of stomata development and for characterizing physiology and growth in the absence of

  10. Analysis of metabolic flux phenotypes for two Arabidopsis mutants with severe impairment in seed storage lipid synthesis

    SciTech Connect

    Lonien, J.; Schwender, J.

    2009-11-01

    Major storage reserves of Arabidopsis (Arabidopsis thaliana) seeds are triacylglycerols (seed oils) and proteins. Seed oil content is severely reduced for the regulatory mutant wrinkled1 (wri1-1; At3g54320) and for a double mutant in two isoforms of plastidic pyruvate kinase (pkp{beta}{sub 1}pkp{alpha}; At5g52920 and At3g22960). Both already biochemically well-characterized mutants were now studied by {sup 13}C metabolic flux analysis of cultured developing embryos based on comparison with their respective genetic wild-type backgrounds. For both mutations, in seeds as well as in cultured embryos, the oil fraction was strongly reduced while the fractions of proteins and free metabolites increased. Flux analysis in cultured embryos revealed changes in nutrient uptakes and fluxes into biomass as well as an increase in tricarboxylic acid cycle activity for both mutations. While in both wild types plastidic pyruvate kinase (PK{sub p}) provides most of the pyruvate for plastidic fatty acid synthesis, the flux through PK{sub p} is reduced in pkp{beta}{sub 1}pkp{alpha} by 43% of the wild-type value. In wri1-1, PK{sub p} flux is even more reduced (by 82%), although the genes PKp{beta}{sub 1} and PKp{alpha} are still expressed. Along a common paradigm of metabolic control theory, it is hypothesized that a large reduction in PK{sub p} enzyme activity in pkp{beta}{sub 1}pkp{alpha} has less effect on PK{sub p} flux than multiple smaller reductions in glycolytic enzymes in wri1-1. In addition, only in the wri1-1 mutant is the large reduction in PK{sub p} flux compensated in part by an increased import of cytosolic pyruvate and by plastidic malic enzyme. No such limited compensatory bypass could be observed in pkp{beta}{sub 1}pkp{alpha}.

  11. Arabidopsis mutants in sphingolipid synthesis as tools to understand the structure and function of membrane microdomains in plasmodesmata

    PubMed Central

    González-Solís, Ariadna; Cano-Ramírez, Dora L.; Morales-Cedillo, Francisco; Tapia de Aquino, Cinthya; Gavilanes-Ruiz, Marina

    2013-01-01

    Plasmodesmata—intercellular channels that communicate adjacent cells—possess complex membranous structures. Recent evidences indicate that plasmodesmata contain membrane microdomains. In order to understand how these submembrane regions collaborate to plasmodesmata function, it is necessary to characterize their size, composition and dynamics. An approach that can shed light on these microdomain features is based on the use of Arabidopsis mutants in sphingolipid synthesis. Sphingolipids are canonical components of microdomains together with sterols and some glycerolipids. Moreover, sphingolipids are transducers in pathways that display programmed cell death as a defense mechanism against pathogens. The study of Arabidopsis mutants would allow determining which structural features of the sphingolipids are important for the formation and stability of microdomains, and if defense signaling networks using sphingoid bases as second messengers are associated to plasmodesmata operation. Such studies need to be complemented by analysis of the ultrastructure and the use of protein probes for plasmodesmata microdomains and may constitute a very valuable source of information to analyze these membrane structures. PMID:24478783

  12. Methyl jasmonate inhibition of root growth and induction of a leaf protein are decreased in an Arabidopsis thaliana mutant.

    PubMed Central

    Staswick, P E; Su, W; Howell, S H

    1992-01-01

    Jasmonic acid and its methyl ester, methyl jasmonate (MeJA), are plant signaling molecules that affect plant growth and gene expression. Primary root growth of wild-type Arabidopsis thaliana seedlings was inhibited 50% when seedlings were grown on agar medium containing 0.1 M MeJA. An ethyl methanesulfonate mutant (jar1) with decreased sensitivity to MeJA inhibition of root elongation was isolated and characterized. Genetic data indicated the trait was recessive and controlled by a single Mendelian factor. MeJA-induced polypeptides were detected in Arabidopsis leaves by antiserum to a MeJA-inducible vegetative storage protein from soybean. The induction of these proteins by MeJA in the mutant was at least 4-fold less in jar1 compared to wild type. In contrast, seeds of jar1 plants were more sensitive than wild type to inhibition of germination by abscisic acid. These results suggest that the defect in jar1 affects a general jasmonate response pathway, which may regulate multiple genes in different plant organs. Images PMID:11607311

  13. Characterization of a JAZ7 activation-tagged Arabidopsis mutant with increased susceptibility to the fungal pathogen Fusarium oxysporum.

    PubMed

    Thatcher, Louise F; Cevik, Volkan; Grant, Murray; Zhai, Bing; Jones, Jonathan D G; Manners, John M; Kazan, Kemal

    2016-04-01

    In Arabidopsis, jasmonate (JA)-signaling plays a key role in mediating Fusarium oxysporum disease outcome. However, the roles of JASMONATE ZIM-domain (JAZ) proteins that repress JA-signaling have not been characterized in host resistance or susceptibility to this pathogen. Here, we found most JAZ genes are induced following F. oxysporum challenge, and screening T-DNA insertion lines in Arabidopsis JAZ family members identified a highly disease-susceptible JAZ7 mutant (jaz7-1D). This mutant exhibited constitutive JAZ7 expression and conferred increased JA-sensitivity, suggesting activation of JA-signaling. Unlike jaz7 loss-of-function alleles, jaz7-1D also had enhanced JA-responsive gene expression, altered development and increased susceptibility to the bacterial pathogen PstDC3000 that also disrupts host JA-responses. We also demonstrate that JAZ7 interacts with transcription factors functioning as activators (MYC3, MYC4) or repressors (JAM1) of JA-signaling and contains a functional EAR repressor motif mediating transcriptional repression via the co-repressor TOPLESS (TPL). We propose through direct TPL recruitment, in wild-type plants JAZ7 functions as a repressor within the JA-response network and that in jaz7-1D plants, misregulated ectopic JAZ7 expression hyper-activates JA-signaling in part by disturbing finely-tuned COI1-JAZ-TPL-TF complexes.

  14. Leaf hydraulic conductance varies with vein anatomy across Arabidopsis thaliana wild-type and leaf vein mutants.

    PubMed

    Caringella, Marissa A; Bongers, Franca J; Sack, Lawren

    2015-12-01

    Leaf venation is diverse across plant species and has practical applications from paleobotany to modern agriculture. However, the impact of vein traits on plant performance has not yet been tested in a model system such as Arabidopsis thaliana. Previous studies analysed cotyledons of A. thaliana vein mutants and identified visible differences in their vein systems from the wild type (WT). We measured leaf hydraulic conductance (Kleaf ), vein traits, and xylem and mesophyll anatomy for A. thaliana WT (Col-0) and four vein mutants (dot3-111 and dot3-134, and cvp1-3 and cvp2-1). Mutant true leaves did not possess the qualitative venation anomalies previously shown in the cotyledons, but varied quantitatively in vein traits and leaf anatomy across genotypes. The WT had significantly higher mean Kleaf . Across all genotypes, there was a strong correlation of Kleaf with traits related to hydraulic conductance across the bundle sheath, as influenced by the number and radial diameter of bundle sheath cells and vein length per area. These findings support the hypothesis that vein traits influence Kleaf , indicating the usefulness of this mutant system for testing theory that was primarily established comparatively across species, and supports a strong role for the bundle sheath in influencing Kleaf .

  15. Isolation of an Arabidopsis mutant lacking vitamin E and identification of a cyclase essential for all tocopherol biosynthesis.

    PubMed

    Porfirova, Svetlana; Bergmuller, Eveline; Tropf, Susanne; Lemke, Rainer; Dormann, Peter

    2002-09-17

    Tocopherol (vitamin E) is a plant chloroplast lipid presumed to be involved in the response to oxidative stress. A tocopherol-deficient mutant (vte1) was isolated from Arabidopsis thaliana by using a TLC-based screening approach. Mutant plants lacked all four tocopherol forms and were deficient in tocopherol cyclase activity. Genetic mapping of vte1 and a genomics-based approach led to the identification of the ORF At4g32770 as a candidate gene for tocopherol cyclase. In vte1, At4g32770 contains a splicing site mutation and the corresponding mRNA expression is reduced. Expression of VTE1 in Escherichia coli resulted in the production of a protein with high tocopherol cyclase and tocotrienol cyclase activity. The VTE1 sequence shows no similarities to genes with known function, but is similar to that of SXD1, a gene that was recently isolated based on the availability of the sucrose export defective1 maize mutant (sxd1). Growth of the vte1 mutant, chlorophyll content, and photosynthetic quantum yield were similar to wild type under optimal growth conditions. Therefore, absence of tocopherol has no large impact on photosynthesis or plant viability, suggesting that other antioxidants can compensate for the loss of tocopherol. During photo-oxidative stress, chlorophyll content and photosynthetic quantum yield were slightly reduced in vte1 as compared with wild type indicating a potential role for tocopherol in maintaining an optimal photosynthesis rate under high-light stress.

  16. The Arabidopsis salt overly sensitive 4 Mutants Uncover a Critical Role for Vitamin B6 in Plant Salt Tolerance

    PubMed Central

    Shi, Huazhong; Xiong, Liming; Stevenson, Becky; Lu, Tiegang; Zhu, Jian-Kang

    2002-01-01

    Salt stress is a major environmental factor influencing plant growth and development. To identify salt tolerance determinants, a genetic screen for salt overly sensitive (sos) mutants was performed in Arabidopsis. We present here the characterization of sos4 mutants and the positional cloning of the SOS4 gene. sos4 mutant plants are hypersensitive to Na+, K+, and Li+ ions. Under NaCl stress, sos4 plants accumulate more Na+ and retain less K+ compared with wild-type plants. SOS4 encodes a pyridoxal kinase that is involved in the biosynthesis of pyridoxal-5-phosphate, an active form of vitamin B6. The expression of SOS4 cDNAs complements an Escherichia coli mutant defective in pyridoxal kinase. Supplementation of pyridoxine but not pyridoxal in the growth medium can partially rescue the sos4 defect in salt tolerance. SOS4 is expressed ubiquitously in all plant tissues. As a result of alternative splicing, two transcripts are derived from the SOS4 gene, the relative abundance of which is modulated by development and environmental stresses. Besides being essential cofactors for numerous enzymes, as shown by pharmacological studies in animal cells, pyridoxal-5-phosphate and its derivatives are also ligands for P2X receptor ion channels. Our results demonstrate that pyridoxal kinase is a novel salt tolerance determinant important for the regulation of Na+ and K+ homeostasis in plants. We propose that pyridoxal-5-phosphate regulates Na+ and K+ homeostasis by modulating the activities of ion transporters. PMID:11910005

  17. Dissection of the Complex Phenotype in Cuticular Mutants of Arabidopsis Reveals a Role of SERRATE as a Mediator

    PubMed Central

    Voisin, Derry; Nawrath, Christiane; Kurdyukov, Sergey; Franke, Rochus B.; Reina-Pinto, José J.; Efremova, Nadia; Will, Isa; Schreiber, Lukas; Yephremov, Alexander

    2009-01-01

    Mutations in LACERATA (LCR), FIDDLEHEAD (FDH), and BODYGUARD (BDG) cause a complex developmental syndrome that is consistent with an important role for these Arabidopsis genes in cuticle biogenesis. The genesis of their pleiotropic phenotypes is, however, poorly understood. We provide evidence that neither distorted depositions of cutin, nor deficiencies in the chemical composition of cuticular lipids, account for these features, instead suggesting that the mutants alleviate the functional disorder of the cuticle by reinforcing their defenses. To better understand how plants adapt to these mutations, we performed a genome-wide gene expression analysis. We found that apparent compensatory transcriptional responses in these mutants involve the induction of wax, cutin, cell wall, and defense genes. To gain greater insight into the mechanism by which cuticular mutations trigger this response in the plants, we performed an overlap meta-analysis, which is termed MASTA (MicroArray overlap Search Tool and Analysis), of differentially expressed genes. This suggested that different cell integrity pathways are recruited in cesA cellulose synthase and cuticular mutants. Using MASTA for an in silico suppressor/enhancer screen, we identified SERRATE (SE), which encodes a protein of RNA–processing multi-protein complexes, as a likely enhancer. In confirmation of this notion, the se lcr and se bdg double mutants eradicate severe leaf deformations as well as the organ fusions that are typical of lcr and bdg and other cuticular mutants. Also, lcr does not confer resistance to Botrytis cinerea in a se mutant background. We propose that there is a role for SERRATE-mediated RNA signaling in the cuticle integrity pathway. PMID:19876373

  18. Auxin transport in an auxin-resistant mutant of arabidopsis thaliana

    SciTech Connect

    Lincoln, C.; Benning, C.; Estelle, M.

    1987-04-01

    The authors are studying a group of allelic recessive mutations in Arabidopsis called axr-1. Homozygous axr-1 plants are resistant to exogenously applied auxin. In addition, axr-1 mutations all confer a number of development abnormalities including an apparent reduction in apical dominance, loss of normal geotropic response, and a failure to self-fertilize due to a decrease in stamen elongation. In order to determine whether this pleiotropic phenotype is due to an alteration in auxin transport they have adapted the agar block transport assay for use in Arabidopsis stem segments. Their results indicate that as in other plant species, auxin transport is strongly polar in Arabidopsis stem segments. In addition transport is inhibited by the well characterized auxin transport inhibitor N-1-naphthylphthalamic acid and the artificial auxin 2,4-D. These results as well as the characterization of transport in axr-1 plants will be presented.

  19. Membrane function in lipid mutants of Arabidopsis. First year progress report

    SciTech Connect

    Browse, J.A.

    1993-06-01

    Progress on the biochemical characterization of the fad3 mutants deficient in 18:3 fatty acid synthesis and the fab2 mutant that accumulates increased amounts of 18:0 is described. Studies of the cell biology and physiology of the fab2 and fad2 mutants have provided evidence for some of the critical roles played by unsaturated fatty acids as components of plant membranes. Finally, the fab2 mutant has allowed us to carry out the first isolation and characterization of intergenic suppressor mutations in a higher plant.

  20. Iron-dependent modifications of the flower transcriptome, proteome, metabolome, and hormonal content in an Arabidopsis ferritin mutant

    PubMed Central

    Sudre, Damien; Gutierrez-Carbonell, Elain; Lattanzio, Giuseppe; Rellán-Álvarez, Rubén; Gaymard, Frédéric; Wohlgemuth, Gert; Fiehn, Oliver; Álvarez-Fernández, Ana; Zamarreño, Angel M.; Bacaicoa, Eva; Duy, Daniela; García-Mina, Jose-María; Abadía, Javier; Philippar, Katrin; López-Millán, Ana-Flor; Briat, Jean-François

    2013-01-01

    Iron homeostasis is an important process for flower development and plant fertility. The role of plastids in these processes has been shown to be essential. To document the relationships between plastid iron homeostasis and flower biology further, a global study (transcriptome, proteome, metabolome, and hormone analysis) was performed of Arabidopsis flowers from wild-type and triple atfer1-3-4 ferritin mutant plants grown under iron-sufficient or excess conditions. Some major modifications in specific functional categories were consistently observed at these three omic levels, although no significant overlaps of specific transcripts and proteins were detected. These modifications concerned redox reactions and oxidative stress, as well as amino acid and protein catabolism, this latter point being exemplified by an almost 10-fold increase in urea concentration of atfer1-3-4 flowers from plants grown under iron excess conditions. The mutant background caused alterations in Fe–haem redox proteins located in membranes and in hormone-responsive proteins. Specific effects of excess Fe in the mutant included further changes in these categories, supporting the idea that the mutant is facing a more intense Fe/redox stress than the wild type. The mutation and/or excess Fe had a strong impact at the membrane level, as denoted by the changes in the transporter and lipid metabolism categories. In spite of the large number of genes and proteins responsive to hormones found to be regulated in this study, changes in the hormonal balance were restricted to cytokinins, especially in the mutant plants grown under Fe excess conditions. PMID:23682113

  1. Characterization of a Cruciferin Deficient Mutant of Arabidopsis and Its Utility for Overexpression of Foreign Proteins in Plants

    PubMed Central

    Lin, Yimei; Pajak, Agnieszka; Marsolais, Frédéric; McCourt, Peter; Riggs, C. Daniel

    2013-01-01

    Plant seeds naturally accumulate storage reserves (proteins, carbohydrates, lipids) that are mobilized during germination to provide energy and raw materials to support early seedling growth. Seeds have been exploited as bioreactors for the production to foreign materials, but stable, high level expression has been elusive, in part due to the intrinsic bias for producing the natural reserves in their typical proportions. To identify mutants governing seed filling, we screened a population of mutagenized Arabidopsis plants for a mutant that failed to fill its seeds. Here we report the identification of ssp1, a recessive, viable mutant that accumulates approximately 15% less protein than wildtype seeds. Molecular analyses revealed that ssp1 is due to the introduction of a premature stop codon in CRU3, one of the major cruciferin genes. Unlike many other reserve mutants or transgenic lines in which seed storage protein levels are reduced by antisense/RNAi technologies, ssp1 exhibits low level compensation by other reserves, and represents a mutant background that might prove useful for high level expression of foreign proteins. To test this hypothesis, we used a bean phytohemagglutinin (PHA) gene as a reporter and compared PHA expression levels in single copy insertion lines in ssp1 vs. wildtype. These near isogenic lines allow reporter protein levels to be compared without the confounding and sometimes unknown influences of transgene copy number and position effects on gene expression. The ssp1 lines consistently accumulated more PHA than the backcrossed counterparts, with increases ranging from 12% to 126%. This proof of principle study suggests that similar strategies in crop plants may improve the yield of foreign proteins of agronomic and economic interest. PMID:23724110

  2. Iron-dependent modifications of the flower transcriptome, proteome, metabolome, and hormonal content in an Arabidopsis ferritin mutant.

    PubMed

    Sudre, Damien; Gutierrez-Carbonell, Elain; Lattanzio, Giuseppe; Rellán-Álvarez, Rubén; Gaymard, Frédéric; Wohlgemuth, Gert; Fiehn, Oliver; Alvarez-Fernández, Ana; Zamarreño, Angel M; Bacaicoa, Eva; Duy, Daniela; García-Mina, Jose-María; Abadía, Javier; Philippar, Katrin; López-Millán, Ana-Flor; Briat, Jean-François

    2013-07-01

    Iron homeostasis is an important process for flower development and plant fertility. The role of plastids in these processes has been shown to be essential. To document the relationships between plastid iron homeostasis and flower biology further, a global study (transcriptome, proteome, metabolome, and hormone analysis) was performed of Arabidopsis flowers from wild-type and triple atfer1-3-4 ferritin mutant plants grown under iron-sufficient or excess conditions. Some major modifications in specific functional categories were consistently observed at these three omic levels, although no significant overlaps of specific transcripts and proteins were detected. These modifications concerned redox reactions and oxidative stress, as well as amino acid and protein catabolism, this latter point being exemplified by an almost 10-fold increase in urea concentration of atfer1-3-4 flowers from plants grown under iron excess conditions. The mutant background caused alterations in Fe-haem redox proteins located in membranes and in hormone-responsive proteins. Specific effects of excess Fe in the mutant included further changes in these categories, supporting the idea that the mutant is facing a more intense Fe/redox stress than the wild type. The mutation and/or excess Fe had a strong impact at the membrane level, as denoted by the changes in the transporter and lipid metabolism categories. In spite of the large number of genes and proteins responsive to hormones found to be regulated in this study, changes in the hormonal balance were restricted to cytokinins, especially in the mutant plants grown under Fe excess conditions.

  3. The Evolutionarily Conserved Protein PHOTOSYNTHESIS AFFECTED MUTANT71 Is Required for Efficient Manganese Uptake at the Thylakoid Membrane in Arabidopsis

    PubMed Central

    Steinberger, Iris; Herdean, Andrei; Gandini, Chiara; Labs, Mathias; Flügge, Ulf-Ingo; Geimer, Stefan; Schmidt, Sidsel Birkelund; Husted, Søren; Spetea, Cornelia; Leister, Dario

    2016-01-01

    In plants, algae, and cyanobacteria, photosystem II (PSII) catalyzes the light-driven oxidation of water. The oxygen-evolving complex of PSII is a Mn4CaO5 cluster embedded in a well-defined protein environment in the thylakoid membrane. However, transport of manganese and calcium into the thylakoid lumen remains poorly understood. Here, we show that Arabidopsis thaliana PHOTOSYNTHESIS AFFECTED MUTANT71 (PAM71) is an integral thylakoid membrane protein involved in Mn2+ and Ca2+ homeostasis in chloroplasts. This protein is required for normal operation of the oxygen-evolving complex (as evidenced by oxygen evolution rates) and for manganese incorporation. Manganese binding to PSII was severely reduced in pam71 thylakoids, particularly in PSII supercomplexes. In cation partitioning assays with intact chloroplasts, Mn2+ and Ca2+ ions were differently sequestered in pam71, with Ca2+ enriched in pam71 thylakoids relative to the wild type. The changes in Ca2+ homeostasis were accompanied by an increased contribution of the transmembrane electrical potential to the proton motive force across the thylakoid membrane. PSII activity in pam71 plants and the corresponding Chlamydomonas reinhardtii mutant cgld1 was restored by supplementation with Mn2+, but not Ca2+. Furthermore, PAM71 suppressed the Mn2+-sensitive phenotype of the yeast mutant Δpmr1. Therefore, PAM71 presumably functions in Mn2+ uptake into thylakoids to ensure optimal PSII performance. PMID:27020959

  4. Liver and colon DNA oxidative damage and gene expression profiles of rats fed Arabidopsis thaliana mutant seeds containing contrasted flavonoids.

    PubMed

    Luceri, Cristina; Giovannelli, Lisa; Pitozzi, Vanessa; Toti, Simona; Castagnini, Cinzia; Routaboul, Jean-Marc; Lepiniec, Loic; Larrosa, Mar; Dolara, Piero

    2008-04-01

    Plant polyphenols, such as flavonoids, comprise many compounds, ranging from simple phenolic molecules (i.e. flavonols, anthocyanins) to polymeric structures with high molecular weight (as proanthocyanidins, PAs). We investigated the effects of flavonoids by feeding Wistar rats Arabidopsis thaliana seeds carrying mutations in key enzymes of the flavonoid biosynthetic pathway (15% w/w seeds for 4 weeks). The seeds used were: Ws-2 wild-type containing flavonols and PAs, tt3-4 mutant containing flavonols only, ban-5 accumulating flavonols and anthocyanins, tt4-8 mutant, deprived of flavonoids. DNA oxidative damage was significantly reduced only in the liver of rats fed tt3-4 mutant seeds. Microarray analysis of the liver revealed down-regulation of genes associated with oxidative stress, Krebs cycle, electron transport and proteasome degradation in all experimental groups compared to the tt4-8-fed reference rats; therefore, these effects were due to the flavonol content and not to high molecular weight compounds. We observed a down-regulation of inflammatory response genes in the colon mucosa in ban-5- fed rats, probably due to anthocyanin content. In conclusion, flavonols exhibited antioxidant effects at systemic level, whereas high molecular weight flavonoids affected only the colon, probably due to their limited absorption.

  5. Analysis of knockout mutants reveals non-redundant functions of poly(ADP-ribose)polymerase isoforms in Arabidopsis.

    PubMed

    Pham, Phuong Anh; Wahl, Vanessa; Tohge, Takayuki; de Souza, Laise Rosado; Zhang, Youjun; Do, Phuc Thi; Olas, Justyna J; Stitt, Mark; Araújo, Wagner L; Fernie, Alisdair R

    2015-11-01

    The enzyme poly(ADP-ribose)polymerase (PARP) has a dual function being involved both in the poly(ADP-ribosyl)ation and being a constituent of the NAD(+) salvage pathway. To date most studies, both in plant and non-plant systems, have focused on the signaling role of PARP in poly(ADP-ribosyl)ation rather than any role that can be ascribed to its metabolic function. In order to address this question we here used a combination of expression, transcript and protein localization studies of all three PARP isoforms of Arabidopsis alongside physiological analysis of the corresponding mutants. Our analyses indicated that whilst all isoforms of PARP were localized to the nucleus they are also present in non-nuclear locations with parp1 and parp3 also localised in the cytosol, and parp2 also present in the mitochondria. We next isolated and characterized insertional knockout mutants of all three isoforms confirming a complete knockout in the full length transcript levels of the target genes as well as a reduced total leaf NAD hydrolase activity in the two isoforms (PARP1, PARP2) that are highly expressed in leaves. Physiological evaluation of the mutant lines revealed that they displayed distinctive metabolic and root growth characteristics albeit unaltered leaf morphology under optimal growth conditions. We therefore conclude that the PARP isoforms play non-redundant non-nuclear metabolic roles and that their function is highly important in rapidly growing tissues such as the shoot apical meristem, roots and seeds.

  6. Morphological analysis of seed shape in Arabidopsis thaliana reveals altered polarity in mutants of the ethylene signaling pathway.

    PubMed

    Robert, Céline; Noriega, Arturo; Tocino, Angel; Cervantes, Emilio

    2008-06-16

    The shape of Arabidopsis thaliana dry seed is described here as a prolate spheroid. The accuracy of this approximation is discussed. Considering its limitations, it allows a geometric approximation to the analysis of changes occurring in seed shape during imbibition prior to seed germination as well as the differences in shape between genotypes and their changes during imbibition. The triple mutant ein2-1, ers1-2, etr1-7 presents notable alterations in seed shape. In addition, seeds of this and other mutants in the ethylene signaling pathway (ctr1-1, eto1-1, etr1-1, ein2-1) show different response to imbibition than the wild type. Imbibed seeds of the wild type increase their asymmetry compared with the dry seeds. This is detected by the relative changes in the curvature values in both poles. Thus, during imbibition of the wild-type seeds, the reduction in curvature values observed in the basal pole gives them an ovoid shape. In contrast, in the seeds of the ethylene mutants, reduction in curvature values occurs in both basal and apical poles, and its shape remains as a prolate spheroid. Our data indicate that the ethylene signaling pathway is involved, in general, in the complex regulation of seed shape and, in particular, in the establishment of polarity in seeds, controlling curvature values in the seed poles.

  7. The influence of microgravity and spaceflight on columella cell ultrastructure in starch-deficient mutants of Arabidopsis

    NASA Technical Reports Server (NTRS)

    Guisinger, M. M.; Kiss, J. Z.

    1999-01-01

    The ultrastructure of root cap columella cells was studied by morphometric analysis in wild-type, a reduced-starch mutant, and a starchless mutant of Arabidopsis grown in microgravity (F-microgravity) and compared to ground 1g (G-1g) and flight 1g (F-1g) controls. Seedlings of the wild-type and reduced-starch mutant that developed during an experiment on the Space Shuttle (both the F-microgravity samples and the F-lg control) exhibited a decreased starch content in comparison to the G-1g control. These results suggest that some factor associated with spaceflight (and not microgravity per se) affects starch metabolism. Elevated levels of ethylene were found during the experiments on the Space Shuttle, and analysis of ground controls with added ethylene demonstrated that this gas was responsible for decreased starch levels in the columella cells. This is the first study to use an on-board centrifuge as a control when quantifying starch in spaceflight-grown plants. Furthermore, our results show that ethylene levels must be carefully considered and controlled when designing experiments with plants for the International Space Station.

  8. Uncoupling PR gene expression from NPR1 and bacterial resistance: characterization of the dominant Arabidopsis cpr6-1 mutant.

    PubMed Central

    Clarke, J D; Liu, Y; Klessig, D F; Dong, X

    1998-01-01

    In Arabidopsis, NPR1 mediates the salicylic acid (SA)-induced expression of pathogenesis-related (PR) genes and systemic acquired resistance (SAR). Here, we report the identification of another component, CPR 6, that may function with NPR1 in regulating PR gene expression. The dominant CPR 6-1 mutant expresses the SA/NPR1-regulated PR genes (PR-1, BGL 2, and PR-5) and displays enhanced resistance to Pseudomonas syringae pv maculicola ES4326 and Peronospora parasitica Noco2 in the absence of SAR induction. cpr 6-1-induced PR gene expression is not suppressed in the cpr 6-1 npr1-1 double mutant but is suppressed when SA is removed by salicylate hydroxylase. Thus, constitutive PR gene expression in cpr 6-1 requires SA but not NPR1. In addition, resistance to P. s. maculicola ES4326 is suppressed in the cpr 6-1 npr1-1 double mutant, despite expression of PR-1, BGL 2, and PR-5. Resistance to P. s. maculicola ES4326 must therefore be accomplished through unidentified antibacterial gene products that are regulated through NPR1. These results show that CPR 6 is an important regulator of multiple signal transduction pathways involved in plant defense. PMID:9548982

  9. Use of the "gl1" Mutant and the "CA-rop2" Transgenic Plants of "Arabidopsis thaliana" in the Biology Laboratory Course

    ERIC Educational Resources Information Center

    Zheng, Zhi-Liang

    2006-01-01

    This article describes the use of the "glabrous1 (g11)" mutant and constitutively active "(CA)-rop2" transgenic plants of "Arabidopsis thaliana" in teaching genetics laboratory for both high school and undergraduate students. The experiments provide students with F[subscript 1] and F[subscript 2] generations within a semester for genetic and…

  10. The mur4 mutant of arabidopsis is partially defective in the de novo synthesis of uridine diphospho L-arabinose

    SciTech Connect

    Burget, E.G.; Reiter, W.D.

    1999-10-01

    To obtain information on the synthesis and function of arabinosylated glycans, the mur4 mutant of arabidopsis was characterized. This mutation leads to a 50% reduction in the monosaccharide L-arabinose in most organs and affects arabinose-containing pectic cell wall polysaccharides and arabinogalactan proteins. Feeding L-arabinose to mur4 plants restores the cell wall composition to wild-type levels, suggesting a partial defect in the de novo synthesis of UDP-L-arabinose, the activated sugar used by arabinosyltransferases. The defect was traced to the conversion of UDP-D-xylose to UDP-L-arabinose in the microsome fraction of leaf material, indicating that mur4 plants are defective in a membrane-bound UDP-D-xylose 4-epimerase.

  11. The impact of PEPC phosphorylation on growth and development of Arabidopsis thaliana: molecular and physiological characterization of PEPC kinase mutants.

    PubMed

    Meimoun, Patrice; Gousset-Dupont, Aurélie; Lebouteiller, Bénédicte; Ambard-Bretteville, Françoise; Besin, Evelyne; Lelarge, Caroline; Mauve, Caroline; Hodges, Michael; Vidal, Jean

    2009-05-19

    Two phosphoenolpyruvate carboxylase (PEPC) kinase genes (PPCk1 and PPCk2) are present in the Arabidopsis genome; only PPCk1 is expressed in rosette leaves. Homozygous lines of two independent PPCk1 T-DNA-insertional mutants showed very little (dln1), or no (csi8) light-induced PEPC phosphorylation and a clear retard in growth under our greenhouse conditions. A mass-spectrometry-based analysis revealed significant changes in metabolite profiles. However, the anaplerotic pathway initiated by PEPC was only moderately altered. These data establish the PPCk1 gene product as responsible for leaf PEPC phosphorylation in planta and show that the absence of PEPC phosphorylation has pleiotropic consequences on plant metabolism.

  12. Genome-wide Hi-C analyses in wild type and mutants reveal high-resolution chromatin interactions in Arabidopsis

    PubMed Central

    Feng, Suhua; Cokus, Shawn J.; Schubert, Veit; Zhai, Jixian; Pellegrini, Matteo; Jacobsen, Steven E.

    2015-01-01

    SUMMARY Chromosomes form three-dimensional structures that are critical to the regulation of cellular and genetic processes. Here, we present a study of global chromatin interaction patterns in Arabidopsis thaliana. Our genome-wide approach confirmed interactions that were previously observed by other methods as well as uncovered previously unknown long-range interactions such as those among small heterochromatic regions embedded in euchromatic arms. We also found that interactions are correlated with various epigenetic marks that are localized in active or silenced chromatin. Arabidopsis chromosomes do not contain large local interactive domains that resemble the topological domains described in animals, but instead contain relatively small interactive regions scattered around the genome that contain H3K27me3 or H3K9me2. We generated interaction maps in mutants that are defective in specific epigenetic pathways and found altered interaction patterns that correlate with changes in the epigenome. These analyses provide further insights into molecular mechanisms of epigenetic regulation of the genome. PMID:25132175

  13. Characterization of photosynthesis in Arabidopsis ER-to-plastid lipid trafficking mutants.

    PubMed

    Li, Ziru; Gao, Jinpeng; Benning, Christoph; Sharkey, Thomas D

    2012-04-01

    Vascular plants use two pathways to synthesize galactolipids, the predominant lipid species in chloroplasts-a prokaryotic pathway that resides entirely in the chloroplast, and a eukaryotic pathway that involves assembly in the endoplasmic reticulum. Mutants deficient in the endoplasmic reticulum pathway, trigalactosyldiacylglycerol (tgd1-1 and tgd2-1) mutants, had been previously identified with reduced contents of monogalactosyldiacylglycerol and digalactosyldiacylglycerol, and altered lipid molecular species composition. Here, we report that the altered lipid composition affected photosynthesis in lipid trafficking mutants. It was found that proton motive force as measured by electrochromic shift was reduced by ~40% in both tgd mutants. This effect was accompanied by an increase in thylakoid conductance attributable to ATPase activity and so the rate of ATP synthesis was nearly unchanged. Thylakoid conductance to ions also increased in tgd mutants. However, gross carbon assimilation in tgd mutants as measured by gas exchange was only marginally affected. Rubisco activity, electron transport rate, and photosystem I and II oxidation status were not altered. Despite the large differences in proton motive force, responses to heat and high light stress were similar between tgd mutants and the wild type.

  14. The Arabidopsis szl1 mutant reveals a critical role of β-carotene in photosystem I photoprotection.

    PubMed

    Cazzaniga, Stefano; Li, Zhirong; Niyogi, Krishna K; Bassi, Roberto; Dall'Osto, Luca

    2012-08-01

    Carotenes and their oxygenated derivatives, the xanthophylls, are structural determinants in both photosystems (PS) I and II. They bind and stabilize photosynthetic complexes, increase the light-harvesting capacity of chlorophyll-binding proteins, and have a major role in chloroplast photoprotection. Localization of carotenoid species within each PS is highly conserved: Core complexes bind carotenes, whereas peripheral light-harvesting systems bind xanthophylls. The specific functional role of each xanthophyll species has been recently described by genetic dissection, however the in vivo role of carotenes has not been similarly defined. Here, we have analyzed the function of carotenes in photosynthesis and photoprotection, distinct from that of xanthophylls, by characterizing the suppressor of zeaxanthin-less (szl) mutant of Arabidopsis (Arabidopsis thaliana) which, due to the decreased activity of the lycopene-β-cyclase, shows a lower carotene content than wild-type plants. When grown at room temperature, mutant plants showed a lower content in PSI light-harvesting complex I complex than the wild type, and a reduced capacity for chlorophyll fluorescence quenching, the rapidly reversible component of nonphotochemical quenching. When exposed to high light at chilling temperature, szl1 plants showed stronger photoxidation than wild-type plants. Both PSI and PSII from szl1 were similarly depleted in carotenes and yet PSI activity was more sensitive to light stress than PSII as shown by the stronger photoinhibition of PSI and increased rate of singlet oxygen release from isolated PSI light-harvesting complex I complexes of szl1 compared with the wild type. We conclude that carotene depletion in the core complexes impairs photoprotection of both PS under high light at chilling temperature, with PSI being far more affected than PSII.

  15. CRISPR/Cas9-Induced Double-Strand Break Repair in Arabidopsis Nonhomologous End-Joining Mutants

    PubMed Central

    Shen, Hexi; Strunks, Gary D.; Klemann, Bart J. P. M.; Hooykaas, Paul J. J.; de Pater, Sylvia

    2016-01-01

    Double-strand breaks (DSBs) are one of the most harmful DNA lesions. Cells utilize two main pathways for DSB repair: homologous recombination (HR) and nonhomologous end-joining (NHEJ). NHEJ can be subdivided into the KU-dependent classical NHEJ (c-NHEJ) and the more error-prone KU-independent backup-NHEJ (b-NHEJ) pathways, involving the poly (ADP-ribose) polymerases (PARPs). However, in the absence of these factors, cells still seem able to adequately maintain genome integrity, suggesting the presence of other b-NHEJ repair factors or pathways independent from KU and PARPs. The outcome of DSB repair by NHEJ pathways can be investigated by using artificial sequence-specific nucleases such as CRISPR/Cas9 to induce DSBs at a target of interest. Here, we used CRISPR/Cas9 for DSB induction at the Arabidopsis cruciferin 3 (CRU3) and protoporphyrinogen oxidase (PPO) genes. DSB repair outcomes via NHEJ were analyzed using footprint analysis in wild-type plants and plants deficient in key factors of c-NHEJ (ku80), b-NHEJ (parp1 parp2), or both (ku80 parp1 parp2). We found that larger deletions of >20 bp predominated after DSB repair in ku80 and ku80 parp1 parp2 mutants, corroborating with a role of KU in preventing DSB end resection. Deletion lengths did not significantly differ between ku80 and ku80 parp1 parp2 mutants, suggesting that a KU- and PARP-independent b-NHEJ mechanism becomes active in these mutants. Furthermore, microhomologies and templated insertions were observed at the repair junctions in the wild type and all mutants. Since these characteristics are hallmarks of polymerase θ-mediated DSB repair, we suggest a possible role for this recently discovered polymerase in DSB repair in plants. PMID:27866150

  16. Genome stability of Arabidopsis atm, ku80 and rad51b mutants: somatic and transgenerational responses to stress.

    PubMed

    Yao, Youli; Bilichak, Andriy; Titov, Viktor; Golubov, Andrey; Kovalchuk, Igor

    2013-06-01

    DNA double-strand breaks (DSBs) can be repaired via two main mechanisms: non-homologous end joining (NHEJ) and homologous recombination (HR). Our previous work showed that exposure to abiotic stresses resulted in an increase in point mutation frequency (PMF) and homologous recombination frequency (HRF), and these changes were heritable. We hypothesized that mutants impaired in DSB recognition and repair would also be deficient in somatic and transgenerational changes in PMF and HRF. To test this hypothesis, we analyzed the genome stability of the Arabidopsis thaliana mutants deficient in ATM (communication between DNA strand break recognition and the repair machinery), KU80 (deficient in NHEJ) and RAD51B (deficient in HR repair) genes. We found that all three mutants exhibited higher levels of DSBs. Plants impaired in ATM had a lower spontaneous PMF and HRF, whereas ku80 plants had higher frequencies. Plants impaired in RAD51B had a lower HRF. HRF in wild-type, atm and rad51b plants increased in response to several abiotic stressors, whereas it did not increase in ku80 plants. The progeny of stressed wild-type and ku80 plants exhibited an increase in HRF in response to all stresses, and the increase was higher in ku80 plants. The progeny of atm plants showed an increase in HRF only when the parental generation was exposed to cold or flood, whereas the progeny of rad51b plants completely lacked a transgenerational increase in HRF. Our experiments showed that mutants impaired in the recognition and repair of DSBs exhibited changes in the efficiency of DNA repair as reflected by changes in strand breaks, point mutation and HRF. They also showed that the HR RAD51B protein and the protein ATM that recognized damaged DNA might play an important role in transgenerational changes in HRF.

  17. Arabidopsis mutants resistant to the auxin effects of indole-3-acetonitrile are defective in the nitrilase encoded by the NIT1 gene.

    PubMed Central

    Normanly, J; Grisafi, P; Fink, G R; Bartel, B

    1997-01-01

    Indole-3-acetonitrile (IAN) is a candidate precursor of the plant growth hormone indole-3-acetic acid (IAA). We demonstrated that IAN has auxinlike effects on Arabidopsis seedlings and that exogenous IAN is converted to IAA in vivo. We isolated mutants with reduced sensitivity to IAN that remained sensitive to IAA. These mutants were recessive and fell into a single complementation group that mapped to chromosome 3, within 0.5 centimorgans of a cluster of three nitrilase-encoding genes, NIT1, NIT2, and NIT3. Each of the three mutants contained a single base change in the coding region of the NIT1 gene, and the expression pattern of NIT1 is consistent with the IAN insensitivity observed in the nit1 mutant alleles. The half-life of IAN and levels of IAA and IAN were unchanged in the nit1 mutant, confirming that Arabidopsis has other functional nitrilases. Overexpressing NIT2 in transgenic Arabidopsis caused increased sensitivity to IAN and faster turnover of exogenous IAN in vivo. PMID:9368415

  18. An Effective Strategy for Reliably Isolating Heritable and Cas9-Free Arabidopsis Mutants Generated by CRISPR/Cas9-Mediated Genome Editing.

    PubMed

    Gao, Xiuhua; Chen, Jilin; Dai, Xinhua; Zhang, Da; Zhao, Yunde

    2016-07-01

    Mutations generated by CRISPR/Cas9 in Arabidopsis (Arabidopsis thaliana) are often somatic and are rarely heritable. Isolation of mutations in Cas9-free Arabidopsis plants can ensure the stable transmission of the identified mutations to next generations, but the process is laborious and inefficient. Here, we present a simple visual screen for Cas9-free T2 seeds, allowing us to quickly obtain Cas9-free Arabidopsis mutants in the T2 generation. To demonstrate this in principle, we targeted two sites in the AUXIN-BINDING PROTEIN1 (ABP1) gene, whose function as a membrane-associated auxin receptor has been challenged recently. We obtained many T1 plants with detectable mutations near the target sites, but only a small fraction of T1 plants yielded Cas9-free abp1 mutations in the T2 generation. Moreover, the mutations did not segregate in Mendelian fashion in the T2 generation. However, mutations identified in the Cas9-free T2 plants were stably transmitted to the T3 generation following Mendelian genetics. To further simplify the screening procedure, we simultaneously targeted two sites in ABP1 to generate large deletions, which can be easily identified by PCR. We successfully generated two abp1 alleles that contained 1,141- and 711-bp deletions in the ABP1 gene. All of the Cas9-free abp1 alleles we generated were stable and heritable. The method described here allows for effectively isolating Cas9-free heritable CRISPR mutants in Arabidopsis.

  19. Chloroplast localization of methylerythritol 4-phosphate pathway enzymes and regulation of mitochondrial genes in ispD and ispE albino mutants in Arabidopsis.

    PubMed

    Hsieh, Ming-Hsiun; Chang, Chiung-Yun; Hsu, Shih-Jui; Chen, Ju-Jiun

    2008-04-01

    Plant isoprenoids are derived from two independent pathways, the cytosolic mevalonate pathway and the plastid methylerythritol 4-phosphate (MEP) pathway. We used green fluorescent fusion protein assays to demonstrate that the Arabidopsis MEP pathway enzymes are localized to the chloroplast. We have also characterized three Arabidopsis albino mutants, ispD-1, ispD-2 and ispE-1, which have T-DNA insertions in the IspD and IspE genes of the MEP pathway. Levels of photosynthetic pigments are almost undetectable in these albino mutants. Instead of thylakoids, the ispD and ispE mutant chloroplasts are filled with large vesicles. Impairments in chloroplast development and functions may signal changes in the expression of nuclear, chloroplast and mitochondrial genes. We used northern blot analysis to examine the expression of photosynthetic and respiratory genes in the ispD and ispE albino mutants. Steady-state mRNA levels of nucleus- and chloroplast-encoded photosynthetic genes are significantly decreased in the albino mutants. In contrast, transcript levels of nuclear and mitochondrial genes encoding subunits of the mitochondrial electron transport chain are increased or not affected in these mutants. Genomic Southern blot analysis revealed that the DNA amounts of mitochondrial genes are not enhanced in the ispD and ispE albino mutants. These results support the notion that the functional state of chloroplasts may affect the expression of nuclear and mitochondrial genes. The up-regulation of mitochondrial genes in the albino mutants is not caused by changes of mitochondrial DNA copy number in Arabidopsis.

  20. Regulated Ethylene Insensitivity through the Inducible Expression of the Arabidopsis etr1-1 Mutant Ethylene Receptor in Tomato1[OA

    PubMed Central

    Gallie, Daniel R.

    2010-01-01

    Ethylene serves as an important hormone controlling several aspects of plant growth and development, including fruit ripening and leaf and petal senescence. Ethylene is perceived following its binding to membrane-localized receptors, resulting in their inactivation and the induction of ethylene responses. Five distinct types of receptors are expressed in Arabidopsis (Arabidopsis thaliana), and mutant receptors have been described that repress ethylene signaling in a dominant negative manner. One such mutant, ethylene resistant1-1 (etr1-1), results in a strong ethylene-insensitive phenotype in Arabidopsis. In this study, regulated expression of the Arabidopsis etr1-1 in tomato (Solanum lycopersicum) was achieved using an inducible promoter. In the absence of the inducer, transgenic seedlings remained sensitive to ethylene, but in its presence, a state of ethylene insensitivity was induced, resulting in the elongation of the hypocotyl and root in dark-grown seedlings in the presence of ethylene, a reduction or absence of an apical hook, and repression of ethylene-inducible E4 expression. The level of ethylene sensitivity could be controlled by the amount of inducer used, demonstrating a linear relationship between the degree of insensitivity and etr1-1 expression. Induction of etr1-1 expression also repressed the epinastic response to ethylene as well as delayed fruit ripening. Restoration of ethylene sensitivity was achieved following the cessation of the induction. These results demonstrate the ability to control ethylene responses temporally and in amount through the control of mutant receptor expression. PMID:20181754

  1. LHC II protein phosphorylation in leaves of Arabidopsis thaliana mutants deficient in non-photochemical quenching.

    PubMed

    Breitholtz, Hanna-Leena; Srivastava, Renu; Tyystjärvi, Esa; Rintamäki, Eevi

    2005-06-01

    Phosphorylation of the light-harvesting chlorophyll a/b complex II (LHC II) proteins is induced in light via activation of the LHC II kinase by reduction of cytochrome b(6)f complex in thylakoid membranes. We have recently shown that, besides this activation, the LHC II kinase can be regulated in vitro by a thioredoxin-like component, and H2O2 that inserts an inhibitory loop in the regulation of LHC II protein phosphorylation in the chloroplast. In order to disclose the complex network for LHC II protein phosphorylation in vivo, we studied phosphorylation of LHC II proteins in the leaves of npq1-2 and npq4-1 mutants of Arabidopis thaliana. In comparison to wild-type, these mutants showed reduced non-photochemical quenching and increased excitation pressure of Photosystem II (PS II) under physiological light intensities. Peculiar regulation of LHC II protein phosphorylation was observed in mutant leaves under illumination. The npq4-1 mutant was able to maintain a high amount of phosphorylated LHC II proteins in thylakoid membranes at light intensities that induced inhibition of phosphorylation in wild-type leaves. Light intensity-dependent changes in the level of LHC II protein phosphorylation were smaller in the npq1-2 mutant compared to the wild-type. No significant differences in leaf thickness, dry weight, chlorophyll content, or the amount of LHC II proteins were observed between the two mutant and wild-type lines. We propose that the reduced capacity of the mutant lines to dissipate excess excitation energy induces changes in the production of reactive oxygen species in chloroplasts, which consequently affects the regulation of LHC II protein phosphorylation.

  2. Evidence for a Ustilago maydis steroid 5alpha-reductase by functional expression in Arabidopsis det2-1 mutants.

    PubMed

    Basse, Christoph W; Kerschbamer, Christine; Brustmann, Markus; Altmann, Thomas; Kahmann, Regine

    2002-06-01

    We have identified a gene (udh1) in the basidiomycete Ustilago maydis that is induced during the parasitic interaction with its host plant maize (Zea mays). udh1 encodes a protein with high similarity to mammalian and plant 5alpha-steroid reductases. Udh1 differs from those of known 5alpha-steroid reductases by six additional domains, partially predicted to be membrane-spanning. A fusion protein of Udh1 and the green fluorescent protein provided evidence for endoplasmic reticulum localization in U. maydis. The function of the Udh1 protein was demonstrated by complementing Arabidopsis det2-1 mutants, which display a dwarf phenotype due to a mutation in the 5alpha-steroid reductase encoding DET2 gene. det2-1 mutant plants expressing either the udh1 or the DET2 gene controlled by the cauliflower mosaic virus 35S promoter differed from wild-type Columbia plants by accelerated stem growth, flower and seed development and a reduction in size and number of rosette leaves. The accelerated growth phenotype of udh1 transgenic plants was stably inherited and was favored under reduced light conditions. Truncation of the N-terminal 70 amino acids of the Udh1 protein abolished the ability to restore growth in det2-1 plants. Our results demonstrate the existence of a 5alpha-steroid reductase encoding gene in fungi and suggest a common ancestor between fungal, plant, and mammalian proteins.

  3. Genome-wide analysis of mutations in mutant lineages selected following fast-neutron irradiation mutagenesis of Arabidopsis thaliana

    PubMed Central

    Belfield, Eric J.; Gan, Xiangchao; Mithani, Aziz; Brown, Carly; Jiang, Caifu; Franklin, Keara; Alvey, Elizabeth; Wibowo, Anjar; Jung, Marko; Bailey, Kit; Kalwani, Sharan; Ragoussis, Jiannis; Mott, Richard; Harberd, Nicholas P.

    2012-01-01

    Ionizing radiation has long been known to induce heritable mutagenic change in DNA sequence. However, the genome-wide effect of radiation is not well understood. Here we report the molecular properties and frequency of mutations in phenotypically selected mutant lines isolated following exposure of the genetic model flowering plant Arabidopsis thaliana to fast neutrons (FNs). Previous studies suggested that FNs predominantly induce deletions longer than a kilobase in A. thaliana. However, we found a higher frequency of single base substitution than deletion mutations. While the overall frequency and molecular spectrum of fast-neutron (FN)–induced single base substitutions differed substantially from those of “background” mutations arising spontaneously in laboratory-grown plants, G:C>A:T transitions were favored in both. We found that FN-induced G:C>A:T transitions were concentrated at pyrimidine dinucleotide sites, suggesting that FNs promote the formation of mutational covalent linkages between adjacent pyrimidine residues. In addition, we found that FNs induced more single base than large deletions, and that these single base deletions were possibly caused by replication slippage. Our observations provide an initial picture of the genome-wide molecular profile of mutations induced in A. thaliana by FN irradiation and are particularly informative of the nature and extent of genome-wide mutation in lines selected on the basis of mutant phenotypes from FN-mutagenized A. thaliana populations. PMID:22499668

  4. Elemental Concentrations in the Seed of Mutants and Natural Variants of Arabidopsis thaliana Grown under Varying Soil Conditions

    PubMed Central

    McDowell, Stephen C.; Akmakjian, Garo; Sladek, Chris; Mendoza-Cozatl, David; Morrissey, Joe B.; Saini, Nick; Mittler, Ron; Baxter, Ivan; Salt, David E.; Ward, John M.; Schroeder, Julian I.; Guerinot, Mary Lou; Harper, Jeffrey F.

    2013-01-01

    The concentrations of mineral nutrients in seeds are critical to both the life cycle of plants as well as human nutrition. These concentrations are strongly influenced by soil conditions, as shown here by quantifying the concentration of 14 elements in seeds from Arabidopsis thaliana plants grown under four different soil conditions: standard, or modified with NaCl, heavy metals, or alkali. Each of the modified soils resulted in a unique change to the seed ionome (the mineral nutrient content of the seeds). To help identify the genetic networks regulating the seed ionome, changes in elemental concentrations were evaluated using mutants corresponding to 760 genes as well as 10 naturally occurring accessions. The frequency of ionomic phenotypes supports an estimate that as much as 11% of the A. thaliana genome encodes proteins of functional relevance to ion homeostasis in seeds. A subset of mutants were analyzed with two independent alleles, providing five examples of genes important for regulation of the seed ionome: SOS2, ABH1, CCC, At3g14280 and CNGC2. In a comparison of nine different accessions to a Col-0 reference, eight accessions were observed to have reproducible differences in elemental concentrations, seven of which were dependent on specific soil conditions. These results indicate that the A. thaliana seed ionome is distinct from the vegetative ionome, and that elemental analysis is a sensitive approach to identify genes controlling ion homeostasis, including those that regulate gene expression, phospho-regulation, and ion transport. PMID:23671651

  5. Elevated Levels of High-Melting-Point Phosphatidylglycerols Do Not Induce Chilling Sensitivity in an Arabidopsis Mutant.

    PubMed Central

    Wu, J.; Browse, J.

    1995-01-01

    Molecular species of phosphatidylglycerol that contain only 16:0, 18:0, and 16:1-trans fatty acids undergo the transition from liquid crystalline phase to gel phase at temperatures well above 20[deg]C. Several lines of evidence have been used to implicate elevated proportions of these high-melting-point molecular species as a major cause of plant chilling sensitivity. In the fatty acid biosynthesis 1 (fab1) mutant of Arabidopsis, leaf phosphatidylglycerol contained 43% high-melting-point molecular species[mdash]a higher percentage than is found in many chilling-sensitive plants. Nevertheless, the mutant was completely unaffected (when compared with wild-type controls) by a range of low-temperature treatments that quickly led to the death of cucumber and other chilling-sensitive plants. Our results clearly demonstrate that high-melting-point phosphatidylglycerols do not mediate classic chilling damage. However, growth of fab1 plants was compromised by long-term (>2 weeks) exposure to 2[deg]C. This finding and other observations are consistent with a proposition that plants native to tropical and subtropical regions have evolved many traits that are incompatible with long-term growth or development in cooler climates but that may confer selective advantages at high temperatures. PMID:12242349

  6. Transmission electron microscopy and serial reconstructions reveal novel meiotic phenotypes for the ahp2 mutant of Arabidopsis thaliana.

    PubMed

    Pathan, Nazia; Stronghill, Patti; Hasenkampf, Clare

    2013-03-01

    We have found novel phenotypes for the previously studied Arabidopsis thaliana (L.) Heynh. meiotic mutant ahp2. These phenotypes were revealed by analysis of reconstructions of normal and ahp2 nuclei that were imaged using transmission electron microscopy. Previous studies of the ahp2 mutant demonstrated that it has a general failure to form synaptonemal complexes, except for the nucleolus organizing regions, and it fails to complete reciprocal genetic exchange. Here, we show that even though the ahp2 chromosome axes have only 5% of the normal amount of synaptonemal complex formation, it nonetheless has slightly more than 40% of the axes involved in close alignment. We also observed two striking nuclear envelope associated abnormalities. Wild type nuclei contain two nucleoli, one nucleolus-like structure, and nuclear envelope associated structures that we refer to as nuclear envelope associated disks. The ahp2 nuclei have the two nucleoli, but they lack the third nucleolus-like structure and instead have a previously uncharacterized structure that spans the nuclear envelope. Additionally, ahp2 meiocytes have nuclear envelope associated disks that are narrower and more numerous (∼2×) than those seen in wild type, and unlike the wild type disks, they are in direct contact with the nuclear envelope.

  7. Opposite variations in fumarate and malate dominate metabolic phenotypes of Arabidopsis salicylate mutants with abnormal biomass under chilling.

    PubMed

    Scott, Ian M; Ward, Jane L; Miller, Sonia J; Beale, Michael H

    2014-12-01

    In chilling conditions (5°C), salicylic acid (SA)-deficient mutants (sid2, eds5 and NahG) of Arabidopsis thaliana produced more biomass than wild type (Col-0), whereas the SA overproducer cpr1 was extremely stunted. The hypothesis that these phenotypes were reflected in metabolism was explored using 600 MHz (1) H nuclear magnetic resonance (NMR) analysis of unfractionated polar shoot extracts. Biomass-related metabolic phenotypes were identified as multivariate data models of these NMR 'fingerprints'. These included principal components that correlated with biomass. Also, partial least squares-regression models were found to predict the relative size of plants in previously unseen experiments in different light intensities, or relative size of one genotype from the others. The dominant signal in these models was fumarate, which was high in SA-deficient mutants, intermediate in Col-0 and low in cpr1 at 5°C. Among signals negatively correlated with biomass, malate was prominent. Abundance of transcripts of the FUM2 cytosolic fumarase (At5g50950) showed strong positive correlation with fumarate levels and with biomass, whereas no significant differences were found for the FUM1 mitochondrial fumarase (At2g47510). It was confirmed that the morphological effects of SA under chilling find expression in the metabolome, with a role of fumarate highlighted.

  8. Reduced immunogenicity of Arabidopsis hgl1 mutant N-glycans caused by altered accessibility of xylose and core fucose epitopes.

    PubMed

    Kaulfürst-Soboll, Heidi; Rips, Stephan; Koiwa, Hisashi; Kajiura, Hiroyuki; Fujiyama, Kazuhito; von Schaewen, Antje

    2011-07-01

    Arabidopsis N-glycosylation mutants with enhanced salt sensitivity show reduced immunoreactivity of complex N-glycans. Among them, hybrid glycosylation 1 (hgl1) alleles lacking Golgi α-mannosidase II are unique, because their glycoprotein N-glycans are hardly labeled by anti-complex glycan antibodies, even though they carry β1,2-xylose and α1,3-fucose epitopes. To dissect the contribution of xylose and core fucose residues to plant stress responses and immunogenic potential, we prepared Arabidopsis hgl1 xylT double and hgl1 fucTa fucTb triple mutants by crossing previously established T-DNA insertion lines and verified them by mass spectrometry analyses. Root growth assays revealed that hgl1 fucTa fucTb but not hgl1 xylT plants are more salt-sensitive than hgl1, hinting at the importance of core fucose modification and masking of xylose residues. Detailed immunoblot analyses with anti-β1,2-xylose and anti-α1,3-fucose rabbit immunoglobulin G antibodies as well as cross-reactive carbohydrate determinant-specific human immunoglobulin E antibodies (present in sera of allergy patients) showed that xylose-specific reactivity of hgl1 N-glycans is indeed reduced. Based on three-dimensional modeling of plant N-glycans, we propose that xylose residues are tilted by 30° because of untrimmed mannoses in hgl1 mutants. Glycosidase treatments of protein extracts restored immunoreactivity of hgl1 N-glycans supporting these models. Furthermore, among allergy patient sera, untrimmed mannoses persisting on the α1,6-arm of hgl1 N-glycans were inhibitory to immunoreaction with core fucoses to various degrees. In summary, incompletely trimmed glycoprotein N-glycans conformationally prevent xylose and, to lesser extent, core fucose accessibility. Thus, in addition to N-acetylglucosaminyltransferase I, Golgi α-mannosidase II emerges as a so far unrecognized target for lowering the immunogenic potential of plant-derived glycoproteins.

  9. Analysis of multiple photoreceptor pigments for phototropism in a mutant of Arabidopsis thaliana

    NASA Technical Reports Server (NTRS)

    Konjevic, R.; Khurana, J. P.; Poff, K. L.

    1992-01-01

    The shape of the fluence-response relationship for the phototropic response of the JK224 strain of Arabidopsis thaliana depends on the fluence rate and wavelength of the actinic light. At low fluence rate (0.1 micromole m-2 s-1), the response to 450-nm light is characterized by a single maximum at about 9 micromoles m-2. At higher fluence rate (0.4 micromole m-2 s-1), the response shows two maxima, at 4.5 and 9 micromoles m-2. The response to 510-nm light shows a single maximum at 4.5 micromoles m-2. Unilateral preirradiation with high fluence rate (25 micromoles m-2 s-1) 510-nm light eliminates the maximum at 4.5 micromoles m-2 in the fluence response curve to a subsequent unilateral 450-nm irradiation, while the second maximum at 9 micromoles m-2 is unaffected. Based on these results, it is concluded that a single photoreceptor pigment has been altered in the JK224 strain of Arabidopsis thaliana.

  10. Antioxidant status, peroxidase activity, and PR protein transcript levels in ascorbate-deficient Arabidopsis thaliana vtc mutants.

    PubMed

    Colville, Louise; Smirnoff, Nicholas

    2008-01-01

    Ascorbate is the most abundant small molecule antioxidant in plants and is proposed to function, along with other members of an antioxidant network, in controlling reactive oxygen species. A biochemical and molecular characterization of four ascorbate-deficient (vtc) Arabidopsis thaliana mutants has been carried out to determine if ascorbate deficiency is compensated by changes in the other major antioxidants. Seedlings grown in vitro were used to minimize stress and longer term developmental differences. Comparison was made with the low glutathione cad2 mutant and vtc2-1 treated with D,L-buthionine-[S,R]-sulphoximine to cause combined ascorbate and glutathione deficiency. The pool sizes and oxidation state of ascorbate and glutathione were not altered by deficiency of the other. alpha-Tocopherol and activities of monodehydroascorbate reductase, dehydroascorbate reductase, glutathione reductase, and catalase were little affected. Ascorbate peroxidase activity was higher in vtc1, vtc2-1, and vtc2-2. Ionically bound cell wall peroxidase activity was increased in vtc1, vtc2-1, and vtc4. Supplementation with ascorbate increased cell wall peroxidase activity. 2,6-Dichlorobenzonitrile, an inhibitor of cellulose synthesis, increased cell wall peroxidase activity in the wild type and vtc1. The transcript level of an endochitinase, PR1, and PR2, but not GST6, was increased in vtc1, vtc2-1, and vtc-2-2. Endochitinase transcript levels increased after ascorbate, paraquat, salicylic acid, and UV-C treatment, PR1 after salicylic acid treatment, and PR2 after paraquat and UV-C treatment. Camalexin was higher in vtc1 and the vtc2 alleles. Induction of PR genes, cell wall peroxidase activity, and camalexin in vtc1, vtc2-1, and vtc2-2 suggests that the mutants are affected in pathogen response signalling pathways.

  11. Photosystem II Repair and Plant Immunity: Lessons Learned from Arabidopsis Mutant Lacking the THYLAKOID LUMEN PROTEIN 18.3

    PubMed Central

    Järvi, Sari; Isojärvi, Janne; Kangasjärvi, Saijaliisa; Salojärvi, Jarkko; Mamedov, Fikret; Suorsa, Marjaana; Aro, Eva-Mari

    2016-01-01

    Chloroplasts play an important role in the cellular sensing of abiotic and biotic stress. Signals originating from photosynthetic light reactions, in the form of redox and pH changes, accumulation of reactive oxygen and electrophile species or stromal metabolites are of key importance in chloroplast retrograde signaling. These signals initiate plant acclimation responses to both abiotic and biotic stresses. To reveal the molecular responses activated by rapid fluctuations in growth light intensity, gene expression analysis was performed with Arabidopsis thaliana wild type and the tlp18.3 mutant plants, the latter showing a stunted growth phenotype under fluctuating light conditions (Biochem. J, 406, 415–425). Expression pattern of genes encoding components of the photosynthetic electron transfer chain did not differ between fluctuating and constant light conditions, neither in wild type nor in tlp18.3 plants, and the composition of the thylakoid membrane protein complexes likewise remained unchanged. Nevertheless, the fluctuating light conditions repressed in wild-type plants a broad spectrum of genes involved in immune responses, which likely resulted from shade-avoidance responses and their intermixing with hormonal signaling. On the contrary, in the tlp18.3 mutant plants there was an imperfect repression of defense-related transcripts upon growth under fluctuating light, possibly by signals originating from minor malfunction of the photosystem II (PSII) repair cycle, which directly or indirectly modulated the transcript abundances of genes related to light perception via phytochromes. Consequently, a strong allocation of resources to defense reactions in the tlp18.3 mutant plants presumably results in the stunted growth phenotype under fluctuating light. PMID:27064270

  12. SHORT-ROOT Deficiency Alleviates the Cell Death Phenotype of the Arabidopsis catalase2 Mutant under Photorespiration-Promoting Conditions.

    PubMed

    Waszczak, Cezary; Kerchev, Pavel I; Mühlenbock, Per; Hoeberichts, Frank A; Van Der Kelen, Katrien; Mhamdi, Amna; Willems, Patrick; Denecker, Jordi; Kumpf, Robert P; Noctor, Graham; Messens, Joris; Van Breusegem, Frank

    2016-08-01

    Hydrogen peroxide (H2O2) can act as a signaling molecule that influences various aspects of plant growth and development, including stress signaling and cell death. To analyze molecular mechanisms that regulate the response to increased H2O2 levels in plant cells, we focused on the photorespiration-dependent peroxisomal H2O2 production in Arabidopsis thaliana mutants lacking CATALASE2 (CAT2) activity (cat2-2). By screening for second-site mutations that attenuate the PSII maximum efficiency (Fv'/Fm') decrease and lesion formation linked to the cat2-2 phenotype, we discovered that a mutation in SHORT-ROOT (SHR) rescued the cell death phenotype of cat2-2 plants under photorespiration-promoting conditions. SHR deficiency attenuated H2O2-dependent gene expression, oxidation of the glutathione pool, and ascorbate depletion in a cat2-2 genetic background upon exposure to photorespiratory stress. Decreased glycolate oxidase and catalase activities together with accumulation of glycolate further implied that SHR deficiency impacts the cellular redox homeostasis by limiting peroxisomal H2O2 production. The photorespiratory phenotype of cat2-2 mutants did not depend on the SHR functional interactor SCARECROW and the sugar signaling component ABSCISIC ACID INSENSITIVE4, despite the requirement for exogenous sucrose for cell death attenuation in cat2-2 shr-6 double mutants. Our findings reveal a link between SHR and photorespiratory H2O2 production that has implications for the integration of developmental and stress responses.

  13. Interaction of glucosinolate content of Arabidopsis thaliana mutant lines and feeding and oviposition by generalist and specialist lepidopterans.

    PubMed

    Badenes-Perez, Francisco R; Reichelt, Michael; Gershenzon, Jonathan; Heckel, David G

    2013-02-01

    The diamondback moth, Plutella xylostella L. (Lepidoptera: Plutellidae), is an insect specialized on glucosinolate-containing Brassicaceae that uses glucosinolates in host-plant recognition. We used wild-type and mutants of Arabidopsis thaliana (L.) Heynh. (Brassicaceae) to investigate the interaction between plant glucosinolate and myrosinase content and herbivory by larvae of the generalist Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) and the specialist P. xylostella. We also measured glucosinolate changes as a result of herbivory by these larvae to investigate whether herbivory and glucosinolate induction had an effect on oviposition preference by P. xylostella. Feeding by H. armigera and P. xylostella larvae was 2.1 and 2.5 times less, respectively, on apk1 apk2 plants (with almost no aliphatic glucosinolates) than on wild-type plants. However, there were no differences in feeding by H. armigera and P. xylostella larvae on wild-type, gsm1 (different concentrations of aliphatic glucosinolates compared to wild-type plants), and tgg1 tgg2 plants (lacking major myrosinases). Glucosinolate induction (up to twofold) as a result of herbivory occurred in some cases, depending on both the plant line and the herbivore. For H. armigera, induction, when observed, was noted mostly for indolic glucosinolates, while for P. xylostella, induction was observed in both aliphatic and indolic glucosinolates, but not in all plant lines. For H. armigera, glucosinolate induction, when observed, resulted in an increase of glucosinolate content, while for P. xylostella, induction resulted in both a decrease and an increase in glucosinolate content. Two-choice tests with wild-type and mutant plants were conducted with larvae and ovipositing moths. There were no significant differences in preference of larvae and ovipositing moths between wild-type and gsm1 mutants and between wild-type and tgg1 tgg2 mutants. However, both larvae and ovipositing moths preferred wild-type over apk

  14. Parallel analysis of Arabidopsis circadian clock mutants reveals different scales of transcriptome and proteome regulation

    PubMed Central

    Graf, Alexander; Coman, Diana; Walsh, Sean; Flis, Anna; Stitt, Mark; Gruissem, Wilhelm

    2017-01-01

    The circadian clock regulates physiological processes central to growth and survival. To date, most plant circadian clock studies have relied on diurnal transcriptome changes to elucidate molecular connections between the circadian clock and observable phenotypes in wild-type plants. Here, we have integrated RNA-sequencing and protein mass spectrometry data to comparatively analyse the lhycca1, prr7prr9, gi and toc1 circadian clock mutant rosette at the end of day and end of night. Each mutant affects specific sets of genes and proteins, suggesting that the circadian clock regulation is modular. Furthermore, each circadian clock mutant maintains its own dynamically fluctuating transcriptome and proteome profile specific to subcellular compartments. Most of the measured protein levels do not correlate with changes in their corresponding transcripts. Transcripts and proteins that have coordinated changes in abundance are enriched for carbohydrate- and cold-responsive genes. Transcriptome changes in all four circadian clock mutants also affect genes encoding starch degradation enzymes, transcription factors and protein kinases. The comprehensive transcriptome and proteome datasets demonstrate that future system-driven research of the circadian clock requires multi-level experimental approaches. Our work also shows that further work is needed to elucidate the roles of post-translational modifications and protein degradation in the regulation of clock-related processes. PMID:28250106

  15. Proteome readjustments in the apoplastic space of Arabidopsis thaliana ggt1 mutant leaves exposed to UV-B radiation

    PubMed Central

    Trentin, Anna Rita; Pivato, Micaela; Mehdi, Syed M. M.; Barnabas, Leonard Ebinezer; Giaretta, Sabrina; Fabrega-Prats, Marta; Prasad, Dinesh; Arrigoni, Giorgio; Masi, Antonio

    2015-01-01

    Ultraviolet-B radiation acts as an environmental stimulus, but in high doses it has detrimental effects on plant metabolism. Plasma membranes represent a major target for Reactive Oxygen Species (ROS) generated by this harmful radiation. Oxidative reactions occurring in the apoplastic space are counteracted by antioxidative systems mainly involving ascorbate and, to some extent, glutathione. The occurrence of the latter and its exact role in the extracellular space are not well documented, however. In Arabidopsis thaliana, the gamma-glutamyl transferase isoform (GGT1) bound to the cell wall takes part in the so-called gamma-glutamyl cycle for extracellular glutathione degradation and recovery, and may be implicated in redox sensing and balance. In this work, oxidative conditions were imposed with Ultraviolet-B radiation (UV-B) and studied in redox altered ggt1 mutants. The response of ggt1 knockout Arabidopsis leaves to UV-B radiation was assessed by investigating changes in extracellular glutathione and ascorbate content and their redox state, and in apoplastic protein composition. Our results show that, on UV-B exposure, soluble antioxidants respond to the oxidative conditions in both genotypes. Rearrangements occur in their apoplastic protein composition, suggesting an involvement of Hydrogen Peroxide (H2O2), which may ultimately act as a signal. Other important changes relating to hormonal effects, cell wall remodeling, and redox activities are discussed. We argue that oxidative stress conditions imposed by UV-B and disruption of the gamma-glutamyl cycle result in similar stress-induced responses, to some degree at least. Data are available via ProteomeXchange with identifier PXD001807. PMID:25852701

  16. Analysis of tobamovirus multiplication in Arabidopsis thaliana mutants defective in TOM2A homologues.

    PubMed

    Fujisaki, Koki; Kobayashi, Soko; Tsujimoto, Yayoi; Naito, Satoshi; Ishikawa, Masayuki

    2008-06-01

    The TOM2A gene of Arabidopsis thaliana encodes a four-pass transmembrane protein that is required for efficient multiplication of a tobamovirus, TMV-Cg. In this study, the involvement of three TOM2A homologues in tobamovirus multiplication in A. thaliana was examined. T-DNA insertion mutations in the three homologues, separately or in combination, did not affect TMV-Cg multiplication, whereas, in the tom2a genetic background, some combinations reduced it. This result suggests that the TOM2A homologues are functional in enhancing TMV-Cg multiplication, but their contribution is much less than TOM2A. Interestingly, the multiplication of another tobamovirus, Tomato mosaic virus, was not drastically affected by any combinations of the mutations in TOM2A and its homologues as far as we examined.

  17. Mutants of arabidopsis thaliana that exhibit chlorosis in an atmosphere enriched with CO sub 2

    SciTech Connect

    Artus, N.N ); Somerville, C. )

    1989-04-01

    Two nonallelic, nuclear recessive mutations have been isolated which cause chlorosis of plants grown in an atmosphere enriched to 2% CO{sub 2}. For one of the mutant lines, chlorosis begins at the veins and gradually spreads to the interveinal regions. A minimum light intensity of ca. 50 {mu}E m{sup {minus}2} s{sup {minus}1} was required for this response. For the other mutant line, the yellowing is independent of the light intensity and begins at the basal regions of the leaves and spreads to the tips. Neither line became chlorotic in a low O{sub 2} atmosphere which suppressed photorespiration as effectively as 2% CO{sub 2}. Thus, the mutations do not impose a requirement for photorespiration. Root elongation and callus growth were not affected by high CO{sub 2} for either mutant line. The possibilities that the high CO{sub 2}-sensitive phenotypes are caused by an effect of CO{sub 2} on stomata or ethylene synthesis have also been ruled out.

  18. Mutants of circadian-associated PRR genes display a novel and visible phenotype as to light responses during de-etiolation of Arabidopsis thaliana seedlings.

    PubMed

    Kato, Takahiko; Murakami, Masaya; Nakamura, Yuko; Ito, Shogo; Nakamichi, Norihito; Yamashino, Takafumi; Mizuno, Takeshi

    2007-03-01

    In Arabidopsis thaliana, it is currently accepted that certain mutants with lesions in clock-associated genes commonly display hallmarked phenotypes with regard to three characteristic biological events: (i) altered rhythmic expression of circadian-controlled genes, (ii) changes in flowering time, and (iii) altered sensitivity to red light in elongation of hypocotyls. During the course of examination of the clock-associated mutants of PSEUDO-RESPONSE REGULATORS, PRRs, including TOC1 (PRR1), we found that they commonly show another visible phenotype of anomalous greening responses upon the onset to light exposure of etiolated seedlings. These findings are indicative of a novel link between circadian rhythms and chloroplast development.

  19. Analysis of a sugar response mutant of Arabidopsis identified a novel B3 domain protein that functions as an active transcriptional repressor.

    PubMed

    Tsukagoshi, Hironaka; Saijo, Takanori; Shibata, Daisuke; Morikami, Atsushi; Nakamura, Kenzo

    2005-06-01

    A recessive mutation hsi2 of Arabidopsis (Arabidopsis thaliana) expressing luciferase (LUC) under control of a short promoter derived from a sweet potato (Ipomoea batatas) sporamin gene (Spo(min)LUC) caused enhanced LUC expression under both low- and high-sugar conditions, which was not due to increased level of abscisic acid. The hsi2 mutant contained a nonsense mutation in a gene encoding a protein with B3 DNA-binding domain. HSI2 and two other Arabidopsis proteins appear to constitute a novel subfamily of B3 domain proteins distinct from ABI3, FUS3, and LEC2, which are transcription activators involved in seed development. The C-terminal part of HSI2 subfamily proteins contained a sequence similar to the ERF-associated amphiphilic repression (EAR) motif. Deletion of the C-terminal portion of HSI2 lost in the hsi2 mutant caused reduced nuclear targeting of HSI2. Null allele of HSI2 showed even higher Spo(min)LUC expression than the hsi2 mutant, whereas overexpression of HSI2 reduced the LUC expression. Transient coexpression of 35SHSI2 with Spo(min)LUC in protoplasts repressed the expression of LUC activity, and deletion or mutation of the EAR motif significantly reduced the repression activity of HSI2. These results indicate that HSI2 and related proteins are B3 domain-EAR motif active transcription repressors.

  20. The dominance of the herbicide resistance cost in several Arabidopsis thaliana mutant lines.

    PubMed Central

    Roux, Fabrice; Gasquez, Jacques; Reboud, Xavier

    2004-01-01

    Resistance evolution depends upon the balance between advantage and disadvantage (cost) conferred in treated and untreated areas. By analyzing morphological characters and simple fitness components, the cost associated with each of eight herbicide resistance alleles (acetolactate synthase, cellulose synthase, and auxin-induced target genes) was studied in the model plant Arabidopsis thaliana. The use of allele-specific PCR to discriminate between heterozygous and homozygous plants was used to provide insights into the dominance of the resistance cost, a parameter rarely described. Morphological characters appear more sensitive than fitness (seed production) because 6 vs. 4 differences between resistant and sensitive homozygous plants were detected, respectively. Dominance levels for the fitness cost ranged from recessivity (csr1-1, ixr1-2, and axr1-3) to dominance (axr2-1) to underdominance (aux1-7). Furthermore, the dominance level of the herbicide resistance trait did not predict the dominance level of the cost of resistance. The relationship of our results to theoretical predictions of dominance and the consequences of fitness cost and its dominance in resistance management are discussed. PMID:15020435

  1. Improved growth and stress tolerance in the Arabidopsis oxt1 mutant triggered by altered adenine metabolism.

    PubMed

    Sukrong, Suchada; Yun, Kil-Young; Stadler, Patrizia; Kumar, Charan; Facciuolo, Tony; Moffatt, Barbara A; Falcone, Deane L

    2012-11-01

    Plants perceive and respond to environmental stresses with complex mechanisms that are often associated with the activation of antioxidant defenses. A genetic screen aimed at isolating oxidative stress-tolerant lines of Arabidopsis thaliana has identified oxt1, a line that exhibits improved tolerance to oxidative stress and elevated temperature but displays no apparent deleterious growth effects under non-stress conditions. Oxt1 harbors a mutation that arises from the altered expression of a gene encoding adenine phosphoribosyltransferase (APT1), an enzyme that converts adenine to adenosine monophosphate (AMP), indicating a link between purine metabolism, whole-plant growth responses, and stress acclimation. The oxt1 mutation results in decreased APT1 expression that leads to reduced enzymatic activity. Correspondingly, oxt1 plants possess elevated levels of adenine. Decreased APT enzyme activity directly correlates with stress resistance in transgenic lines that ectopically express APT1. The metabolic alteration in oxt1 plants also alters the expression of several antioxidant defense genes and the response of these genes to oxidative challenge. Finally, it is shown that manipulation of adenine levels can induce stress tolerance to wild-type plants. Collectively, these results show that alterations in cellular adenine levels can trigger stress tolerance and improve growth, leading to increases in plant biomass. The results also suggest that adenine might play a part in the signals that modulate responses to abiotic stress and plant growth.

  2. Strong morphological defects in conditional Arabidopsis abp1 knock-down mutants generated in absence of functional ABP1 protein

    PubMed Central

    Perrot-Rechenmann, Catherine; Friml, Jiří

    2016-01-01

    The Auxin Binding Protein 1 (ABP1) is one of the most studied proteins in plants. Since decades ago, it has been the prime receptor candidate for the plant hormone auxin with a plethora of described functions in auxin signaling and development. The developmental importance of ABP1 has recently been questioned by identification of Arabidopsis thaliana abp1 knock-out alleles that show no obvious phenotypes under normal growth conditions. In this study, we examined the contradiction between the normal growth and development of the abp1 knock-outs and the strong morphological defects observed in three different ethanol-inducible abp1 knock-down mutants ( abp1-AS, SS12K, SS12S). By analyzing segregating populations of abp1 knock-out vs. abp1 knock-down crosses we show that the strong morphological defects that were believed to be the result of conditional down-regulation of ABP1 can be reproduced also in the absence of the functional ABP1 protein. This data suggests that the phenotypes in  abp1 knock-down lines are due to the off-target effects and asks for further reflections on the biological function of ABP1 or alternative explanations for the missing phenotypic defects in the abp1 loss-of-function alleles. PMID:26925228

  3. Transcriptome analysis of Arabidopsis GCR1 mutant reveals its roles in stress, hormones, secondary metabolism and phosphate starvation.

    PubMed

    Chakraborty, Navjyoti; Sharma, Priyanka; Kanyuka, Kostya; Pathak, Ravi R; Choudhury, Devapriya; Hooley, Richard A; Raghuram, Nandula

    2015-01-01

    The controversy over the existence or the need for G-protein coupled receptors (GPCRs) in plant G-protein signalling has overshadowed a more fundamental quest for the role of AtGCR1, the most studied and often considered the best candidate for GPCR in plants. Our whole transcriptome microarray analysis of the GCR1-knock-out mutant (gcr1-5) in Arabidopsis thaliana revealed 350 differentially expressed genes spanning all chromosomes. Many of them were hitherto unknown in the context of GCR1 or G-protein signalling, such as in phosphate starvation, storage compound and fatty acid biosynthesis, cell fate, etc. We also found some GCR1-responsive genes/processes that are reported to be regulated by heterotrimeric G-proteins, such as biotic and abiotic stress, hormone response and secondary metabolism. Thus, GCR1 could have G-protein-mediated as well as independent roles and regardless of whether it works as a GPCR, further analysis of the organism-wide role of GCR1 has a significance of its own.

  4. The absence of chlorophyll b affects lateral mobility of photosynthetic complexes and lipids in grana membranes of Arabidopsis and barley chlorina mutants.

    PubMed

    Tyutereva, Elena V; Evkaikina, Anastasiia I; Ivanova, Alexandra N; Voitsekhovskaja, Olga V

    2017-04-05

    The lateral mobility of integral components of thylakoid membranes, such as plastoquinone, xanthophylls, and pigment-protein complexes, is critical for the maintenance of efficient light harvesting, high rates of linear electron transport, and successful repair of damaged photosystem II (PSII). The packaging of the photosynthetic pigment-protein complexes in the membrane depends on their size and stereometric parameters which in turn depend on the composition of the complexes. Chlorophyll b (Chlb) is an important regulator of antenna size and composition. In this study, the lateral mobility (the mobile fraction size) of pigment-protein complexes and lipids in grana membranes was analyzed in chlorina mutants of Arabidopsis and barley lacking Chlb. In the Arabidopsis ch1-3 mutant, diffusion of membrane lipids decreased as compared to wild-type plants, but the diffusion of photosynthetic complexes was not affected. In the barley chlorina f2 3613 mutant, the diffusion of pigment-protein complexes significantly decreased, while the diffusion of lipids increased, as compared to wild-type plants. We propose that the size of the mobile fractions of pigment-protein complexes in grana membranes in vivo is higher than reported previously. The data are discussed in the context of the protein composition of antennae, characteristics of the plastoquinone pool, and production of reactive oxygen species in leaves of chlorina mutants.

  5. Molecular Phenotyping of the pal1 and pal2 Mutants of Arabidopsis thaliana Reveals Far-Reaching Consequences on Phenylpropanoid, Amino Acid, and Carbohydrate Metabolism

    PubMed Central

    Rohde, Antje; Morreel, Kris; Ralph, John; Goeminne, Geert; Hostyn, Vanessa; De Rycke, Riet; Kushnir, Sergej; Van Doorsselaere, Jan; Joseleau, Jean-Paul; Vuylsteke, Marnik; Van Driessche, Gonzalez; Van Beeumen, Jozef; Messens, Eric; Boerjan, Wout

    2004-01-01

    The first enzyme of the phenylpropanoid pathway, Phe ammonia-lyase (PAL), is encoded by four genes in Arabidopsis thaliana. Whereas PAL function is well established in various plants, an insight into the functional significance of individual gene family members is lacking. We show that in the absence of clear phenotypic alterations in the Arabidopsis pal1 and pal2 single mutants and with limited phenotypic alterations in the pal1 pal2 double mutant, significant modifications occur in the transcriptome and metabolome of the pal mutants. The disruption of PAL led to transcriptomic adaptation of components of the phenylpropanoid biosynthesis, carbohydrate metabolism, and amino acid metabolism, revealing complex interactions at the level of gene expression between these pathways. Corresponding biochemical changes included a decrease in the three major flavonol glycosides, glycosylated vanillic acid, scopolin, and two novel feruloyl malates coupled to coniferyl alcohol. Moreover, Phe overaccumulated in the double mutant, and the levels of many other amino acids were significantly imbalanced. The lignin content was significantly reduced, and the syringyl/guaiacyl ratio of lignin monomers had increased. Together, from the molecular phenotype, common and specific functions of PAL1 and PAL2 are delineated, and PAL1 is qualified as being more important for the generation of phenylpropanoids. PMID:15377757

  6. The cpr5 mutant of Arabidopsis expresses both NPR1-dependent and NPR1-independent resistance.

    PubMed Central

    Bowling, S A; Clarke, J D; Liu, Y; Klessig, D F; Dong, X

    1997-01-01

    The cpr5 mutant was identified from a screen for constitutive expression of systemic acquired resistance (SAR). This single recessive mutation also leads to spontaneous expression of chlorotic lesions and reduced trichome development. The cpr5 plants were found to be constitutively resistant to two virulent pathogens, Pseudomonas syringae pv maculicola ES4326 and Peronospora parasitica Noco2; to have endogenous expression of the pathogenesis-related gene 1 (PR-1); and to have an elevated level of salicylic acid (SA). Lines homozygous for cpr5 and either the SA-degrading bacterial gene nahG or the SA-insensitive mutation npr1 do not express PR-1 or exhibit resistance to P. s. maculicola ES4326. Therefore, we conclude that cpr5 acts upstream of SA in inducing SAR. However, the cpr5 npr1 plants retained heightened resistance to P. parasitica Noco2 and elevated expression of the defensin gene PDF1.2, implying that NPR1-independent resistance signaling also occurs. We conclude that the cpr5 mutation leads to constitutive expression of both an NPR1-dependent and an NPR1-independent SAR pathway. Identification of this mutation indicates that these pathways are connected in early signal transduction steps and that they have overlapping functions in providing resistance. PMID:9338960

  7. Analysis of combinatorial loss-of-function mutants in the Arabidopsis ethylene receptors reveals that the ers1 etr1 double mutant has severe developmental defects that are EIN2 dependent.

    PubMed

    Hall, Anne E; Bleecker, Anthony B

    2003-09-01

    Ethylene responses in Arabidopsis are controlled by the ETR receptor family. The receptors function as negative regulators of downstream signal transduction components and fall into two distinct subfamilies based on sequence similarity. To clarify the levels of functional redundancy between receptor isoforms, combinatorial mutant lines were generated that included the newly isolated ers1-2 allele. Based on the etiolated seedling growth response, all mutant combinations tested exhibited some constitutive ethylene responsiveness but also remained responsive to exogenous ethylene, indicating that all five receptor isoforms can contribute to signaling and no one receptor subtype is essential. On the other hand, light-grown seedlings and adult ers1 etr1 double mutants exhibited severe phenotypes such as miniature rosette size, delayed flowering, and sterility, revealing a distinct role for subfamily I receptors in light-grown plants. Introduction of an ein2 loss-of-function mutation into the ers1 etr1 double mutant line resulted in plants that phenocopy ein2 single mutants, indicating that all phenotypes observed in the ers1 etr1 double mutant are EIN2 dependent.

  8. Analysis of Combinatorial Loss-of-Function Mutants in the Arabidopsis Ethylene Receptors Reveals That the ers1 etr1 Double Mutant Has Severe Developmental Defects That Are EIN2 Dependent

    PubMed Central

    Hall, Anne E.; Bleecker, Anthony B.

    2003-01-01

    Ethylene responses in Arabidopsis are controlled by the ETR receptor family. The receptors function as negative regulators of downstream signal transduction components and fall into two distinct subfamilies based on sequence similarity. To clarify the levels of functional redundancy between receptor isoforms, combinatorial mutant lines were generated that included the newly isolated ers1-2 allele. Based on the etiolated seedling growth response, all mutant combinations tested exhibited some constitutive ethylene responsiveness but also remained responsive to exogenous ethylene, indicating that all five receptor isoforms can contribute to signaling and no one receptor subtype is essential. On the other hand, light-grown seedlings and adult ers1 etr1 double mutants exhibited severe phenotypes such as miniature rosette size, delayed flowering, and sterility, revealing a distinct role for subfamily I receptors in light-grown plants. Introduction of an ein2 loss-of-function mutation into the ers1 etr1 double mutant line resulted in plants that phenocopy ein2 single mutants, indicating that all phenotypes observed in the ers1 etr1 double mutant are EIN2 dependent. PMID:12953109

  9. Identification of the Abundant Hydroxyproline-Rich Glycoproteins in the Root Walls of Wild-Type Arabidopsis, an ext3 Mutant Line, and Its Phenotypic Revertant

    PubMed Central

    Chen, Yuning; Ye, Dening; Held, Michael A.; Cannon, Maura C.; Ray, Tui; Saha, Prasenjit; Frye, Alexandra N.; Mort, Andrew J.; Kieliszewski, Marcia J.

    2015-01-01

    Extensins are members of the cell wall hydroxyproline-rich glycoprotein (HRGP) superfamily that form covalently cross-linked networks in primary cell walls. A knockout mutation in EXT3 (AT1G21310), the gene coding EXTENSIN 3 (EXT3) in Arabidopsis Landsberg erecta resulted in a lethal phenotype, although about 20% of the knockout plants have an apparently normal phenotype (ANP). In this study the root cell wall HRGP components of wild-type, ANP and the ext3 mutant seedlings were characterized by peptide fractionation of trypsin digested anhydrous hydrogen fluoride deglycosylated wall residues and by sequencing using LC-MS/MS. Several HRGPs, including EXT3, were identified in the wild-type root walls but not in walls of the ANP and lethal mutant. Indeed the ANP walls and walls of mutants displaying the lethal phenotype possessed HRGPs, but the profiles suggest that changes in the amount and perhaps type may account for the corresponding phenotypes. PMID:27135319

  10. The Seed Composition of Arabidopsis Mutants for the Group 3 Sulfate Transporters Indicates a Role in Sulfate Translocation within Developing Seeds1[C][W][OA

    PubMed Central

    Zuber, Hélène; Davidian, Jean-Claude; Aubert, Grégoire; Aimé, Delphine; Belghazi, Maya; Lugan, Raphaël; Heintz, Dimitri; Wirtz, Markus; Hell, Rüdiger; Thompson, Richard; Gallardo, Karine

    2010-01-01

    Sulfate is required for the synthesis of sulfur-containing amino acids and numerous other compounds essential for the plant life cycle. The delivery of sulfate to seeds and its translocation between seed tissues is likely to require specific transporters. In Arabidopsis (Arabidopsis thaliana), the group 3 plasmalemma-predicted sulfate transporters (SULTR3) comprise five genes, all expressed in developing seeds, especially in the tissues surrounding the embryo. Here, we show that sulfur supply to seeds is unaffected by T-DNA insertions in the SULTR3 genes. However, remarkably, an increased accumulation of sulfate was found in mature seeds of four mutants out of five. In these mutant seeds, the ratio of sulfur in sulfate form versus total sulfur was significantly increased, accompanied by a reduction in free cysteine content, which varied depending on the gene inactivated. These results demonstrate a reduced capacity of the mutant seeds to metabolize sulfate and suggest that these transporters may be involved in sulfate translocation between seed compartments. This was further supported by sulfate measurements of the envelopes separated from the embryo of the sultr3;2 mutant seeds, which showed differences in sulfate partitioning compared with the wild type. A dissection of the seed proteome of the sultr3 mutants revealed protein changes characteristic of a sulfur-stress response, supporting a role for these transporters in providing sulfate to the embryo. The mutants were affected in 12S globulin accumulation, demonstrating the importance of intraseed sulfate transport for the synthesis and maturation of embryo proteins. Metabolic adjustments were also revealed, some of which could release sulfur from glucosinolates. PMID:20702726

  11. The seed composition of Arabidopsis mutants for the group 3 sulfate transporters indicates a role in sulfate translocation within developing seeds.

    PubMed

    Zuber, Hélène; Davidian, Jean-Claude; Aubert, Grégoire; Aimé, Delphine; Belghazi, Maya; Lugan, Raphaël; Heintz, Dimitri; Wirtz, Markus; Hell, Rüdiger; Thompson, Richard; Gallardo, Karine

    2010-10-01

    Sulfate is required for the synthesis of sulfur-containing amino acids and numerous other compounds essential for the plant life cycle. The delivery of sulfate to seeds and its translocation between seed tissues is likely to require specific transporters. In Arabidopsis (Arabidopsis thaliana), the group 3 plasmalemma-predicted sulfate transporters (SULTR3) comprise five genes, all expressed in developing seeds, especially in the tissues surrounding the embryo. Here, we show that sulfur supply to seeds is unaffected by T-DNA insertions in the SULTR3 genes. However, remarkably, an increased accumulation of sulfate was found in mature seeds of four mutants out of five. In these mutant seeds, the ratio of sulfur in sulfate form versus total sulfur was significantly increased, accompanied by a reduction in free cysteine content, which varied depending on the gene inactivated. These results demonstrate a reduced capacity of the mutant seeds to metabolize sulfate and suggest that these transporters may be involved in sulfate translocation between seed compartments. This was further supported by sulfate measurements of the envelopes separated from the embryo of the sultr3;2 mutant seeds, which showed differences in sulfate partitioning compared with the wild type. A dissection of the seed proteome of the sultr3 mutants revealed protein changes characteristic of a sulfur-stress response, supporting a role for these transporters in providing sulfate to the embryo. The mutants were affected in 12S globulin accumulation, demonstrating the importance of intraseed sulfate transport for the synthesis and maturation of embryo proteins. Metabolic adjustments were also revealed, some of which could release sulfur from glucosinolates.

  12. Impaired Chloroplast Biogenesis in Immutans, an Arabidopsis Variegation Mutant, Modifies Developmental Programming, Cell Wall Composition and Resistance to Pseudomonas syringae

    PubMed Central

    Pogorelko, Gennady V.; Kambakam, Sekhar; Nolan, Trevor; Foudree, Andrew; Zabotina, Olga A.; Rodermel, Steven R.

    2016-01-01

    The immutans (im) variegation mutation of Arabidopsis has green- and white- sectored leaves due to action of a nuclear recessive gene. IM codes for PTOX, a plastoquinol oxidase in plastid membranes. Previous studies have revealed that the green and white sectors develop into sources (green tissues) and sinks (white tissues) early in leaf development. In this report we focus on white sectors, and show that their transformation into effective sinks involves a sharp reduction in plastid number and size. Despite these reductions, cells in the white sectors have near-normal amounts of plastid RNA and protein, and surprisingly, a marked amplification of chloroplast DNA. The maintenance of protein synthesis capacity in the white sectors might poise plastids for their development into other plastid types. The green and white im sectors have different cell wall compositions: whereas cell walls in the green sectors resemble those in wild type, cell walls in the white sectors have reduced lignin and cellulose microfibrils, as well as alterations in galactomannans and the decoration of xyloglucan. These changes promote susceptibility to the pathogen Pseudomonas syringae. Enhanced susceptibility can also be explained by repressed expression of some, but not all, defense genes. We suggest that differences in morphology, physiology and biochemistry between the green and white sectors is caused by a reprogramming of leaf development that is coordinated, in part, by mechanisms of retrograde (plastid-to-nucleus) signaling, perhaps mediated by ROS. We conclude that variegation mutants offer a novel system to study leaf developmental programming, cell wall metabolism and host-pathogen interactions. PMID:27050746

  13. Impaired Chloroplast Biogenesis in Immutans, an Arabidopsis Variegation Mutant, Modifies Developmental Programming, Cell Wall Composition and Resistance to Pseudomonas syringae

    DOE PAGES

    Pogorelko, Gennady V.; Kambakam, Sekhar; Nolan, Trevor; ...

    2016-04-06

    The immutans (im) variegation mutation of Arabidopsis has green- and white- sectored leaves due to action of a nuclear recessive gene. IM codes for PTOX, a plastoquinol oxidase in plastid membranes. Previous studies have revealed that the green and white sectors develop into sources (green tissues) and sinks (white tissues) early in leaf development. In this report we focus on white sectors, and show that their transformation into effective sinks involves a sharp reduction in plastid number and size. Despite these reductions, cells in the white sectors have near-normal amounts of plastid RNA and protein, and surprisingly, a marked amplificationmore » of chloroplast DNA. The maintenance of protein synthesis capacity in the white sectors might poise plastids for their development into other plastid types. The green and white im sectors have different cell wall compositions: whereas cell walls in the green sectors resemble those in wild type, cell walls in the white sectors have reduced lignin and cellulose microfibrils, as well as alterations in galactomannans and the decoration of xyloglucan. These changes promote susceptibility to the pathogen Pseudomonas syringae. Enhanced susceptibility can also be explained by repressed expression of some, but not all, defense genes. We suggest that differences in morphology, physiology and biochemistry between the green and white sectors is caused by a reprogramming of leaf development that is coordinated, in part, by mechanisms of retrograde (plastid-tonucleus) signaling, perhaps mediated by ROS. Lastly, we conclude that variegation mutants offer a novel system to study leaf developmental programming, cell wall metabolism and hostpathogen interactions.« less

  14. Impaired Chloroplast Biogenesis in Immutans, an Arabidopsis Variegation Mutant, Modifies Developmental Programming, Cell Wall Composition and Resistance to Pseudomonas syringae

    SciTech Connect

    Pogorelko, Gennady V.; Kambakam, Sekhar; Nolan, Trevor; Foudree, Andrew; Zabotina, Olga A.; Rodermel, Steven R.

    2016-04-06

    The immutans (im) variegation mutation of Arabidopsis has green- and white- sectored leaves due to action of a nuclear recessive gene. IM codes for PTOX, a plastoquinol oxidase in plastid membranes. Previous studies have revealed that the green and white sectors develop into sources (green tissues) and sinks (white tissues) early in leaf development. In this report we focus on white sectors, and show that their transformation into effective sinks involves a sharp reduction in plastid number and size. Despite these reductions, cells in the white sectors have near-normal amounts of plastid RNA and protein, and surprisingly, a marked amplification of chloroplast DNA. The maintenance of protein synthesis capacity in the white sectors might poise plastids for their development into other plastid types. The green and white im sectors have different cell wall compositions: whereas cell walls in the green sectors resemble those in wild type, cell walls in the white sectors have reduced lignin and cellulose microfibrils, as well as alterations in galactomannans and the decoration of xyloglucan. These changes promote susceptibility to the pathogen Pseudomonas syringae. Enhanced susceptibility can also be explained by repressed expression of some, but not all, defense genes. We suggest that differences in morphology, physiology and biochemistry between the green and white sectors is caused by a reprogramming of leaf development that is coordinated, in part, by mechanisms of retrograde (plastid-tonucleus) signaling, perhaps mediated by ROS. Lastly, we conclude that variegation mutants offer a novel system to study leaf developmental programming, cell wall metabolism and hostpathogen interactions.

  15. Transcriptomic Profiling of Arabidopsis thaliana Mutant pad2.1 in Response to Combined Cold and Osmotic Stress

    PubMed Central

    Kumar, Deepak; Datta, Riddhi; Hazra, Saptarshi; Sultana, Asma; Mukhopadhyay, Ria; Chattopadhyay, Sharmila

    2015-01-01

    The contribution of glutathione (GSH) in stress tolerance, defense response and antioxidant signaling is an established fact. In this study transcriptome analysis of pad2.1, an Arabidopsis thaliana mutant, after combined osmotic and cold stress treatment has been performed to explore the intricate position of GSH in the stress and defense signaling network in planta. Microarray data revealed the differential regulation of about 1674 genes in pad2.1 amongst which 973 and 701 were significantly up- and down-regulated respectively. Gene enrichment, functional pathway analysis by DAVID and MapMan analysis identified various stress and defense related genes viz. members of heat shock protein family, peptidyl prolyl isomerase (PPIase), thioredoxin peroxidase (TPX2), glutathione-S-transferase (GST), NBS-LRR type resistance protein etc. as down-regulated. The expression pattern of the above mentioned stress and defense related genes and APETALA were also validated by comparative proteomic analysis of combined stress treated Col-0 and pad2.1. Functional annotation noted down-regulation of UDP-glycosyl transferase, 4-coumarate CoA ligase 8, cinnamyl alcohol dehydrogenase 4 (CAD4), ACC synthase and ACC oxidase which are the important enzymes of phenylpropanoid, lignin and ethylene (ET) biosynthetic pathway respectively. Since the only difference between Col-0 (Wild type) and pad2.1 is the content of GSH, so, this study suggested that in addition to its association with specific stress responsive genes and proteins, GSH provides tolerance to plants by its involvement with phenylpropanoid, lignin and ET biosynthesis under stress conditions. PMID:25822199

  16. Impaired Chloroplast Biogenesis in Immutans, an Arabidopsis Variegation Mutant, Modifies Developmental Programming, Cell Wall Composition and Resistance to Pseudomonas syringae.

    PubMed

    Pogorelko, Gennady V; Kambakam, Sekhar; Nolan, Trevor; Foudree, Andrew; Zabotina, Olga A; Rodermel, Steven R

    2016-01-01

    The immutans (im) variegation mutation of Arabidopsis has green- and white- sectored leaves due to action of a nuclear recessive gene. IM codes for PTOX, a plastoquinol oxidase in plastid membranes. Previous studies have revealed that the green and white sectors develop into sources (green tissues) and sinks (white tissues) early in leaf development. In this report we focus on white sectors, and show that their transformation into effective sinks involves a sharp reduction in plastid number and size. Despite these reductions, cells in the white sectors have near-normal amounts of plastid RNA and protein, and surprisingly, a marked amplification of chloroplast DNA. The maintenance of protein synthesis capacity in the white sectors might poise plastids for their development into other plastid types. The green and white im sectors have different cell wall compositions: whereas cell walls in the green sectors resemble those in wild type, cell walls in the white sectors have reduced lignin and cellulose microfibrils, as well as alterations in galactomannans and the decoration of xyloglucan. These changes promote susceptibility to the pathogen Pseudomonas syringae. Enhanced susceptibility can also be explained by repressed expression of some, but not all, defense genes. We suggest that differences in morphology, physiology and biochemistry between the green and white sectors is caused by a reprogramming of leaf development that is coordinated, in part, by mechanisms of retrograde (plastid-to-nucleus) signaling, perhaps mediated by ROS. We conclude that variegation mutants offer a novel system to study leaf developmental programming, cell wall metabolism and host-pathogen interactions.

  17. Expansion and Functional Divergence of AP2 Group Genes in Spermatophytes Determined by Molecular Evolution and Arabidopsis Mutant Analysis

    PubMed Central

    Wang, Pengkai; Cheng, Tielong; Lu, Mengzhu; Liu, Guangxin; Li, Meiping; Shi, Jisen; Lu, Ye; Laux, Thomas; Chen, Jinhui

    2016-01-01

    The APETALA2 (AP2) genes represent the AP2 group within a large group of DNA-binding proteins called AP2/EREBP. The AP2 gene is functional and necessary for flower development, stem cell maintenance, and seed development, whereas the other members of AP2 group redundantly affect flowering time. Here we study the phylogeny of AP2 group genes in spermatophytes. Spermatophyte AP2 group genes can be classified into AP2 and TOE types, six clades, and we found that the AP2 group homologs in gymnosperms belong to the AP2 type, whereas TOE types are absent, which indicates the AP2 type gene are more ancient and TOE type was split out of AP2 type and losing the major function. In Brassicaceae, the expansion of AP2 and TOE type lead to the gene number of AP2 group were up to six. Purifying selection appears to have been the primary driving force of spermatophyte AP2 group evolution, although positive selection occurred in the AP2 clade. The transition from exon to intron of AtAP2 in Arabidopsis mutant leads to the loss of gene function and the same situation was found in AtTOE2. Combining this evolutionary analysis and published research, the results suggest that typical AP2 group genes may first appear in gymnosperms and diverged in angiosperms, following expansion of group members and functional differentiation. In angiosperms, AP2 genes (AP2 clade) inherited key functions from ancestors and other genes of AP2 group lost most function but just remained flowering time controlling in gene formation. In this study, the phylogenies of AP2 group genes in spermatophytes was analyzed, which supported the evidence for the research of gene functional evolution of AP2 group. PMID:27703459

  18. Pale-Green Phenotype of atl31 atl6 Double Mutant Leaves Is Caused by Disruption of 5-Aminolevulinic Acid Biosynthesis in Arabidopsis thaliana

    PubMed Central

    Maekawa, Shugo; Takabayashi, Atsushi; Huarancca Reyes, Thais; Yamamoto, Hiroko; Tanaka, Ayumi; Sato, Takeo; Yamaguchi, Junji

    2015-01-01

    Arabidopsis ubiquitin ligases ATL31 and homologue ATL6 control the carbon/nitrogen nutrient and pathogen responses. A mutant with the loss-of-function of both atl31 and atl6 developed light intensity-dependent pale-green true leaves, whereas the single knockout mutants did not. Plastid ultrastructure and Blue Native-PAGE analyses revealed that pale-green leaves contain abnormal plastid structure with highly reduced levels of thylakoid proteins. In contrast, the pale-green leaves of the atl31/atl6 mutant showed normal Fv/Fm. In the pale-green leaves of the atl31/atl6, the expression of HEMA1, which encodes the key enzyme for 5-aminolevulinic acid synthesis, the rate-limiting step in chlorophyll biosynthesis, was markedly down-regulated. The expression of key transcription factor GLK1, which directly promotes HEMA1 transcription, was also significantly decreased in atl31/atl6 mutant. Finally, application of 5-aminolevulinic acid to the atl31/atl6 mutants resulted in recovery to a green phenotype. Taken together, these findings indicate that the 5-aminolevulinic acid biosynthesis step was inhibited through the down-regulation of chlorophyll biosynthesis-related genes in the pale-green leaves of atl31/atl6 mutant. PMID:25706562

  19. Analysis of a Partial Male-Sterile Mutant of Arabidopsis thaliana Isolated from a Low-Energy Argon Ion Beam Mutagenized Pool

    NASA Astrophysics Data System (ADS)

    Xu, Min; Bian, Po; Wu, Yuejin; Yu, Zengliang

    2008-04-01

    A screen for Arabidopsis fertility mutants, mutagenized by low-energy argon ion beam, yielded two partial male-sterile mutants tc243-1 and tc243-2 which have similar phenotypes. tc243-2 was investigated in detail. The segregation ratio of the mutant phenotypes in the M2 pools suggested that mutation behaved as single Mendelian recessive mutations. tc243 showed a series of mutant phenotypes, among which partial male-sterile was its striking mutant characteristic. Phenotype analysis indicates that there are four factors leading to male sterility. a. Floral organs normally develop inside the closed bud, but the anther filaments do not elongate sufficiently to position the locules above the stigma at anthesis. b. The anther locules do not dehisce at the time of flower opening (although limited dehiscence occurs later). c. Pollens of mutant plants develop into several types of pollens at the trinucleated stage, as determined by staining with DAPI (4',6-diamidino-2-phenylindole), which shows a variable size, shape and number of nucleus. d. The viability of pollens is lower than that of the wild type on the germination test in vivo and vitro.

  20. Gravity-dependent differentiation and root coils in Arabidopsis thaliana wild type and phospholipase-A-I knockdown mutant grown on the International Space Station.

    PubMed

    Scherer, G F E; Pietrzyk, P

    2014-01-01

    Arabidopsis roots on 45° tilted agar in 1-g grow in wave-like figures. In addition to waves, formation of root coils is observed in several mutants compromised in gravitropism and/or auxin transport. The knockdown mutant ppla-I-1 of patatin-related phospholipase-A-I is delayed in root gravitropism and forms increased numbers of root coils. Three known factors contribute to waving: circumnutation, gravisensing and negative thigmotropism. In microgravity, deprivation of wild type (WT) and mutant roots of gravisensing and thigmotropism and circumnutation (known to slow down in microgravity, and could potentially lead to fewer waves or increased coiling in both WT and mutant). To resolve this, mutant ppla-I-1 and WT were grown in the BIOLAB facility in the International Space Station. In 1-g, roots of both types only showed waving. In the first experiment in microgravity, the mutant after 9 days formed far more coils than in 1-g but the WT also formed several coils. After 24 days in microgravity, in both types the coils were numerous with slightly more in the mutant. In the second experiment, after 9 days in microgravity only the mutant formed coils and the WT grew arcuated roots. Cell file rotation (CFR) on the mutant root surface in microgravity decreased in comparison to WT, and thus was not important for coiling. Several additional developmental responses (hypocotyl elongation, lateral root formation, cotyledon expansion) were found to be gravity-influenced. We tentatively discuss these in the context of disturbances in auxin transport, which are known to decrease through lack of gravity.

  1. Arabidopsis pab1, a mutant with reduced anthocyanins in immature seeds from banyuls, harbors a mutation in the MATE transporter FFT.

    PubMed

    Kitamura, Satoshi; Oono, Yutaka; Narumi, Issay

    2016-01-01

    Forward genetics approaches have helped elucidate the anthocyanin biosynthetic pathway in plants. Here, we used the Arabidopsis banyuls (ban) mutant, which accumulates anthocyanins, instead of colorless proanthocyanidin precursors, in immature seeds. In contrast to standard screens for mutants lacking anthocyanins in leaves/stems, we mutagenized ban plants and screened for mutants showing differences in pigmentation of immature seeds. The pale banyuls1 (pab1) mutation caused reduced anthocyanin pigmentation in immature seeds compared with ban. Immature pab1 ban seeds contained less anthocyanins and flavonols than ban, but showed normal expression of anthocyanin biosynthetic genes. In contrast to pab1, introduction of a flavonol-less mutation into ban did not produce paler immature seeds. Map-based cloning showed that two independent pab1 alleles disrupted the MATE-type transporter gene FFT/DTX35. Complementation of pab1 with FFT confirmed that mutation in FFT causes the pab1 phenotype. During development, FFT promoter activity was detected in the seed-coat layers that accumulate flavonoids. Anthocyanins accumulate in the vacuole and FFT fused to GFP mainly localized in the vacuolar membrane. Heterologous expression of grapevine MATE-type anthocyanin transporter gene partially complemented the pab1 phenotype. These results suggest that FFT acts at the vacuolar membrane in anthocyanin accumulation in the Arabidopsis seed coat, and that our screening strategy can reveal anthocyanin-related genes that have not been found by standard screening.

  2. The Cell Wall Arabinose-Deficient Arabidopsis thaliana Mutant murus5 Encodes a Defective Allele of REVERSIBLY GLYCOSYLATED POLYPEPTIDE21[OPEN

    PubMed Central

    Dugard, Christopher K.; Olek, Anna T.; Cooper, Bruce R.

    2016-01-01

    Traditional marker-based mapping and next-generation sequencing was used to determine that the Arabidopsis (Arabidopsis thaliana) low cell wall arabinose mutant murus5 (mur5) encodes a defective allele of REVERSIBLY GLYCOSYLATED POLYPEPTIDE2 (RGP2). Marker analysis of 13 F2 confirmed mutant progeny from a recombinant mapping population gave a rough map position on the upper arm of chromosome 5, and deep sequencing of DNA from these 13 lines gave five candidate genes with G→A (C→T) transitions predicted to result in amino acid changes. Of these five, only insertional mutant alleles of RGP2, a gene that encodes a UDP-arabinose mutase that interconverts UDP-arabinopyranose and UDP-arabinofuranose, exhibited the low cell wall arabinose phenotype. The identities of mur5 and two SALK insertional alleles were confirmed by allelism tests and overexpression of wild-type RGP2 complementary DNA placed under the control of the 35S promoter in the three alleles. The mur5 mutation results in the conversion of cysteine-257 to tyrosine-257 within a conserved hydrophobic cluster predicted to be distal to the active site and essential for protein stability and possible heterodimerization with other isoforms of RGP. PMID:27217494

  3. Differential abscisic acid regulation of guard cell slow anion channels in Arabidopsis wild-type and abi1 and abi2 mutants.

    PubMed Central

    Pei, Z M; Kuchitsu, K; Ward, J M; Schwarz, M; Schroeder, J I

    1997-01-01

    Abscisic acid (ABA) regulates vital physiological responses, and a number of events in the ABA signaling cascade remain to be identified. To allow quantitative analysis of genetic signaling mutants, patch-clamp experiments were developed and performed with the previously inaccessible Arabidopsis guard cells from the wild type and ABA-insensitive (abi) mutants. Slow anion channels have been proposed to play a rate-limiting role in ABA-induced stomatal closing. We now directly demonstrate that ABA strongly activates slow anion channels in wild-type guard cells. Furthermore, ABA-induced anion channel activation and stomatal closing were suppressed by protein phosphatase inhibitors. In abi1-1 and abi2-1 mutant guard cells, ABA activation of slow anion channels and ABA-induced stomatal closing were abolished. These impairments in ABA signaling were partially rescued by kinase inhibitors in abi1 but not in abi2 guard cells. These data provide cell biological evidence that the abi2 locus disrupts early ABA signaling, that abi1 and abi2 affect ABA signaling at different steps in the cascade, and that protein kinases act as negative regulators of ABA signaling in Arabidopsis. New models for ABA signaling pathways and roles for abi1, abi2, and protein kinases and phosphatases are discussed. PMID:9090884

  4. A starch deficient mutant of Arabidopsis thaliana with low ADPglucose pyrophosphorylase activity lacks one of the two subunits of the enzyme

    SciTech Connect

    Lin, Tsanpiao; Caspar, T.; Somerville, C.R.; Preiss, J. )

    1988-12-01

    A starch deficient mutant of Arabidopsis thaliana (L.) Heynh. has been isolated in which leaf extracts contain only about 5% as much activity of ADPglucose pyrophosphorylase (EC 2.7.7.27) as the wild type. A single, nuclear mutation at a previously undescribed locus designated adg2 is responsible for the mutant phenotype. Although the mutant contained only 5% as much ADPglucose pyrophosphorylase activity as the wild type, it accumulated 40% as much starch when grown in a 12 hour photoperiod. The mutant also contained about 40% as much starch as the wild type when grown in continuous light, suggesting that the rate of synthesis regulates its steady state accumulation. Immunological analysis of leaf extracts using antibodies against the spinach 54 and 51 kilodalton (kD) ADPglucose pyrophosphorylase subunits indicated that the mutant is deficient in a cross-reactive 54 kD polypeptide and has only about 4% as much as the wild type of a cross-reactive 51 kD polypeptide. This result and genetic studies suggested that adg2 is a structural gene which codes for the 54 kD polypeptide, and provides the first functional evidence that the 54 kD polypeptide is a required component of the native ADPglucose pyrophosphorylase enzyme.

  5. Accumulation of Zeaxanthin in Abscisic Acid-Deficient Mutants of Arabidopsis Does Not Affect Chlorophyll Fluorescence Quenching or Sensitivity to Photoinhibition in Vivo.

    PubMed Central

    Hurry, V.; Anderson, J. M.; Chow, W. S.; Osmond, C. B.

    1997-01-01

    Abscisic acid (ABA)-deficient mutants of Arabidopsis do not synthesize the epoxy-xanthophylls antheraxanthin, violaxanthin, or neoxanthin. However, thylakoid membranes from these mutants contain 3-fold more zeaxanthin than wild-type plants. This increase in zeaxanthin occurs as a stoichiometric replacement of the missing violaxanthin and neoxanthin within the pigment-protein complexes of both photosystem I and photosystem II (PSII). The retention of zeaxanthin in the dark by ABA-deficient mutants sensitizes the leaves to the development of nonphotochemical quenching (NPQ) during the first 2 to 4 min following a dark-light transition. However, the increase in pool size does not result in any increase in steady-state NPQ. When we exposed wild-type and ABA-deficient mutants leaves to twice growth irradiance, the mutants developed lower maximal NPQ but suffered similar photoinhibition to wildtype, measured both as a decline in the ratio of variable to maximal fluorescence and as a loss of functional PSII centers from oxygen flash yield measurements. These results suggest that only a few of the zeaxanthin molecules present within the light-harvesting antenna of PSII may be involved in NPQ and neither the accumulation of a large pool of zeaxanthin within the antenna of PSII nor an increase in conversion of violaxanthin to zeaxanthin will necessarily enhance photoprotective energy dissipation. PMID:12223632

  6. Partially dissecting the steady-state electron fluxes in Photosystem I in wild-type and pgr5 and ndh mutants of Arabidopsis

    PubMed Central

    Kou, Jiancun; Takahashi, Shunichi; Fan, Da-Yong; Badger, Murray R.; Chow, Wah S.

    2015-01-01

    Cyclic electron flux (CEF) around Photosystem I (PS I) is difficult to quantify. We obtained the linear electron flux (LEFO2) through both photosystems and the total electron flux through PS I (ETR1) in Arabidopsis in CO2-enriched air. ΔFlux = ETR1 – LEFO2 is an upper estimate of CEF, which consists of two components, an antimycin A-sensitive, PGR5 (proton gradient regulation 5 protein)-dependent component and an insensitive component facilitated by a chloroplastic nicotinamide adenine dinucleotide dehydrogenase-like complex (NDH). Using wild type as well as pgr5 and ndh mutants, we observed that (1) 40% of the absorbed light was partitioned to PS I; (2) at high irradiance a substantial antimycin A-sensitive CEF occurred in the wild type and the ndh mutant; (3) at low irradiance a sizable antimycin A-sensitive CEF occurred in the wild type but not in the ndh mutant, suggesting an enhancing effect of NDH in low light; and (4) in the pgr5 mutant, and the wild type and ndh mutant treated with antimycin A, a residual ΔFlux existed at high irradiance, attributable to charge recombination and/or pseudo-cyclic electron flow. Therefore, in low-light-acclimated plants exposed to high light, ΔFlux has contributions from various paths of electron flow through PS I. PMID:26442071

  7. Herbicide-resistant forms of Arabidopsis thaliana acetohydroxyacid synthase: characterization of the catalytic properties and sensitivity to inhibitors of four defined mutants.

    PubMed Central

    Chang, A K; Duggleby, R G

    1998-01-01

    Acetohydroxyacid synthase (AHAS) catalyses the first step in the synthesis of the branched-chain amino acids and is the target of several classes of herbicides. Four mutants (A122V, W574S, W574L and S653N) of the AHAS gene from Arabidopsis thaliana were constructed, expressed in Escherichia coli, and the enzymes were purified. Each mutant form and wild-type was characterized with respect to its catalytic properties and sensitivity to nine herbicides. Each enzyme had a pH optimum near 7.5. The specific activity varied from 13% (A122V) to 131% (W574L) of the wild-type and the Km for pyruvate of the mutants was similar to the wild-type, except for W574L where it was five-fold higher. The activation by cofactors (FAD, Mg2+ and thiamine diphosphate) was examined. A122V showed reduced affinity for all three cofactors, whereas S653N bound FAD more strongly than wild-type AHAS. Six sulphonylurea herbicides inhibited A122V to a similar degree as the wild-type but S653N showed a somewhat greater reduction in sensitivity to these compounds. In contrast, the W574 mutants were insensitive to these sulphonylureas, with increases in the Kiapp (apparent inhibition constant) of several hundred fold. All four mutants were resistant to three imidazolinone herbicides with decreases in sensitivity ranging from 100-fold to more than 1000-fold. PMID:9677339

  8. The Arabidopsis sickle mutant exhibits altered circadian clock responses to cool tempatures and tempature-dependent alternative splicing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The circadian clock allows plants to anticipate and respond to daily changes in ambient temperature. Mechanisms establishing the timing of circadian rhythms in Arabidopsis thaliana through temperature entrainment remain unclear. Also incompletely understood is the temperature compensation mechanism ...

  9. Modeling the Arabidopsis seed shape by a cardioid: efficacy of the adjustment with a scale change with factor equal to the Golden Ratio and analysis of seed shape in ethylene mutants.

    PubMed

    Cervantes, Emilio; Javier Martín, José; Ardanuy, Ramón; de Diego, Juana G; Tocino, Angel

    2010-03-15

    A new model for the description of Arabidopsis seed shape based on the comparison of the outline of its longitudinal section with a transformed cardioid is presented. The transformation consists of scaling the horizontal axis by a factor equal to the Golden Ratio. The elongated cardioid approximates the shape of the Arabidopsis seed with more accuracy than other figures. The length to width ratio in wild-type Columbia Arabidopsis dry seeds is close to the Golden Ratio and decreases over the course of imbibition. Dry seeds of etr1-1 mutants presented a reduced length to width ratio. Application of the new model based on the cardioid allows for comparison of shape between wild-type and mutant genotypes, revealing other general alterations in the seeds in ethylene signaling pathway mutants (etr1-1).

  10. Chlorophyll Synthesis in a Deetiolated (det340) Mutant of Arabidopsis without NADPH-Protochlorophyllide (PChlide) Oxidoreductase (POR) A and Photoactive PChlide-F655.

    PubMed Central

    Lebedev, N.; Van Cleve, B.; Armstrong, G.; Apel, K.

    1995-01-01

    Chlorophyll (Chl) synthesis in Arabidopsis is controlled by two light-dependent NADPH-protochlorophyllide (PChlide) oxidoreductases (PORs), one (POR A) that is active transiently in etiolated seedlings at the beginning of illumination and another (POR B) that also operates in green plants. The function of these two enzymes during the light-induced greening of dark-grown seedlings has been studied in the wild type and a deetiolated (det340) mutant of Arabidopsis. One of the consequences of the det mutation is that POR A is constitutively down-regulated, and therefore, synthesis of the POR A enzyme is shut off. When grown in the dark, the det340 mutant lacks POR A and the photoactive PChlide-F655 species but maintains the second PChlide reductase, POR B. Previously, photoactive PChlide-F655 has often been considered to be the only PChlide form that leads to Chl formation. Despite its deficiency in POR A and photoactive PChlide-F655, the det340 mutant is able to green when placed in the light. Chl accumulation, however, proceeds abnormally. At the beginning of illumination, seedlings of det340 mutants are extremely susceptible to photooxidative damage and accumulate Chl only at extremely low light intensities. They form core complexes of photosystems I and II but are almost completely devoid of light-harvesting structures. The results of this study demonstrate that in addition to the route of Chl synthesis that has been studied extensively in illuminated dark-grown wild-type plants, a second branch of Chl synthesis exists that is driven by POR B and does not require POR A. PMID:12242369

  11. Arabidopsis AtDjA3 Null Mutant Shows Increased Sensitivity to Abscisic Acid, Salt, and Osmotic Stress in Germination and Post-germination Stages

    PubMed Central

    Salas-Muñoz, Silvia; Rodríguez-Hernández, Aída A.; Ortega-Amaro, Maria A.; Salazar-Badillo, Fatima B.; Jiménez-Bremont, Juan F.

    2016-01-01

    DnaJ proteins are essential co-chaperones involved in abiotic and biotic stress responses. Arabidopsis AtDjA3 gene encodes a molecular co-chaperone of 420 amino acids, which belongs to the J-protein family. In this study, we report the functional characterization of the AtDjA3 gene using the Arabidopsis knockout line designated j3 and the 35S::AtDjA3 overexpression lines. Loss of AtDjA3 function was associated with small seed production. In fact, j3 mutant seeds showed a reduction of 24% in seed weight compared to Col-0 seeds. Expression analysis showed that the AtDjA3 gene was modulated in response to NaCl, glucose, and abscisic acid (ABA). The j3 line had increased sensitivity to NaCl and glucose treatments in the germination and cotyledon development in comparison to parental Col-0. Furthermore, the j3 mutant line exhibited higher ABA sensitivity in comparison to parental Col-0 and 35S::AtDjA3 overexpression lines. In addition, we examined the expression of ABI3 gene, which is a central regulator in ABA signaling, in j3 mutant and 35S::AtDjA3 overexpression lines. Under 5 μM ABA treatment at 24 h, j3 mutant seedlings displayed higher ABI3 expression, whereas in 35S::AtDjA3 overexpression lines, ABI3 gene expression was repressed. Taken together, these results demonstrate that the AtDjA3 gene is involved in seed development and abiotic stress tolerance. PMID:26941772

  12. Effect of hypergravity on lignin formation and expression of lignin-related genes in inflorescence stems of an ethylene-insensitive Arabidopsis mutant ein3-1

    NASA Astrophysics Data System (ADS)

    Karahara, Ichirou; Kobayashi, Mai; Tamaoki, Daisuke; Kamisaka, Seiichiro

    Our previous studies have shown that hypergravity inhibits growth and promotes lignin forma-tion in inflorescence stems of Arabidopsis thaliana by up-regulation of genes involved in lignin biosynthesis (Tamaoki et al. 2006, 2009). In the present study, we have examined whether ethylene is involved in these responses using an ethylene-insensitive Arabidopsis mutant ein3-1. Our results revealed that hypergravity treatment at 300 G for 24 h significantly inhibited growth of inflorescence stems, promoted both deposition of acetyl bromide extractable lignin and gene expression involved in lignin formation in inflorescence stems of wild type plants. Growth inhibition of inflorescence stems was also observed in ein3-1. However, the effects of hypergravity on the promotion of the deposition of acetyl bromide lignin and the expression of genes involved in lignin formation were not observed in ein3-1, indicating that ethylene sig-naling is involved in the up-regulation of the expression of lignin-related genes as well as the promotion of deposition of lignin by hypergravity in Arabidopsis inflorescence stems.

  13. Arabidopsis sos1 mutant in a salt-tolerant accession revealed an importance of salt acclimation ability in plant salt tolerance.

    PubMed

    Ariga, Hirotaka; Katori, Taku; Yoshihara, Ryouhei; Hase, Yoshihiro; Nozawa, Shigeki; Narumi, Issay; Iuchi, Satoshi; Kobayashi, Masatomo; Tezuka, Kenji; Sakata, Yoichi; Hayashi, Takahisa; Taji, Teruaki

    2013-07-01

    An analysis of the salinity tolerance of 354 Arabidopsis thaliana accessions showed that some accessions were more tolerant to salt shock than the reference accession, Col-0, when transferred from 0 to 225 mM NaCl. In addition, several accessions, including Zu-0, showed marked acquired salt tolerance after exposure to moderate salt stress. It is likely therefore that Arabidopsis plants have at least two types of tolerance, salt shock tolerance and acquired salt tolerance. To evaluate a role of well-known salt shock tolerant gene SOS1 in acquired salt tolerance, we isolated a sos1 mutant from ion-beam-mutagenized Zu-0 seedlings. The mutant showed severe growth inhibition under salt shock stress owing to a single base deletion in the SOS1 gene and was even more salt sensitive than Col-0. Nevertheless, it was able to survive after acclimation on 100 mM NaCl for 7 d followed by 750 mM sorbitol for 20 d, whereas Col-0 became chlorotic under the same conditions. We propose that genes for salt acclimation ability are different from genes for salt shock tolerance and play an important role in the acquisition of salt or osmotic tolerance.

  14. Arabidopsis sos1 mutant in a salt-tolerant accession revealed an importance of salt acclimation ability in plant salt tolerance

    PubMed Central

    Ariga, Hirotaka; Katori, Taku; Yoshihara, Ryouhei; Hase, Yoshihiro; Nozawa, Shigeki; Narumi, Issay; Iuchi, Satoshi; Kobayashi, Masatomo; Tezuka, Kenji; Sakata, Yoichi; Hayashi, Takahisa; Taji, Teruaki

    2013-01-01

    An analysis of the salinity tolerance of 354 Arabidopsis thaliana accessions showed that some accessions were more tolerant to salt shock than the reference accession, Col-0, when transferred from 0 to 225 mM NaCl. In addition, several accessions, including Zu-0, showed marked acquired salt tolerance after exposure to moderate salt stress. It is likely therefore that Arabidopsis plants have at least two types of tolerance, salt shock tolerance and acquired salt tolerance. To evaluate a role of well-known salt shock tolerant gene SOS1 in acquired salt tolerance, we isolated a sos1 mutant from ion-beam-mutagenized Zu-0 seedlings. The mutant showed severe growth inhibition under salt shock stress owing to a single base deletion in the SOS1 gene and was even more salt sensitive than Col-0. Nevertheless, it was able to survive after acclimation on 100 mM NaCl for 7 d followed by 750 mM sorbitol for 20 d, whereas Col-0 became chlorotic under the same conditions. We propose that genes for salt acclimation ability are different from genes for salt shock tolerance and play an important role in the acquisition of salt or osmotic tolerance. PMID:23656872

  15. Spatiotemporal modelling of hormonal crosstalk explains the level and patterning of hormones and gene expression in Arabidopsis thaliana wild-type and mutant roots.

    PubMed

    Moore, Simon; Zhang, Xiaoxian; Mudge, Anna; Rowe, James H; Topping, Jennifer F; Liu, Junli; Lindsey, Keith

    2015-09-01

    Patterning in Arabidopsis root development is coordinated via a localized auxin concentration maximum in the root tip, requiring the regulated expression of specific genes. However, little is known about how hormone and gene expression patterning is generated. Using a variety of experimental data, we develop a spatiotemporal hormonal crosstalk model that describes the integrated action of auxin, ethylene and cytokinin signalling, the POLARIS protein, and the functions of PIN and AUX1 auxin transporters. We also conduct novel experiments to confirm our modelling predictions. The model reproduces auxin patterning and trends in wild-type and mutants; reveals that coordinated PIN and AUX1 activities are required to generate correct auxin patterning; correctly predicts shoot to root auxin flux, auxin patterning in the aux1 mutant, the amounts of cytokinin, ethylene and PIN protein, and PIN protein patterning in wild-type and mutant roots. Modelling analysis further reveals how PIN protein patterning is related to the POLARIS protein through ethylene signalling. Modelling prediction of the patterning of POLARIS expression is confirmed experimentally. Our combined modelling and experimental analysis reveals that a hormonal crosstalk network regulates the emergence of patterns and levels of hormones and gene expression in wild-type and mutants.

  16. The Arabidopsis pi4kIIIβ1β2 double mutant is salicylic acid-overaccumulating: a new example of salicylic acid influence on plant stature

    PubMed Central

    Janda, Martin; Šašek, Vladimír; Ruelland, Eric

    2014-01-01

    Growth is the best visible sign of plant comfort. If plants are under stress, a difference in growth with control conditions can indicate that something is going wrong (or better). Phytohormones such as auxin, cytokinins, brassinosteroids or giberellins, are important growth regulators and their role in plant growth was extensively studied. On the other hand the role of salicylic acid (SA), a phytohormone commonly connected with plant defense responses, in plant growth is under-rated. However, studies with SA-overaccumulating mutants directly showed an influence of SA on plant growth. Recently we characterized an Arabidopsis SA-overaccumulating mutant impaired in phosphatidylinositol-4-kinases (pi4kIIIβ1β2). This mutant is dwarf. The crossing with mutants impaired in SA signaling revealed that pi4kIIIβ1β2 stunted rosette is due to high SA, while the short root length is not. This brings into evidence that upper and lower parts of the plants, even though they may share common phenotypes, are differently regulated. PMID:25482755

  17. The role of cell wall-based defences in the early restriction of non-pathogenic hrp mutant bacteria in Arabidopsis.

    PubMed

    Mitchell, Kathy; Brown, Ian; Knox, Paul; Mansfield, John

    2015-04-01

    We have investigated the cause of the restricted multiplication of hrp mutant bacteria in leaves of Arabidopsis. Our focus was on early interactions leading to differentiation between virulent wild-type and non-pathogenic hrpA mutant strains of Pseudomonas syringae pv. tomato. An initial drop in recoverable bacteria detected 0-4 h after inoculation with either strain was dependent on a functional FLS2 receptor and H2O2 accumulation in challenged leaves. Wild-type bacteria subsequently multiplied rapidly whereas the hrpA mutant was restricted within 6 h. Despite the early restriction, the hrpA mutant was still viable several days after inoculation. Analysis of intercellular washing fluids (IWFs), showed that high levels of nutrients were readily available to bacteria in the apoplast and that no diffusible inhibitors were produced in response to bacterial challenge. Histochemical and immunocytochemical methods were used to detect changes in polysaccharides (callose, two forms of cellulose, and pectin), arabinogalactan proteins (AGPs), H2O2 and peroxidase. Quantitative analysis showed very similar changes in localisation of AGPs, cellulose epitopes and callose 2 and 4 h after inoculation with either strain. However from 6 to 12 h after inoculation papillae expanded only next to the hrp mutant. In contrast to the similar patterns of secretory activity recorded from mesophyll cells, accumulation of H2O2 and peroxidase was significantly greater around the hrpA mutant within the first 4h after inoculation. A striking differential accumulation of H2O2 was also found in chloroplasts in cells next to the mutant. Ascorbate levels were lower in the IWFs recovered from sites inoculated with the hrp mutant than with wild-type bacteria. The critical response, observed at the right time and place to explain the observed differential behaviour of wild-type and hrpA mutant bacteria was the accumulation of H2O2, probably generated through Type III peroxidase activity and in

  18. Characterization of the snowy cotyledon 1 mutant of Arabidopsis thaliana: the impact of chloroplast elongation factor G on chloroplast development and plant vitality.

    PubMed

    Albrecht, Verónica; Ingenfeld, Anke; Apel, Klaus

    2006-03-01

    During seedling development chloroplast formation marks the transition from heterotrophic to autotrophic growth. The development and activity of chloroplasts may differ in cotyledons that initially serve as a storage organ and true leaves whose primary function is photosynthesis. A genetic screen was used for the identification of genes that affect selectively chloroplast function in cotyledons of Arabidopsis thaliana. Several mutants exhibiting pale cotyledons and green true leaves were isolated and dubbed snowy cotyledon (sco). One of the mutants, sco1, was characterized in more detail. The mutated gene was identified using map-based cloning. The mutant contains a point mutation in a gene encoding the chloroplast elongation factor G, leading to an amino acid exchange within the predicted 70S ribosome-binding domain. The mutation results in a delay in the onset of germination. At this early developmental stage embryos still contain undifferentiated proplastids, whose proper function seems necessary for seed germination. In light-grown sco1 seedlings the greening of cotyledons is severely impaired, whereas the following true leaves develop normally as in wild-type plants. Despite this apparent similarity of chloroplast development in true leaves of mutant and wild-type plants various aspects of mature plant development are also affected by the sco1 mutation such as the onset of flowering, the growth rate, and seed production. The onset of senescence in the mutant and the wild-type plants occurs, however, at the same time, suggesting that in the mutant this particular developmental step does not seem to suffer from reduced protein translation efficiency in chloroplasts.

  19. A combination of gene expression ranking and co-expression network analysis increases discovery rate in large-scale mutant screens for novel Arabidopsis thaliana abiotic stress genes.

    PubMed

    Ransbotyn, Vanessa; Yeger-Lotem, Esti; Basha, Omer; Acuna, Tania; Verduyn, Christoph; Gordon, Michal; Chalifa-Caspi, Vered; Hannah, Matthew A; Barak, Simon

    2015-05-01

    As challenges to food security increase, the demand for lead genes for improving crop production is growing. However, genetic screens of plant mutants typically yield very low frequencies of desired phenotypes. Here, we present a powerful computational approach for selecting candidate genes for screening insertion mutants. We combined ranking of Arabidopsis thaliana regulatory genes according to their expression in response to multiple abiotic stresses (Multiple Stress [MST] score), with stress-responsive RNA co-expression network analysis to select candidate multiple stress regulatory (MSTR) genes. Screening of 62 T-DNA insertion mutants defective in candidate MSTR genes, for abiotic stress germination phenotypes yielded a remarkable hit rate of up to 62%; this gene discovery rate is 48-fold greater than that of other large-scale insertional mutant screens. Moreover, the MST score of these genes could be used to prioritize them for screening. To evaluate the contribution of the co-expression analysis, we screened 64 additional mutant lines of MST-scored genes that did not appear in the RNA co-expression network. The screening of these MST-scored genes yielded a gene discovery rate of 36%, which is much higher than that of classic mutant screens but not as high as when picking candidate genes from the co-expression network. The MSTR co-expression network that we created, AraSTressRegNet is publicly available at http://netbio.bgu.ac.il/arnet. This systems biology-based screening approach combining gene ranking and network analysis could be generally applicable to enhancing identification of genes regulating additional processes in plants and other organisms provided that suitable transcriptome data are available.

  20. Detection of a gravitropism phenotype in glutamate receptor-like 3.3 mutants of Arabidopsis thaliana using machine vision and computation.

    PubMed

    Miller, Nathan D; Durham Brooks, Tessa L; Assadi, Amir H; Spalding, Edgar P

    2010-10-01

    Gene disruption frequently produces no phenotype in the model plant Arabidopsis thaliana, complicating studies of gene function. Functional redundancy between gene family members is one common explanation but inadequate detection methods could also be responsible. Here, newly developed methods for automated capture and processing of time series of images, followed by computational analysis employing modified linear discriminant analysis (LDA) and wavelet-based differentiation, were employed in a study of mutants lacking the Glutamate Receptor-Like 3.3 gene. Root gravitropism was selected as the process to study with high spatiotemporal resolution because the ligand-gated Ca(2+)-permeable channel encoded by GLR3.3 may contribute to the ion fluxes associated with gravity signal transduction in roots. Time series of root tip angles were collected from wild type and two different glr3.3 mutants across a grid of seed-size and seedling-age conditions previously found to be important to gravitropism. Statistical tests of average responses detected no significant difference between populations, but LDA separated both mutant alleles from the wild type. After projecting the data onto LDA solution vectors, glr3.3 mutants displayed greater population variance than the wild type in all four conditions. In three conditions the projection means also differed significantly between mutant and wild type. Wavelet analysis of the raw response curves showed that the LDA-detected phenotypes related to an early deceleration and subsequent slower-bending phase in glr3.3 mutants. These statistically significant, heritable, computation-based phenotypes generated insight into functions of GLR3.3 in gravitropism. The methods could be generally applicable to the study of phenotypes and therefore gene function.

  1. Elemental concentrations in the seed of mutants and natural variants of Arabidopsis thaliana grown under varying soil conditions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The concentrations of mineral nutrients in seeds are critical to both the life cycle of plants as well as human nutrition. These concentrations are strongly influenced by soil conditions, as shown here by quantifying the concentration of 14 elements in seeds from Arabidopsis thaliana plants grown un...

  2. A Novel fry1 Allele Reveals the Existence of a Mutant Phenotype Unrelated to 5′->3′ Exoribonuclease (XRN) Activities in Arabidopsis thaliana Roots

    PubMed Central

    Hirsch, Judith; Estavillo, Gonzalo M.; Javot, Hélène; Chiarenza, Serge; Mallory, Allison C.; Maizel, Alexis; Declerck, Marie; Pogson, Barry J.; Vaucheret, Hervé; Crespi, Martin; Desnos, Thierry; Thibaud, Marie-Christine; Nussaume, Laurent; Marin, Elena

    2011-01-01

    Background Mutations in the FRY1/SAL1 Arabidopsis locus are highly pleiotropic, affecting drought tolerance, leaf shape and root growth. FRY1 encodes a nucleotide phosphatase that in vitro has inositol polyphosphate 1-phosphatase and 3′,(2′),5′-bisphosphate nucleotide phosphatase activities. It is not clear which activity mediates each of the diverse biological functions of FRY1 in planta. Principal Findings A fry1 mutant was identified in a genetic screen for Arabidopsis mutants deregulated in the expression of Pi High affinity Transporter 1;4 (PHT1;4). Histological analysis revealed that, in roots, FRY1 expression was restricted to the stele and meristems. The fry1 mutant displayed an altered root architecture phenotype and an increased drought tolerance. All of the phenotypes analyzed were complemented with the AHL gene encoding a protein that converts 3′-polyadenosine 5′-phosphate (PAP) into AMP and Pi. PAP is known to inhibit exoribonucleases (XRN) in vitro. Accordingly, an xrn triple mutant with mutations in all three XRNs shared the fry1 drought tolerance and root architecture phenotypes. Interestingly these two traits were also complemented by grafting, revealing that drought tolerance was primarily conferred by the rosette and that the root architecture can be complemented by long-distance regulation derived from leaves. By contrast, PHT1 expression was not altered in xrn mutants or in grafting experiments. Thus, PHT1 up-regulation probably resulted from a local depletion of Pi in the fry1 stele. This hypothesis is supported by the identification of other genes modulated by Pi deficiency in the stele, which are found induced in a fry1 background. Conclusions/Significance Our results indicate that the 3′,(2′),5′-bisphosphate nucleotide phosphatase activity of FRY1 is involved in long-distance as well as local regulatory activities in roots. The local up-regulation of PHT1 genes transcription in roots likely results from local depletion of Pi

  3. Structural plasticity and functional implications of internal cavities in distal mutants of type 1 non-symbiotic hemoglobin AHb1 from Arabidopsis thaliana.

    PubMed

    Faggiano, Serena; Abbruzzetti, Stefania; Spyrakis, Francesca; Grandi, Elena; Viappiani, Cristiano; Bruno, Stefano; Mozzarelli, Andrea; Cozzini, Pietro; Astegno, Alessandra; Dominici, Paola; Brogioni, Silvia; Feis, Alessandro; Smulevich, Giulietta; Carrillo, Oliver; Schmidtke, Peter; Bidon-Chanal, Axel; Luque, F Javier

    2009-12-10

    The increasing number of nonsymbiotic plant hemoglobins discovered in genomic studies in the past decade raises intriguing questions about their physiological role. Among them, the nonsymbiotic hemoglobin AHb1 from Arabidopsis thaliana deserves particular attention, as it combines an extremely high oxygen affinity with an internal hexacoordination of the distal histidine HisE7 to the heme iron in the absence of exogenous ligands. In order to gain insight into the structure-function relationships of the protein, the ligand binding properties of mutants of two conserved residues of the distal cavity, HisE7 --> Leu and PheB10 --> Leu, were investigated by experimental and computational studies and compared to results determined for the wild type (wt) protein. The Fe(2+)-deoxy HisE7 --> Leu mutant exists, as expected, in the pentacoordinated form, while a mixture of penta- and hexacoordinated forms is found for the PheB10 --> Leu mutant, with an equilibrium shifted toward the pentacoordinated form with respect to the wt protein. Spectroscopic studies of the complexes of CO and CN(-) with AHb1 and its mutants show a subtle interplay of steric and electrostatic effects by distal residues on the ligand binding to the heme. Moreover, stopped-flow and flash photolysis experiments reveal substantial kinetic differences triggered by those mutations, which are particularly manifested in the enhanced geminate rebinding and bimolecular association rate. These findings are discussed in light of the drastic alterations found by molecular dynamics simulations in the nature and distribution of internal cavities in the protein matrix of the mutants, revealing an extremely large sensitivity of the protein structure to changes in distal HisE7 and PheB10 residues. Overall, data are consistent with the putative NO-dioxygenase activity attributed to AHb1.

  4. Molecular Characterization of the glauce Mutant: A Central Cell–Specific Function Is Required for Double Fertilization in Arabidopsis[W

    PubMed Central

    Leshem, Yehoram; Johnson, Cameron; Wuest, Samuel E.; Song, Xiaoya; Ngo, Quy A.; Grossniklaus, Ueli; Sundaresan, Venkatesan

    2012-01-01

    Double fertilization of the egg cell and the central cell by two sperm cells, resulting in the formation of the embryo and the endosperm, respectively, is a defining characteristic of flowering plants. The Arabidopsis thaliana female gametophytic mutant glauce (glc) can exhibit embryo development without any endosperm. Here, we show that in glc mutant embryo sacs one sperm cell successfully fuses with the egg cell but the second sperm cell fails to fuse with the central cell, resulting in single fertilization. Complementation studies using genes from the glc deletion interval identified an unusual genomic locus having homology to BAHD (for BEAT, AHCT, HCBT, and DAT) acyl-transferases with dual transcription units and alternative splicing that could rescue the sterility defect of glc. Expression of these transcripts appears restricted to the central cell, and expression within the central cell is sufficient to restore fertility. We conclude that the central cell actively promotes its own fertilization by the sperm cell through a signaling mechanism involving products of At1g65450. Successful fertilization of the egg cell is not blocked in the glc mutant, suggesting that evolution of double fertilization in flowering plants involved acquisition of specific functions by the central cell to enable its role as a second female gamete. PMID:22872756

  5. The Arabidopsis pxa1 Mutant Is Defective in an ATP-Binding Cassette Transporter-Like Protein Required for Peroxisomal Fatty Acid β-Oxidation1

    PubMed Central

    Zolman, Bethany K.; Silva, Illeana D.; Bartel, Bonnie

    2001-01-01

    Peroxisomes are important organelles in plant metabolism, containing all the enzymes required for fatty acid β-oxidation. More than 20 proteins are required for peroxisomal biogenesis and maintenance. The Arabidopsis pxa1 mutant, originally isolated because it is resistant to the auxin indole-3-butyric acid (IBA), developmentally arrests when germinated without supplemental sucrose, suggesting defects in fatty acid β-oxidation. Because IBA is converted to the more abundant auxin, indole-3-acetic acid (IAA), in a mechanism that parallels β-oxidation, the mutant is likely to be IBA resistant because it cannot convert IBA to IAA. Adult pxa1 plants grow slowly compared with wild type, with smaller rosettes, fewer leaves, and shorter inflorescence stems, indicating that PXA1 is important throughout development. We identified the molecular defect in pxa1 using a map-based positional approach. PXA1 encodes a predicted peroxisomal ATP-binding cassette transporter that is 42% identical to the human adrenoleukodystrophy (ALD) protein, which is defective in patients with the demyelinating disorder X-linked ALD. Homology to ALD protein and other human and yeast peroxisomal transporters suggests that PXA1 imports coenzyme A esters of fatty acids and IBA into the peroxisome for β-oxidation. The pxa1 mutant makes fewer lateral roots than wild type, both in response to IBA and without exogenous hormones, suggesting that the IAA derived from IBA during seedling development promotes lateral root formation. PMID:11706205

  6. Higher order Arabidopsis 14-3-3 mutants show 14-3-3 involvement in primary root growth both under control and abiotic stress conditions

    PubMed Central

    van Kleeff, P. J. M.; Jaspert, N.; Li, K. W.; Rauch, S.; Oecking, C.; de Boer, A. H.

    2014-01-01

    Arabidopsis 14-3-3 proteins are a family of conserved proteins that interact with numerous partner proteins in a phospho-specific manner, and can affect the target proteins in a number of ways; e.g. modification of enzymatic activity. We isolated T-DNA insertion lines in six 14-3-3 genes within the non-epsilon group that phylogenetically group in three closely related gene pairs. In total, 6 single, 3 double, 12 triple, and 3 quadruple mutants were generated. The mutants were phenotyped for primary root growth on control plates: single and double mutants were indistinguishable from WT, whereas six triples and all quadruples showed a shorter primary root. In addition, length of the first epidermal cell with a visible root hair bulge (LEH) was used to determine primary root elongation on medium containing mannitol and 1-aminocyclopropane-1-carboxylic acid (ACC). This analysis showed clear differences depending on the stress and 14-3-3 gene combinations. Next to the phenotypic growth analyses, a 14-3-3 pull-down assay on roots treated with and without mannitol showed that mannitol stress strongly affects the 14-3-3 interactome. In conclusion, we show gene specificity and functional redundancy among 14-3-3 proteins in primary root elongation under control and under abiotic stress conditions and changes in the 14-3-3 interactome during the onset of stress adaptation. PMID:25189593

  7. Annexin-like protein from Arabidopsis thaliana rescues delta oxyR mutant of Escherichia coli from H2O2 stress.

    PubMed Central

    Gidrol, X; Sabelli, P A; Fern, Y S; Kush, A K

    1996-01-01

    Reactive oxygen species are common causes of cellular damages in all aerobic organisms. In Escherichia coli, the oxyR gene product is a positive regulator of the oxyR regulon that is induced in response to H2O2 stress. To identify genes involved in counteracting oxidative stress in plants, we transformed a delta oxyR mutant of E. coli with an Arabidopsis thaliana cDNA library and selected for clones that restored the ability of the delta oxyR mutant to grow in the presence of H2O2. Using this approach, we isolated a cDNA that has strong homology with the annexin super-gene family. The complemented mutant showed higher catalase activity. mRNA expression of the annexin gene in A. thaliana was higher in roots as compared with other organs and was also increased when the plants were exposed to H2O2 stress or salicylic acid. Based on the results presented in this study, we propose a novel physiological role for annexin in counteracting H2O2 stress. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 7 Fig. 8 PMID:8855345

  8. A role for katanin in plant cell division: microtubule organization in dividing root cells of fra2 and lue1Arabidopsis thaliana mutants.

    PubMed

    Panteris, Emmanuel; Adamakis, Ioannis-Dimosthenis S; Voulgari, Georgia; Papadopoulou, Galini

    2011-07-01

    Severing of microtubules by katanin has proven to be crucial for cortical microtubule organization in elongating and differentiating plant cells. On the contrary, katanin is currently not considered essential during cell division in plants as it is in animals. However, defects in cell patterning have been observed in katanin mutants, implying a role for it in dividing plant cells. Therefore, microtubule organization was studied in detail by immunofluorescence in dividing root cells of fra2 and lue1 katanin mutants of Arabidopsis thaliana. In both, early preprophase bands consisted of poorly aligned microtubules, prophase spindles were multipolar, and the microtubules of expanding phragmoplasts were elongated, bended toward and connected to the surface of daughter nuclei. Accordingly, severing by katanin seems to be necessary for the proper organization of these microtubule arrays. In both fra2 and lue1, metaphase/anaphase spindles and initiating phragmoplasts exhibited typical organization. However, they were obliquely oriented more frequently than in the wild type. It is proposed that this oblique orientation may be due to prophase spindle multipolarity and results in a failure of the cell plate to follow the predetermined division plane, during cytokinesis, producing oblique cell walls in the roots of both mutants. It is therefore concluded that, like in animal cells, katanin is important for plant cell division, influencing the organization of several microtubule arrays. Moreover, failure in microtubule severing indirectly affects the orientation of the division plane.

  9. Flavonol-induced changes in PIN2 polarity and auxin transport in the Arabidopsis thaliana rol1-2 mutant require phosphatase activity

    PubMed Central

    Kuhn, Benjamin M.; Nodzyński, Tomasz; Errafi, Sanae; Bucher, Rahel; Gupta, Shibu; Aryal, Bibek; Dobrev, Petre; Bigler, Laurent; Geisler, Markus; Zažímalová, Eva; Friml, Jiří; Ringli, Christoph

    2017-01-01

    The phytohormone auxin is a major determinant and regulatory component important for plant development. Auxin transport between cells is mediated by a complex system of transporters such as AUX1/LAX, PIN, and ABCB proteins, and their localization and activity is thought to be influenced by phosphatases and kinases. Flavonols have been shown to alter auxin transport activity and changes in flavonol accumulation in the Arabidopsis thaliana rol1-2 mutant cause defects in auxin transport and seedling development. A new mutation in ROOTS CURL IN NPA 1 (RCN1), encoding a regulatory subunit of the phosphatase PP2A, was found to suppress the growth defects of rol1-2 without changing the flavonol content. rol1-2 rcn1-3 double mutants show wild type-like auxin transport activity while levels of free auxin are not affected by rcn1-3. In the rol1-2 mutant, PIN2 shows a flavonol-induced basal-to-apical shift in polar localization which is reversed in the rol1-2 rcn1-3 to basal localization. In vivo analysis of PINOID action, a kinase known to influence PIN protein localization in a PP2A-antagonistic manner, revealed a negative impact of flavonols on PINOID activity. Together, these data suggest that flavonols affect auxin transport by modifying the antagonistic kinase/phosphatase equilibrium. PMID:28165500

  10. Characterization of a NADH-dependent glutamate dehydrogenase mutant of Arabidopsis demonstrates the key role of this enzyme in root carbon and nitrogen metabolism.

    PubMed

    Fontaine, Jean-Xavier; Tercé-Laforgue, Thérèse; Armengaud, Patrick; Clément, Gilles; Renou, Jean-Pierre; Pelletier, Sandra; Catterou, Manuella; Azzopardi, Marianne; Gibon, Yves; Lea, Peter J; Hirel, Bertrand; Dubois, Frédéric

    2012-10-01

    The role of NADH-dependent glutamate dehydrogenase (GDH) was investigated by studying the physiological impact of a complete lack of enzyme activity in an Arabidopsis thaliana plant deficient in three genes encoding the enzyme. This study was conducted following the discovery that a third GDH gene is expressed in the mitochondria of the root companion cells, where all three active GDH enzyme proteins were shown to be present. A gdh1-2-3 triple mutant was constructed and exhibited major differences from the wild type in gene transcription and metabolite concentrations, and these differences appeared to originate in the roots. By placing the gdh triple mutant under continuous darkness for several days and comparing it to the wild type, the evidence strongly suggested that the main physiological function of NADH-GDH is to provide 2-oxoglutarate for the tricarboxylic acid cycle. The differences in key metabolites of the tricarboxylic acid cycle in the triple mutant versus the wild type indicated that, through metabolic processes operating mainly in roots, there was a strong impact on amino acid accumulation, in particular alanine, γ-aminobutyrate, and aspartate in both roots and leaves. These results are discussed in relation to the possible signaling and physiological functions of the enzyme at the interface of carbon and nitrogen metabolism.

  11. Reduced naphthylphthalamic acid binding in the tir3 mutant of Arabidopsis is associated with a reduction in polar auxin transport and diverse morphological defects

    NASA Technical Reports Server (NTRS)

    Ruegger, M.; Dewey, E.; Hobbie, L.; Brown, D.; Bernasconi, P.; Turner, J.; Muday, G.; Estelle, M.

    1997-01-01

    Polar auxin transport plays a key role in the regulation of plant growth and development. To identify genes involved in this process, we have developed a genetic procedure to screen for mutants of Arabidopsis that are altered in their response to auxin transport inhibitors. We recovered a total of 16 independent mutants that defined seven genes, called TRANSPORT INHIBITOR RESPONSE (TIR) genes. Recessive mutations in one of these genes, TIR3, result in altered responses to transport inhibitors, a reduction in polar auxin transport, and a variety of morphological defects that can be ascribed to changes in indole-3-acetic acid distribution. Most dramatically, tir3 seedlings are strongly deficient in lateral root production, a process that is known to depend on polar auxin transport from the shoot into the root. In addition, tir3 plants display a reduction in apical dominance as well as decreased elongation of siliques, pedicels, roots, and the inflorescence. Biochemical studies indicate that tir3 plants have a reduced number of N-1-naphthylphthalamic (NPA) binding sites, suggesting that the TIR3 gene is required for expression, localization, or stabilization of the NPA binding protein (NBP). Alternatively, the TIR3 gene may encode the NBP. Because the tir3 mutants have a substantial defect in NPA binding, their phenotype provides genetic evidence for a role for the NBP in plant growth and development.

  12. Proteomic Analysis of Different Mutant Genotypes of Arabidopsis Led to the Identification of 11 Proteins Correlating with Adventitious Root Development1[W

    PubMed Central

    Sorin, Céline; Negroni, Luc; Balliau, Thierry; Corti, Hélène; Jacquemot, Marie-Pierre; Davanture, Marlène; Sandberg, Göran; Zivy, Michel; Bellini, Catherine

    2006-01-01

    A lack of competence to form adventitious roots by cuttings or explants in vitro occurs routinely and is an obstacle for the clonal propagation and rapid fixation of elite genotypes. Adventitious rooting is known to be a quantitative genetic trait. We performed a proteomic analysis of Arabidopsis (Arabidopsis thaliana) mutants affected in their ability to develop adventitious roots in order to identify associated molecular markers that could be used to select genotypes for their rooting ability and/or to get further insight into the molecular mechanisms controlling adventitious rooting. Comparison of two-dimensional gel electrophoresis protein profiles resulted in the identification of 11 proteins whose abundance could be either positively or negatively correlated with endogenous auxin content, the number of adventitious root primordia, and/or the number of mature adventitious roots. One protein was negatively correlated only to the number of root primordia and two were negatively correlated to the number of mature adventitious roots. Two putative chaperone proteins were positively correlated only to the number of primordia, and, interestingly, three auxin-inducible GH3-like proteins were positively correlated with the number of mature adventitious roots. The others were correlated with more than one parameter. The 11 proteins are predicted to be involved in different biological processes, including the regulation of auxin homeostasis and light-associated metabolic pathways. The results identify regulatory pathways associated with adventitious root formation and represent valuable markers that might be used for the future identification of genotypes with better rooting abilities. PMID:16377752

  13. Intronic T-DNA Insertion Renders Arabidopsis opr3 a Conditional Jasmonic Acid-Producing Mutant1[C][W][OA

    PubMed Central

    Chehab, E. Wassim; Kim, Se; Savchenko, Tatyana; Kliebenstein, Daniel; Dehesh, Katayoon; Braam, Janet

    2011-01-01

    Jasmonic acid and its derived metabolites (JAs) orchestrate plant defense against insects and fungi. 12-Oxo-phytodienoic acid (OPDA), a JA precursor, has also been implicated in plant defense. We sought to define JAs and OPDA functions through comparative defense susceptibility characteristics of three Arabidopsis (Arabidopsis thaliana) genotypes: aos, lacking JAs and OPDA; opda reductase3 (opr3), deficient in JA production but can accumulate OPDA; and transgenics that overexpress OPR3. opr3, like aos, is susceptible to cabbage loopers (Trichoplusia ni) but, relative to aos, opr3 has enhanced resistance to a necrotrophic fungus. Gas chromatography-mass spectrometry reveals that opr3 produces OPDA but no detectable JAs following wounding and looper infestation; unexpectedly, substantial levels of JAs accumulate in opr3 upon fungal infection. Full-length OPR3 transcripts accumulate in fungal-infected opr3, potentially through splicing of the T-DNA containing intron. Fungal resistance correlates with levels of JAs not OPDA; therefore, opr3 resistance to some pests is likely due to JA accumulation, and signaling activities ascribed to OPDA should be reassessed because opr3 can produce JAs. Together these data (1) reinforce the primary role JAs play in plant defense against insects and necrotrophic fungi, (2) argue for a reassessment of signaling activities ascribed to OPDA, and (3) provide evidence that mutants with intron insertions can retain gene function. PMID:21487047

  14. Analysis of an Arabidopsis heat-sensitive mutant reveals that chlorophyll synthase is involved in reutilization of chlorophyllide during chlorophyll turnover.

    PubMed

    Lin, Yao-Pin; Lee, Tsung-yuan; Tanaka, Ayumi; Charng, Yee-yung

    2014-10-01

    Chlorophylls, the most abundant pigments in the photosynthetic apparatus, are constantly turned over as a result of the degradation and replacement of the damage-prone reaction center D1 protein of photosystem II. Results from isotope labeling experiments suggest that chlorophylls are recycled by reutilization of chlorophyllide and phytol, but the underlying mechanism is unclear. In this study, by characterization of a heat-sensitive Arabidopsis mutant we provide evidence of a salvage pathway for chlorophyllide a. A missense mutation in CHLOROPHYLL SYNTHASE (CHLG) was identified and confirmed to be responsible for a light-dependent, heat-induced cotyledon bleaching phenotype. Following heat treatment, mutant (chlg-1) but not wild-type seedlings accumulated a substantial level of chlorophyllide a, which resulted in a surge of phototoxic singlet oxygen. Immunoblot analysis suggested that the mutation destabilized the chlorophyll synthase proteins and caused a conditional blockage of esterification of chlorophyllide a after heat stress. Accumulation of chlorophyllide a after heat treatment occurred during recovery in the dark in the light-grown but not the etiolated seedlings, suggesting that the accumulated chlorophyllides were not derived from de novo biosynthesis but from de-esterification of the existing chlorophylls. Further analysis of the triple mutant harboring the CHLG mutant allele and null mutations of CHLOROPHYLLASE1 (CLH1) and CLH2 indicated that the known chlorophyllases are not responsible for the accumulation of chlorophyllide a in chlg-1. Taken together, our results show that chlorophyll synthase acts in a salvage pathway for chlorophyll biosynthesis by re-esterifying the chlorophyllide a produced during chlorophyll turnover.

  15. Expression of Camelina WRINKLED1 Isoforms Rescue the Seed Phenotype of the Arabidopsis wri1 Mutant and Increase the Triacylglycerol Content in Tobacco Leaves

    PubMed Central

    An, Dahee; Kim, Hyojin; Ju, Seulgi; Go, Young Sam; Kim, Hyun Uk; Suh, Mi Chung

    2017-01-01

    Triacylglycerol (TAG) is an energy-rich reserve in plant seeds that is composed of glycerol esters with three fatty acids. Since TAG can be used as a feedstock for the production of biofuels and bio-chemicals, producing TAGs in vegetative tissue is an alternative way of meeting the increasing demand for its usage. The WRINKLED1 (WRI1) gene is a well-established key transcriptional regulator involved in the upregulation of fatty acid biosynthesis in developing seeds. WRI1s from Arabidopsis and several other crops have been previously employed for increasing TAGs in seed and vegetative tissues. In the present study, we first identified three functional CsWRI1 genes (CsWRI1A. B, and C) from the Camelina oil crop and tested their ability to induce TAG synthesis in leaves. The amino acid sequences of CsWRI1s exhibited more than 90% identity with those of Arabidopsis WRI1. The transcript levels of the three CsWRI1 genes showed higher expression levels in developing seeds than in vegetative and floral tissues. When the CsWRI1A. B, or C was introduced into Arabidopsis wri1-3 loss-of-function mutant, the fatty acid content was restored to near wild-type levels and percentages of the wrinkled seeds were remarkably reduced in the transgenic lines relative to wri1-3 mutant line. In addition, the fluorescent signals of the enhanced yellow fluorescent protein (eYFP) fused to the CsWRI1 genes were observed in the nuclei of Nicotiana benthamiana leaf epidermal cells. Nile red staining indicated that the transient expression of CsWRI1A. B, or C caused an enhanced accumulation of oil bodies in N. benthamiana leaves. The levels of TAGs was higher by approximately 2.5- to 4.0-fold in N. benthamiana fresh leaves expressing CsWRI1 genes than in the control leaves. These results suggest that the three Camelina WRI1s can be used as key transcriptional regulators to increase fatty acids in biomass. PMID:28174580

  16. An UPLC-ESI-MS/MS Assay Using 6-Aminoquinolyl-N-Hydroxysuccinimidyl Carbamate Derivatization for Targeted Amino Acid Analysis: Application to Screening of Arabidopsis thaliana Mutants

    PubMed Central

    Salazar, Carolina; Armenta, Jenny M.; Shulaev, Vladimir

    2012-01-01

    In spite of the large arsenal of methodologies developed for amino acid assessment in complex matrices, their implementation in metabolomics studies involving wide-ranging mutant screening is hampered by their lack of high-throughput, sensitivity, reproducibility, and/or wide dynamic range. In response to the challenge of developing amino acid analysis methods that satisfy the criteria required for metabolomic studies, improved reverse-phase high-performance liquid chromatography-mass spectrometry (RPHPLC-MS) methods have been recently reported for large-scale screening of metabolic phenotypes. However, these methods focus on the direct analysis of underivatized amino acids and, therefore, problems associated with insufficient retention and resolution are observed due to the hydrophilic nature of amino acids. It is well known that derivatization methods render amino acids more amenable for reverse phase chromatographic analysis by introducing highly-hydrophobic tags in their carboxylic acid or amino functional group. Therefore, an analytical platform that combines the 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) pre-column derivatization method with ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) is presented in this article. For numerous reasons typical amino acid derivatization methods would be inadequate for large scale metabolic projects. However, AQC derivatization is a simple, rapid and reproducible way of obtaining stable amino acid adducts amenable for UPLC-ESI-MS/MS and the applicability of the method for high-throughput metabolomic analysis in Arabidopsis thaliana is demonstrated in this study. Overall, the major advantages offered by this amino acid analysis method include high-throughput, enhanced sensitivity and selectivity; characteristics that showcase its utility for the rapid screening of the preselected plant metabolites without compromising the quality of the metabolic data. The

  17. Robust root growth in altered hydrotropic response1 (ahr1) mutant of Arabidopsis is maintained by high rate of cell production at low water potential gradient.

    PubMed

    Salazar-Blas, Amed; Noriega-Calixto, Laura; Campos, María E; Eapen, Delfeena; Cruz-Vázquez, Tania; Castillo-Olamendi, Luis; Sepulveda-Jiménez, Gabriela; Porta, Helena; Dubrovsky, Joseph G; Cassab, Gladys I

    2017-01-01

    Hydrotropism is the directional root growth response determined by water stimulus. In a water potential gradient system (WPGS) the roots of the Arabidopsis wild type have a diminished root growth compared to normal medium (NM). In contrast, the altered hydrotropic response1 (ahr1) mutant roots maintain their robust growth in the same WPGS. The aims of this work were to ascertain how ahr1 roots could sustain growth in the WPGS, with a special focus on the integration of cellular processes involved in the signaling that determines root growth during abiotic stress and their relation to hydrotropism. Cellular analysis of the root apical meristem of ahr1 mutant contrary to the wild type showed an absence of changes in the meristem length, the elongation zone length, the length of fully elongated cells, and the cell cycle duration. The robust and steady root growth of ahr1 seedlings in the WPGS is explained by the mutant capacity to maintain cell production and cell elongation at the same level as in the NM. Analysis of auxin response at a transcriptional level showed that roots of the ahr1 mutant had a lower auxin response when grown in the WPGS, compared to wild type, indicating that auxin signaling participates in attenuation of root growth under water stress conditions. Also, wild type plants exhibited a high increase in proline content while ahr1 mutants showed minimum changes in the Normal Medium→Water Stress Medium (NM→WSM), a lower water potential gradient system than the WPGS. Accordingly, in this condition, gene expression of Δ1-6 Pyrroline-5-Carboxylate Synthetase1 (P5CS1) involved in proline synthesis strongly increased in wild type but not in ahr1 seedlings. The ahr1 phenotype shows unique features since the mutant root cells continue to proliferate and grow in the presence of a progressively negative water potential gradient at a level comparable to wild type growing in the NM. As such, it represents an exceptional resource for understanding

  18. Double knockout mutants of Arabidopsis grown under normal conditions reveal that the plastidial phosphorylase isozyme participates in transitory starch metabolism.

    PubMed

    Malinova, Irina; Mahlow, Sebastian; Alseekh, Saleh; Orawetz, Tom; Fernie, Alisdair R; Baumann, Otto; Steup, Martin; Fettke, Joerg

    2014-02-01

    In leaves of two starch-related single-knockout lines lacking either the cytosolic transglucosidase (also designated as disproportionating enzyme 2, DPE2) or the maltose transporter (MEX1), the activity of the plastidial phosphorylase isozyme (PHS1) is increased. In both mutants, metabolism of starch-derived maltose is impaired but inhibition is effective at different subcellular sites. Two constitutive double knockout mutants were generated (designated as dpe2-1×phs1a and mex1×phs1b) both lacking functional PHS1. They reveal that in normally grown plants, the plastidial phosphorylase isozyme participates in transitory starch degradation and that the central carbon metabolism is closely integrated into the entire cell biology. All plants were grown either under continuous illumination or in a light-dark regime. Both double mutants were compromised in growth and, compared with the single knockout plants, possess less average leaf starch when grown in a light-dark regime. Starch and chlorophyll contents decline with leaf age. As revealed by transmission electron microscopy, mesophyll cells degrade chloroplasts, but degradation is not observed in plants grown under continuous illumination. The two double mutants possess similar but not identical phenotypes. When grown in a light-dark regime, mesophyll chloroplasts of dpe2-1×phs1a contain a single starch granule but under continuous illumination more granules per chloroplast are formed. The other double mutant synthesizes more granules under either growth condition. In continuous light, growth of both double mutants is similar to that of the parental single knockout lines. Metabolite profiles and oligoglucan patterns differ largely in the two double mutants.

  19. Global Metabolic Profiling of Arabidopsis Polyamine Oxidase 4 (AtPAO4) Loss-of-Function Mutants Exhibiting Delayed Dark-Induced Senescence

    PubMed Central

    Sequera-Mutiozabal, Miren I.; Erban, Alexander; Kopka, Joachim; Atanasov, Kostadin E.; Bastida, Jaume; Fotopoulos, Vasileios; Alcázar, Rubén; Tiburcio, Antonio F.

    2016-01-01

    Early and more recent studies have suggested that some polyamines (PAs), and particularly spermine (Spm), exhibit anti-senescence properties in plants. In this work, we have investigated the role of Arabidopsis Polyamine Oxidase 4 (PAO4), encoding a PA back-conversion oxidase, during dark-induced senescence. Two independent PAO4 (pao4-1 and pao4-2) loss-of-function mutants have been found that accumulate 10-fold higher Spm, and this associated with delayed entry into senescence under dark conditions. Mechanisms underlying pao4 delayed senescence have been studied using global metabolic profiling by GC-TOF/MS. pao4 mutants exhibit constitutively higher levels of important metabolites involved in redox regulation, central metabolism and signaling that support a priming status against oxidative stress. During senescence, interactions between PAs and oxidative, sugar and nitrogen metabolism have been detected that additively contribute to delayed entry into senescence. Our results indicate the occurrence of metabolic interactions between PAs, particularly Spm, with cell oxidative balance and transport/biosynthesis of amino acids as a strategy to cope with oxidative damage produced during senescence. PMID:26925084

  20. Root-growth behavior of the Arabidopsis mutant rgr1. Roles of gravitropism and circumnutation in the waving/coiling phenomenon

    NASA Technical Reports Server (NTRS)

    Mullen, J. L.; Turk, E.; Johnson, K.; Wolverton, C.; Ishikawa, H.; Simmons, C.; Soll, D.; Evans, M. L.

    1998-01-01

    In this study we investigated the kinetics of the gravitropic response of the Arabidopsis mutant rgr1 (reduced root gravitropism). Although the rate of curvature in rgr1, which is allelic to axr4, was smaller than in the wild type (ecotype Wassilewskija), curvature was initiated in the same region of the root, the distal elongation zone. The time lag for the response was unaffected in the mutant; however, the gravitropic response of rgr1 contained a feature not found in the wild type: when roots growing along the surface of an agar plate were gravistimulated, there was often an upward curvature that initiated in the central elongation zone. Because this response was dependent on the tactile environment of the root, it most likely resulted from the superposition of the waving/coiling phenomenon onto the gravitropic response. We found that the frequency of the waving pattern and circumnutation, a cyclic endogenous pattern of root growth, was the same in rgr1 and in the wild type, so the waving/coiling phenomenon is likely governed by circumnutation patterns. The amplitudes of these oscillations may then be selectively amplified by tactile stimulation to provide a directional preference to the slanting.

  1. Five components of the ethylene-response pathway identified in a screen for weak ethylene-insensitive mutants in Arabidopsis.

    PubMed

    Alonso, Jose M; Stepanova, Anna N; Solano, Roberto; Wisman, Ellen; Ferrari, Simone; Ausubel, Frederick M; Ecker, Joseph R

    2003-03-04

    Five ethylene-insensitive loci (wei1-wei5) were identified by using a low-dose screen for "weak" ethylene-insensitive mutants. wei1, wei2, and wei3 seedlings showed hormone insensitivity only in roots, whereas wei4 and wei5 displayed insensitivity in both roots and hypocotyls. The genes corresponding to wei1, wei4, and wei5 were isolated using a positional cloning approach. The wei1 mutant harbored a recessive mutation in TIR1, which encodes a component of the SCF protein ubiquitin ligase involved in the auxin response. wei4, a dominant mutant, resulted from a mutation in the ethylene receptor ERS, whereas wei5, a semidominant mutant, was caused by a mutation in the EIN3-related transcription factor gene EIL1. The simultaneous loss of functional WEI5EIL1 and EIN3 nearly completely abolished the ethylene response in etiolated seedlings, and adult plants were highly susceptible to infection by the necrotrophic fungal pathogen Botrytis cinerea. Moreover, wei5eil1 ein3 double mutants were able to fully suppress constitutive signaling caused by ctr1, suggesting a synergistic interaction among these gene products. Unlike previously known root ethylene-insensitive mutants, wei2 and wei3 were not affected in their response to auxin and showed a normal response to gravity. Genetic mapping studies indicate that wei2 and wei3 correspond to previously unidentified ethylene pathway genes that may control cell-elongation processes functioning at the intersection of the ethylene and auxin response pathways.

  2. Aberrant gene expression in the Arabidopsis SULTR1;2 mutants suggests a possible regulatory role for this sulfate transporter in response to sulfur nutrient status.

    PubMed

    Zhang, Bo; Pasini, Rita; Dan, Hanbin; Joshi, Naveen; Zhao, Yihong; Leustek, Thomas; Zheng, Zhi-Liang

    2014-01-01

    Sulfur is required for the biosynthesis of cysteine, methionine and numerous other metabolites, and thus is critical for cellular metabolism and various growth and developmental processes. Plants are able to sense their physiological state with respect to sulfur availability, but the sensor remains to be identified. Here we report the isolation and characterization of two novel allelic mutants of Arabidopsis thaliana, sel1-15 and sel1-16, which show increased expression of a sulfur deficiency-activated gene β-glucosidase 28 (BGLU28). The mutants, which represent two different missense alleles of SULTR1;2, which encodes a high-affinity sulfate transporter, are defective in sulfate transport and as a result have a lower cellular sulfate level. However, when treated with a very high dose of sulfate, sel1-15 and sel1-16 accumulated similar amounts of internal sulfate and its metabolite glutathione (GSH) to wild-type, but showed higher expression of BGLU28 and other sulfur deficiency-activated genes than wild-type. Reduced sensitivity to inhibition of gene expression was also observed in the sel1 mutants when fed with the sulfate metabolites Cys and GSH. In addition, a SULTR1;2 knockout allele also exhibits reduced inhibition in response to sulfate, Cys and GSH, consistent with the phenotype of sel1-15 and sel1-16. Taken together, the genetic evidence suggests that, in addition to its known function as a high-affinity sulfate transporter, SULTR1;2 may have a regulatory role in response to sulfur nutrient status. The possibility that SULTR1;2 may function as a sensor of sulfur status or a component of a sulfur sensory mechanism is discussed.

  3. THI1, a protein involved in the biosynthesis of thiamin in Arabidopsis thaliana: structural analysis of THI1(A140V) mutant.

    PubMed

    Garcia, Assuero F; Dyszy, Fabio; Munte, Claudia E; Demarco, Ricardo; Beltramini, Leila M; Oliva, Glaucius; Costa-Filho, Antonio J; Araujo, Ana P U

    2014-06-01

    In eukaryotes, there are still steps of the vitamin B1 biosynthetic pathway not completely understood. In Arabidopsis thaliana, THI1 protein has been associated with the synthesis of the thiazole ring, a finding supported by the identification of a thiamine pyrophosphate (TPP)-like compound in its structure. Here, we investigated THI1 and its mutant THI1(A140V), responsible for the thiamin auxotrophy in a A. thaliana mutant line, aiming to clarify the impact of this mutation in the stability and activity of THI1. Recently, the THI1 orthologue (THI4) was revealed to be responsible for the donation of the sulfur atom from a cysteine residue to the thiazole ring in the thiamine intermediate. In this context, we carried out a cysteine quantification in THI1 and THI1(A140V) using electron spin resonance (ESR). These data showed that THI1(A140V) contains more sulfur-containing cysteines than THI1, indicating that the function as a sulfur donor is conserved, but the rate of donation reaction is somehow affected. Also, the bound compounds were isolated from both proteins and are present in different amounts in each protein. Unfolding studies presented differences in melting temperatures and also in the concentration of guanidine at which half of the protein unfolds, thus showing that THI1(A140V) has its conformational stability affected by the mutation. Hence, despite keeping its function in the early steps during the synthesis of TPP precursor, our studies have shown a decrease in the THI1(A140V) stability, which might be slowing down the biological activity of the mutant, and thus contributing to thiamin auxotrophy.

  4. Salicylic acid deficiency in NahG transgenic lines and sid2 mutants increases seed yield in the annual plant Arabidopsis thaliana.

    PubMed

    Abreu, Maria Elizabeth; Munné-Bosch, Sergi

    2009-01-01

    Salicylic acid-deficient NahG transgenic lines and sid2 mutants were used to evaluate the role of this compound in the development of the short-lived, annual plant Arabidopsis thaliana, with a particular focus on the interplay between salicylic acid and other phytohormones. Low salicylic acid levels led to increased growth, as well as to smaller abscisic acid levels and reduced damage to PSII (as indicated by F(v)/F(m) ratios) during the reproductive stages in rosette leaves of NahG transgenic lines and sid2 mutants, compared with wild-type plants. Furthermore, salicylic acid deficiency highly influenced seed yield and composition. Seed production increased by 4.4-fold and 3.5-fold in NahG transgenic lines and sid2 mutants, respectively, compared to the wild type. Salicylic acid deficiency also improved seed composition in terms of antioxidant vitamin concentrations, seeds of salicylic acid-deficient plants showing higher levels of alpha- and gamma-tocopherol (vitamin E) and beta-carotene (pro-vitamin A) than seeds of wild-type plants. Seeds of salicylic acid-deficient plants also showed higher nitrogen concentrations than seeds of wild-type plants. It is concluded that (i) the sid2 gene, which encodes for isochorismate synthase, plays a central role in salicylic acid biosynthesis during plant development in A. thaliana, (ii) salicylic acid plays a role in the regulation of growth, senescence, and seed production, (iii) there is a cross-talk between salicylic acid and other phytohormones during plant development, and (iv) the concentrations of antioxidant vitamins in seeds may be influenced by the endogenous levels of salicylic acid in plants.

  5. Functional Analysis of Arabidopsis Mutants Points to Novel Roles for Glutathione in Coupling H2O2 to Activation of Salicylic Acid Accumulation and Signaling

    PubMed Central

    Han, Yi; Chaouch, Sejir; Mhamdi, Amna; Queval, Guillaume; Zechmann, Bernd

    2013-01-01

    Abstract Aims: Through its interaction with H2O2, glutathione is a candidate for transmission of signals in plant responses to pathogens, but identification of signaling roles is complicated by its antioxidant function. Using a genetic approach based on a conditional catalase-deficient Arabidopsis mutant, cat2, this study aimed at establishing whether GSH plays an important functional role in the transmission of signals downstream of H2O2. Results: Introducing the cad2 or allelic mutations in the glutathione synthesis pathway into cat2 blocked H2O2-triggered GSH oxidation and accumulation. While no effects on NADP(H) or ascorbate were observed, and H2O2-induced decreases in growth were maintained, blocking GSH modulation antagonized salicylic acid (SA) accumulation and SA-dependent responses. Other novel double and triple mutants were produced and compared with cat2 cad2 at the levels of phenotype, expression of marker genes, nontargeted metabolite profiling, accumulation of SA, and bacterial resistance. Most of the effects of the cad2 mutation on H2O2-triggered responses were distinct from those produced by mutations for GLUTATHIONE REDUCTASE1 (GR1) or NONEXPRESSOR OF PATHOGENESIS-RELATED GENES 1 (NPR1), and were linked to compromised induction of ISOCHORISMATE SYNTHASE1 (ICS1) and ICS1-dependent SA accumulation. Innovation: A novel genetic approach was used in which GSH content or antioxidative capacity was independently modified in an H2O2 signaling background. Analysis of new double and triple mutants allowed us to infer previously undescribed regulatory roles for GSH. Conclusion: In parallel to its antioxidant role, GSH acts independently of NPR1 to allow increased intracellular H2O2 to activate SA signaling, a key defense response in plants. Antioxid. Redox Signal. 18, 2106–2121. PMID:23148658

  6. Biochemical and quantitative proteomics investigations in Arabidopsis ggt1 mutant leaves reveal a role for the gamma-glutamyl cycle in plant's adaptation to environment.

    PubMed

    Tolin, Serena; Arrigoni, Giorgio; Trentin, Anna Rita; Veljovic-Jovanovic, Sonja; Pivato, Micaela; Zechman, Bernd; Masi, Antonio

    2013-06-01

    The existence of a gamma-glutamyl cycle consisting of intracellular GSH synthesis, extrusion to the apoplastic space and recovery by gamma-glutamyl transferase (GGT)-assisted degradation into its constituent amino acids, has been demonstrated in plants. To address the significance of this cycle in plant cells, we performed integrated biochemical, immunocytochemical, and quantitative proteomics analyses in the Arabidopsis thaliana ggt1 knockout mutant (lacking apoplastic GGT1 isoform) and its corresponding wild-type (WT). The ggt1 knockout leaves exhibited an increased ascorbate and GSH content, increased apoplastic GSH content, and enhanced protein carbonylations in the low-molecular weight range compared to WT. The combined iTRAQ and LC-MS/MS-based quantitative proteomics approach identified 70 proteins (out of 1013 identified proteins) whose abundance was significantly different in leaves of ggt1 mutant compared to WT, with a fold change ≥1.5. Mining of the proteome data for GSH-associated genes showed that disruption of gamma-glutamyl cycle in ggt1 knockout-leaves was associated with the induction of genes encoding four GSTs in the phi class (GSTF2, GSTF6, GSTF9, and GSTF10), a GSH peroxidase (GPX1), and glyoxylase II. Proteins with a lower abundance compared to the WT are involved in chloroplast functions, carbohydrate/maltose metabolism, and vegetative storage protein synthesis. Present findings suggest that GGT1 plays a role in redox signaling. The disruption of the gamma-glutamyl cycle in the ggt1 mutant results in pleiotropic effects related to biotic and abiotic stress response, antioxidant metabolism, senescence, carbohydrate metabolism, and photosynthesis, with strong implications for plant adaptation to the environment.

  7. Modulation of biosynthesis of photosynthetic pigments and light-harvesting complex in wild-type and gun5 mutant of Arabidopsis thaliana during impaired chloroplast development.

    PubMed

    Pattanayak, Gopal K; Tripathy, Baishnab C

    2016-05-01

    Plants in response to different environmental cues need to modulate the expression of nuclear and chloroplast genomes that are in constant communication. To understand the signals that are responsible for inter-organellar communication, levulinic acid (LA), an inhibitor of 5-aminolevulinic acid dehydratase, was used to suppress the synthesis of pyrrole-derived tetrapyrroles chlorophylls. Although, it does not specifically inhibit carotenoid biosynthesis enzymes, LA reduced the carotenoid contents during photomorphogenesis of etiolated Arabidopsis seedlings. The expression of nuclear genes involved in carotenoid biosynthesis, i.e., geranylgeranyl diphosphate synthase, phytoene synthase, and phytoene desaturase, was downregulated in LA-treated seedlings. Similarly, the transcript abundance of nuclear genes, i.e., Lhcb1, PsbO, and RcbS, coding for chloroplastic proteins was severely attenuated in LA-treated samples. In contrast, LA treatment did not affect the transcript abundance of chalcone synthase, a marker gene for cytoplasm, and β-ATP synthase, a marker gene for mitochondria. This demonstrates the retrograde signaling from chloroplast to nucleus to suppress chloroplastic proteins during impaired chloroplast development. However, under identical conditions in LA-treated tetrapyrrole-deficient gun5 mutant, retrograde signal continued. The tetrapyrrole biosynthesis inhibitor LA suppressed formation of all tetrapyrroles both in WT and gun5. This rules out the role of tetrapyrroles as signaling molecules in WT and gun5. The removal of LA from the Arabidopsis seedlings restored the chlorophyll and carotenoid contents and expression of nuclear genes coding for chloroplastic proteins involved in chloroplast biogenesis. Therefore, LA could be used to modulate chloroplast biogenesis at a desired phase of chloroplast development.

  8. Arabidopsis inositol polyphosphate 6-/3-kinase is a nuclear protein that complements a yeast mutant lacking a functional ArgR-Mcm1 transcription complex.

    PubMed

    Xia, Hui-Jun; Brearley, Charles; Elge, Stephan; Kaplan, Boaz; Fromm, Hillel; Mueller-Roeber, Bernd

    2003-02-01

    Inositol 1,4,5-trisphosphate 3-kinase, and more generally inositol polyphosphate kinases (Ipk), play important roles in signal transduction in animal cells; however, their functions in plant cells remain to be elucidated. Here, we report the molecular cloning of a cDNA (AtIpk2beta) from a higher plant, Arabidopsis. Arabidopsis AtIpk2beta is a 33-kD protein that exhibits weak homology ( approximately 25% identical amino acids) with Ipk proteins from animals and yeast and lacks a calmodulin binding site, as revealed by sequence analysis and calmodulin binding assays. However, recombinant AtIpk2beta phosphorylates inositol 1,4,5-trisphosphate to inositol 1,4,5,6-tetrakisphosphate and also converts it to inositol 1,3,4,5,6-pentakisphosphate [Ins(1,3,4,5,6)P(5)]. AtIpk2beta also phosphorylates inositol 1,3,4,5-tetrakisphosphate to Ins(1,3,4,5,6)P(5). Thus, the enzyme is a D3/D6 dual-specificity inositol phosphate kinase. AtIpk2beta complements a yeast ARG82/IPK2 mutant lacking a functional ArgR-Mcm1 transcription complex. This complex is involved in regulating Arg metabolism-related gene expression and requires inositol polyphosphate kinase activity to function. AtIpk2beta was found to be located predominantly in the nucleus of plant cells, as demonstrated by immunolocalization and fusion to green fluorescent protein. RNA gel blot analysis and promoter-beta-glucuronidase reporter gene studies demonstrated AtIpk2beta gene expression in various organs tested. These data suggest a role for AtIpk2beta as a transcriptional control mediator in plants.

  9. Diurnal Changes of Polysome Loading Track Sucrose Content in the Rosette of Wild-Type Arabidopsis and the Starchless pgm Mutant1[W][OA

    PubMed Central

    Pal, Sunil Kumar; Liput, Magdalena; Piques, Maria; Ishihara, Hirofumi; Obata, Toshihiro; Martins, Marina C.M.; Sulpice, Ronan; van Dongen, Joost T.; Fernie, Alisdair R.; Yadav, Umesh Prasad; Lunn, John E.; Usadel, Björn; Stitt, Mark

    2013-01-01

    Growth is driven by newly fixed carbon in the light, but at night it depends on reserves, like starch, that are laid down in the light. Unless plants coordinate their growth with diurnal changes in the carbon supply, they will experience acute carbon starvation during the night. Protein synthesis represents a major component of cellular growth. Polysome loading was investigated during the diurnal cycle, an extended night, and low CO2 in Arabidopsis (Arabidopsis thaliana) Columbia (Col-0) and in the starchless phosphoglucomutase (pgm) mutant. In Col-0, polysome loading was 60% to 70% in the light, 40% to 45% for much of the night, and less than 20% in an extended night, while in pgm, it fell to less than 25% early in the night. Quantification of ribosomal RNA species using quantitative reverse transcription-polymerase chain reaction revealed that polysome loading remained high for much of the night in the cytosol, was strongly light dependent in the plastid, and was always high in mitochondria. The rosette sucrose content correlated with overall and with cytosolic polysome loading. Ribosome abundance did not show significant diurnal changes. However, compared with Col-0, pgm had decreased and increased abundance of plastidic and mitochondrial ribosomes, respectively. Incorporation of label from 13CO2 into protein confirmed that protein synthesis continues at a diminished rate in the dark. Modeling revealed that a decrease in polysome loading at night is required to balance protein synthesis with the availability of carbon from starch breakdown. Costs are also reduced by using amino acids that accumulated in the previous light period. These results uncover a tight coordination of protein synthesis with the momentary supply of carbon. PMID:23674104

  10. Chemically Induced Conditional Rescue of the Reduced Epidermal Fluorescence8 Mutant of Arabidopsis Reveals Rapid Restoration of Growth and Selective Turnover of Secondary Metabolite Pools1[C][OPEN

    PubMed Central

    Kim, Jeong Im; Ciesielski, Peter N.; Donohoe, Bryon S.; Chapple, Clint; Li, Xu

    2014-01-01

    The phenylpropanoid pathway is responsible for the biosynthesis of diverse and important secondary metabolites including lignin and flavonoids. The reduced epidermal fluorescence8 (ref8) mutant of Arabidopsis (Arabidopsis thaliana), which is defective in a lignin biosynthetic enzyme p-coumaroyl shikimate 3′-hydroxylase (C3′H), exhibits severe dwarfism and sterility. To better understand the impact of perturbation of phenylpropanoid metabolism on plant growth, we generated a chemically inducible C3′H expression construct and transformed it into the ref8 mutant. Application of dexamethasone to these plants greatly alleviates the dwarfism and sterility and substantially reverses the biochemical phenotypes of ref8 plants, including the reduction of lignin content and hyperaccumulation of flavonoids and p-coumarate esters. Induction of C3′H expression at different developmental stages has distinct impacts on plant growth. Although early induction effectively restored the elongation of primary inflorescence stem, application to 7-week-old plants enabled them to produce new rosette inflorescence stems. Examination of hypocotyls of these plants revealed normal vasculature in the newly formed secondary xylem, presumably restoring water transport in the mutant. The ref8 mutant accumulates higher levels of salicylic acid than the wild type, but depletion of this compound in ref8 did not relieve the mutant’s growth defects, suggesting that the hyperaccumulation of salicylic acid is unlikely to be responsible for dwarfism in this mutant. PMID:24381065

  11. Amino acid transporter mutants of Arabidopsis provides evidence that a non-mycorrhizal plant acquires organic nitrogen from agricultural soil.

    PubMed

    Ganeteg, Ulrika; Ahmad, Iftikhar; Jämtgård, Sandra; Aguetoni-Cambui, Camila; Inselsbacher, Erich; Svennerstam, Henrik; Schmidt, Susanne; Näsholm, Torgny

    2017-03-01

    Although organic nitrogen (N) compounds are ubiquitous in soil solutions, their potential role in plant N nutrition has been questioned. We performed a range of experiments on Arabidopsis thaliana genetically modified to enhance or reduce root uptake of amino acids. Plants lacking expression of the Lysine Histidine Transporter 1 (LHT1) displayed significantly lower contents of (13) C and (15) N label and of U-(13) C5 ,(15) N2 L-glutamine, as determined by liquid chromatography-mass spectrometry when growing in pots and supplied with dually labelled L-glutamine compared to wild type plants and LHT1-overexpressing plants. Slopes of regressions between accumulation of (13) C-labelled carbon and (15) N-labelled N were higher for LHT1-overexpressing plants than wild type plants, while plants lacking expression of LHT1 did not display a significant regression between the two isotopes. Uptake of labelled organic N from soil tallied with that of labelled ammonium for wild type plants and LHT1-overexpressing plants but was significantly lower for plants lacking expression of LHT1. When grown on agricultural soil plants lacking expression of LHT1 had the lowest, and plants overexpressing LHT1 the highest C/N ratios and natural δ(15) N abundance suggesting their dependence on different N pools. Our data show that LHT1 expression is crucial for plant uptake of organic N from soil.

  12. Null Mutants of Individual RABA Genes Impact the Proportion of Different Cell Wall Components in Stem Tissue of Arabidopsis thaliana

    PubMed Central

    Lunn, Daniel; Gaddipati, Sanyasi R.; Tucker, Gregory A.; Lycett, Grantley W.

    2013-01-01

    In Arabidopsis, and other plants, the RABA GTPases (orthologous to the Rab11a of mammals) have expanded in number and diversity and have been shown to belong to eight sub clades, some of which have been implicated in controlling vesicles that traffic cell wall polymers and enzymes that synthesise or modify them to the cell wall. In order to investigate this, we have investigated whether T-DNA insertion knockouts of individual RABA genes belonging to different sub clades, impact on the composition of the plant cell wall. Single gene knockouts of the RABA1, RABA2 and RABA4 sub clades primarily affected the percentage composition of pectin, cellulose and hemicellulose within the cell wall, respectively, despite having no obvious phenotype in the whole plant. We hypothesise that vesicles carrying specific types of cargoes from the Golgi to the cell surface may be regulated by particular sub types of RABA proteins, a finding that could have wider implications for how trafficking systems work and could be a useful tool in cell wall research and other fields of plant biology. PMID:24124508

  13. Insights into the Composition and Assembly of the Membrane Arm of Plant Complex I through Analysis of Subcomplexes in Arabidopsis Mutant Lines*

    PubMed Central

    Meyer, Etienne H.; Solheim, Cory; Tanz, Sandra K.; Bonnard, Géraldine; Millar, A. Harvey

    2011-01-01

    NADH-ubiquinone oxidoreductase (Complex I, EC 1.6.5.3) is the largest complex of the mitochondrial respiratory chain. In eukaryotes, it is composed of more than 40 subunits that are encoded by both the nuclear and mitochondrial genomes. Plant Complex I differs from the enzyme described in other eukaryotes, most notably due to the large number of plant-specific subunits in the membrane arm of the complex. The elucidation of the assembly pathway of Complex I has been a long-standing research aim in cellular biochemistry. We report the study of Arabidopsis mutants in Complex I subunits using a combination of Blue-Native PAGE and immunodetection to identify stable subcomplexes containing Complex I components, along with mass spectrometry analysis of Complex I components in membrane fractions and two-dimensional diagonal Tricine SDS-PAGE to study the composition of the largest subcomplex. Four subcomplexes of the membrane arm of Complex I with apparent molecular masses of 200, 400, 450, and 650 kDa were observed. We propose a working model for the assembly of the membrane arm of Complex I in plants and assign putative roles during the assembly process for two of the subunits studied. PMID:21606486

  14. A role for nonsense-mediated mRNA decay in plants: pathogen responses are induced in Arabidopsis thaliana NMD mutants.

    PubMed

    Rayson, Samantha; Arciga-Reyes, Luis; Wootton, Lucie; De Torres Zabala, Marta; Truman, William; Graham, Neil; Grant, Murray; Davies, Brendan

    2012-01-01

    Nonsense-mediated mRNA decay (NMD) is a conserved mechanism that targets aberrant mRNAs for destruction. NMD has also been found to regulate the expression of large numbers of genes in diverse organisms, although the biological role for this is unclear and few evolutionarily conserved targets have been identified. Expression analyses of three Arabidopsis thaliana lines deficient in NMD reveal that the vast majority of NMD-targeted transcripts are associated with response to pathogens. Congruently, NMD mutants, in which these transcripts are elevated, confer partial resistance to Pseudomonas syringae. These findings suggest a biological rationale for the regulation of gene expression by NMD in plants and suggest that manipulation of NMD could offer a new approach for crop protection. Amongst the few non-pathogen responsive NMD-targeted genes, one potential NMD targeted signal, the evolutionarily conserved upstream open reading frame (CuORF), was found to be hugely over-represented, raising the possibility that this feature could be used to target specific physiological mRNAs for control by NMD.

  15. Manipulation of Guaiacyl and Syringyl Monomer Biosynthesis in an Arabidopsis Cinnamyl Alcohol Dehydrogenase Mutant Results in Atypical Lignin Biosynthesis and Modified Cell Wall Structure

    PubMed Central

    Anderson, Nickolas A.; Tobimatsu, Yuki; Ciesielski, Peter N.; Ximenes, Eduardo; Ralph, John; Donohoe, Bryon S.; Ladisch, Michael; Chapple, Clint

    2015-01-01

    Modifying lignin composition and structure is a key strategy to increase plant cell wall digestibility for biofuel production. Disruption of the genes encoding both cinnamyl alcohol dehydrogenases (CADs), including CADC and CADD, in Arabidopsis thaliana results in the atypical incorporation of hydroxycinnamaldehydes into lignin. Another strategy to change lignin composition is downregulation or overexpression of ferulate 5-hydroxylase (F5H), which results in lignins enriched in guaiacyl or syringyl units, respectively. Here, we combined these approaches to generate plants enriched in coniferaldehyde-derived lignin units or lignins derived primarily from sinapaldehyde. The cadc cadd and ferulic acid hydroxylase1 (fah1) cadc cadd plants are similar in growth to wild-type plants even though their lignin compositions are drastically altered. In contrast, disruption of CAD in the F5H-overexpressing background results in dwarfism. The dwarfed phenotype observed in these plants does not appear to be related to collapsed xylem, a hallmark of many other lignin-deficient dwarf mutants. cadc cadd, fah1 cadc cadd, and cadd F5H-overexpressing plants have increased enzyme-catalyzed cell wall digestibility. Given that these CAD-deficient plants have similar total lignin contents and only differ in the amounts of hydroxycinnamaldehyde monomer incorporation, these results suggest that hydroxycinnamaldehyde content is a more important determinant of digestibility than lignin content. PMID:26265762

  16. Identification and characterization of an Arabidopsis mutant with altered localization of NIP5;1, a plasma membrane boric acid channel, reveals the requirement for D-galactose in endomembrane organization.

    PubMed

    Uehara, Masataka; Wang, Sheliang; Kamiya, Takehiro; Shigenobu, Shuji; Yamaguchi, Katsushi; Fujiwara, Toru; Naito, Satoshi; Takano, Junpei

    2014-04-01

    Endomembrane organization is important for various aspects of cell physiology, including membrane protein trafficking. To explore the molecular mechanisms regulating the trafficking of plasma membrane-localized proteins in plants, we screened for Arabidopsis mutants with defective localization of green fluorescent protein (GFP)-nodulin 26-like intrinsic protein (NIP)5;1. Fluorescence imaging-based screening led to the isolation of a mutant which accumulated abnormal intracellular aggregates labeled by GFP-NIP5;1. The aggregates appeared in epidermal cells in the root elongation zone and included the trans-Golgi network/early endosomes. Rough mapping and whole-genome sequencing identified the mutant as an allele of UDP-glucose 4-epimerase 4 (uge4)/root hair defective 1 (rhd1) /root epidermal bulgar 1 (reb 1), which was originally defined as a cell wall mutant. The responsible gene encodes UDP-glucose 4-epimerase 4 (UGE4), which functions in the biosynthesis of d-galactose, especially for the synthesis of the cell wall polysaccharide xyloglucan and arabinogalactan proteins (AGPs). The endomembrane aggregates in the mutants were absent in the presence of d-galactose, indicative of a requirement for a d-galactose-containing component in endomembrane organization. Genetic and pharmacological analyses suggested that the aggregates were not caused by the disruption of cell wall polysaccharides or the cytoskeleton. Overall, our results suggest that UGE4 activity in d-galactose synthesis is required for the structure of cell wall polysaccharides and endomembrane organization.

  17. Possible Role of MADS AFFECTING FLOWERING 3 and B-BOX DOMAIN PROTEIN 19 in Flowering Time Regulation of Arabidopsis Mutants with Defects in Nonsense-Mediated mRNA Decay

    PubMed Central

    Nasim, Zeeshan; Fahim, Muhammad; Ahn, Ji Hoon

    2017-01-01

    Eukaryotic cells use nonsense-mediated mRNA decay (NMD) to clear aberrant mRNAs from the cell, thus preventing the accumulation of truncated proteins. In Arabidopsis, two UP-Frameshift (UPF) proteins, UPF1 and UPF3, play a critical role in NMD. Although deficiency of UPF1 and UPF3 leads to various developmental defects, little is known about the mechanism underlying the regulation of flowering time by NMD. Here, we showed that the upf1-5 and upf3-1 mutants had a late-flowering phenotype under long-day conditions and the upf1-5 upf3-1 double mutants had an additive effect in delaying flowering time. RNA sequencing of the upf mutants revealed that UPF3 exerted a stronger effect than UPF1 in the UPF-mediated regulation of flowering time. Among genes known to regulate flowering time, FLOWERING LOCUS C (FLC) mRNA levels increased (up to 8-fold) in upf mutants, as confirmed by qPCR. The upf1-5, upf3-1, and upf1-5 upf3-1 mutants responded to vernalization, suggesting a role of FLC in delayed flowering of upf mutants. Consistent with the high FLC transcript levels and delayed flowering in upf mutants, levels of FLOWERING LOCUS T (FT) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) mRNAs were reduced in the upf mutants. However, RNA-seq did not identify an aberrant FLC transcript containing a premature termination codon (PTC), suggesting that FLC is not a direct target in the regulation of flowering time by NMD. Among flowering time regulators that act in an FLC-dependent manner, we found that MAF3, NF-YA2, NF-YA5, and TAF14 showed increased transcript levels in upf mutants. We also found that BBX19 and ATC, which act in an FLC-independent manner, showed increased transcript levels in upf mutants. An aberrant transcript containing a PTC was identified from MAF3 and BBX19 and the levels of the aberrant transcripts increased in upf mutants. Taking these results together, we propose that the late-flowering phenotype of upf mutants is mediated by at least two different

  18. Possible Role of MADS AFFECTING FLOWERING 3 and B-BOX DOMAIN PROTEIN 19 in Flowering Time Regulation of Arabidopsis Mutants with Defects in Nonsense-Mediated mRNA Decay.

    PubMed

    Nasim, Zeeshan; Fahim, Muhammad; Ahn, Ji Hoon

    2017-01-01

    Eukaryotic cells use nonsense-mediated mRNA decay (NMD) to clear aberrant mRNAs from the cell, thus preventing the accumulation of truncated proteins. In Arabidopsis, two UP-Frameshift (UPF) proteins, UPF1 and UPF3, play a critical role in NMD. Although deficiency of UPF1 and UPF3 leads to various developmental defects, little is known about the mechanism underlying the regulation of flowering time by NMD. Here, we showed that the upf1-5 and upf3-1 mutants had a late-flowering phenotype under long-day conditions and the upf1-5 upf3-1 double mutants had an additive effect in delaying flowering time. RNA sequencing of the upf mutants revealed that UPF3 exerted a stronger effect than UPF1 in the UPF-mediated regulation of flowering time. Among genes known to regulate flowering time, FLOWERING LOCUS C (FLC) mRNA levels increased (up to 8-fold) in upf mutants, as confirmed by qPCR. The upf1-5, upf3-1, and upf1-5 upf3-1 mutants responded to vernalization, suggesting a role of FLC in delayed flowering of upf mutants. Consistent with the high FLC transcript levels and delayed flowering in upf mutants, levels of FLOWERING LOCUS T (FT) and SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) mRNAs were reduced in the upf mutants. However, RNA-seq did not identify an aberrant FLC transcript containing a premature termination codon (PTC), suggesting that FLC is not a direct target in the regulation of flowering time by NMD. Among flowering time regulators that act in an FLC-dependent manner, we found that MAF3, NF-YA2, NF-YA5, and TAF14 showed increased transcript levels in upf mutants. We also found that BBX19 and ATC, which act in an FLC-independent manner, showed increased transcript levels in upf mutants. An aberrant transcript containing a PTC was identified from MAF3 and BBX19 and the levels of the aberrant transcripts increased in upf mutants. Taking these results together, we propose that the late-flowering phenotype of upf mutants is mediated by at least two different

  19. Characterization of multiple SPS knockout mutants reveals redundant functions of the four Arabidopsis sucrose phosphate synthase isoforms in plant viability, and strongly indicates that enhanced respiration and accelerated starch turnover can alleviate the blockage of sucrose biosynthesis.

    PubMed

    Bahaji, Abdellatif; Baroja-Fernández, Edurne; Ricarte-Bermejo, Adriana; Sánchez-López, Ángela María; Muñoz, Francisco José; Romero, Jose M; Ruiz, María Teresa; Baslam, Marouane; Almagro, Goizeder; Sesma, María Teresa; Pozueta-Romero, Javier

    2015-09-01

    We characterized multiple knock-out mutants of the four Arabidopsis sucrose phosphate synthase (SPSA1, SPSA2, SPSB and SPSC) isoforms. Despite their reduced SPS activity, spsa1/spsa2, spsa1/spsb, spsa2/spsb, spsa2/spsc, spsb/spsc, spsa1/spsa2/spsb and spsa2/spsb/spsc mutants displayed wild type (WT) vegetative and reproductive morphology, and showed WT photosynthetic capacity and respiration. In contrast, growth of rosettes, flowers and siliques of the spsa1/spsc and spsa1/spsa2/spsc mutants was reduced compared with WT plants. Furthermore, these plants displayed a high dark respiration phenotype. spsa1/spsb/spsc and spsa1/spsa2/spsb/spsc seeds poorly germinated and produced aberrant and sterile plants. Leaves of all viable sps mutants, except spsa1/spsc and spsa1/spsa2/spsc, accumulated WT levels of nonstructural carbohydrates. spsa1/spsc leaves possessed high levels of metabolic intermediates and activities of enzymes of the glycolytic and tricarboxylic acid cycle pathways, and accumulated high levels of metabolic intermediates of the nocturnal starch-to-sucrose conversion process, even under continuous light conditions. Results presented in this work show that SPS is essential for plant viability, reveal redundant functions of the four SPS isoforms in processes that are important for plant growth and nonstructural carbohydrate metabolism, and strongly indicate that accelerated starch turnover and enhanced respiration can alleviate the blockage of sucrose biosynthesis in spsa1/spsc leaves.

  20. The "Arabidopsis cax3" mutants display altered salt tolerance, pH sensitivity and reduced plasma membrane H(+)-ATPase activity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Perturbing CAX1, an "Arabidopsis" vacuolar H(+)/Ca(2+) antiporter, and the related vacuolar transporter CAX3, has been previously shown to cause severe growth defects; however, the specific function of CAX3 has remained elusive. Here, we describe plant phenotypes that are shared among "cax1" and "ca...

  1. Whole-plant mineral partitioning throughout the life cycle in Arabidopsis thaliana ecotypes Columbia, Landsberg erecta, Cape Verde Islands, and the mutant line ysl1ysl3

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Minimal information exists on whole-plant dynamics of mineral flow through Arabidopsis thaliana or on the source tissues responsible for mineral export to developing seeds. Understanding these phenomena in a model plant could help in the development of nutritionally enhanced crop cultivars. A who...

  2. Defective chloroplast development inhibits maintenance of normal levels of abscisic acid in a mutant of the Arabidopsis RH3 DEAD-box protein during early post-germination growth.

    PubMed

    Lee, Kwang-Hee; Park, Jiyoung; Williams, Donna S; Xiong, Yuqing; Hwang, Inhwan; Kang, Byung-Ho

    2013-03-01

    The plastid has its own translation system, and its ribosomes are assembled through a complex process in which rRNA precursors are processed and ribosomal proteins are inserted into the rRNA backbone. DEAD-box proteins have been shown to play roles in multiple steps in ribosome biogenesis. To investigate the cellular and physiological roles of an Arabidopsis DEAD-box protein, RH3, we examined its expression and localization and the phenotypes of rh3-4, a T-DNA insertion mutant allele of RH3. The promoter activity of RH3 is strongest in the greening tissues of 3-day and 1-week-old seedlings but reduced afterwards. Cotyledons were pale and seedling growth was retarded in the mutant. The most obvious abnormality in the mutant chloroplasts was their lack of normal ribosomes. Electron tomography analysis indicated that ribosome density in the 3-day-old mutant chloroplasts is only 20% that of wild-type chloroplasts, and the ribosomes in the mutant are smaller. These chloroplast defects in rh3-4 were alleviated in 2-week-old cotyledons and true leaves. Interestingly, rh3-4 seedlings have lower amounts of abscisic acid prior to recovery of their chloroplasts, and were more sensitive to abiotic stresses. Transcriptomic analysis indicated that nuclear genes for chloroplast proteins are down-regulated, and proteins mediating chloroplast-localized steps of abscisic acid biosynthesis are expressed to a lower extent in 1-week-old rh3-4 seedlings. Taken together, these results suggest that conversion of eoplasts into chloroplasts in young seedlings is critical for the seedlings to start carbon fixation as well as for maintenance of abscisic acid levels for responding to environmental challenges.

  3. Genetic Screening for Arabidopsis Mutants Defective in STA1 Regulation under Thermal Stress Implicates the Existence of Regulators of Its Specific Expression, and the Genetic Interactions in the Stress Signaling Pathways.

    PubMed

    Yu, Si-In; Han, Jin-Hee; Chhoeun, Chanvotey; Lee, Byeong-Ha

    2016-01-01

    To cope with environmental stresses, plants have developed various stress tolerance mechanisms that involve the induction of many stress responsive genes through stress-specific and common signaling pathways. Stress-specific/common transcription factors, rather than general basal factors, were considered important in this stress tolerance. The Arabidopsis STABILIZED1 (STA1) gene encodes a putative pre-mRNA splicing factor that is similar to the human U5 snRNP-associated 102-kDa protein and the yeast pre-mRNA splicing factors, PRP1p and Prp6p. As pre-mRNA splicing is a necessary process for proper gene expression in eukaryotes, STA1 is expected to be constantly functional in all conditions. Interestingly, STA1 expression is induced by temperature stresses, and STA1 recessive mutation (sta1-1) resulted in temperature stress-specific hypersensitivity. This suggests STA1's stress specific function in addition to its presumed "housekeeping" role. In order to establish the genetic system to understand the regulation of STA1 expression in temperature stresses, we generated a bioluminescent Arabidopsis plant harboring the STA1 promoter fused to the firefly luciferase coding sequence (STA1p-LUC). Through genetic analysis, the bioluminescent Arabidopsis homozygous for one-copy STA1p-LUC was isolated and characterized. In this STA1p-LUC line, the expression patterns of STA1p-LUC were similar to those of the endogenous STA1 gene under cold and heat stresses. The STA1p-LUC line was then chemically mutagenized and screened to isolate the genetic loci of STA1 regulators under cold or heat stresses. Mutants with altered STA1p-LUC luminescence were identified and further confirmed through luminescence imaging in the next generation and analysis of endogenous STA1 expression. The categorization of STA1p-LUC deregulated mutants implicated the existence of cold or heat stress-specific as well as common genetic regulators for STA1 expression. Interestingly, some mutants showed opposite

  4. S-nitrosoglutathione promotes cell wall remodelling, alters the transcriptional profile and induces root hair formation in the hairless root hair defective 6 (rhd6) mutant of Arabidopsis thaliana.

    PubMed

    Moro, Camila Fernandes; Gaspar, Marilia; da Silva, Felipe Rodrigues; Pattathil, Sivakumar; Hahn, Michael G; Salgado, Ione; Braga, Marcia Regina

    2017-03-01

    Nitric oxide (NO) exerts pleiotropic effects on plant development; however, its involvement in cell wall modification during root hair formation (RHF) has not yet been addressed. Here, mutants of Arabidopsis thaliana with altered root hair phenotypes were used to assess the involvement of S-nitrosoglutathione (GSNO), the primary NO source, in cell wall dynamics and gene expression in roots induced to form hairs. GSNO and auxin restored the root hair phenotype of the hairless root hair defective 6 (rhd6) mutant. A positive correlation was observed between increased NO production and RHF induced by auxin in rhd6 and transparent testa glabra (ttg) mutants. Deposition of an epitope within rhamnogalacturonan-I recognized by the CCRC-M2 antibody was delayed in root hair cells (trichoblasts) compared with nonhair cells (atrichoblasts). GSNO, but not auxin, restored the wild-type root glycome and transcriptome profiles in rhd6, modulating the expression of a large number of genes related to cell wall composition and metabolism, as well as those encoding ribosomal proteins, DNA and histone-modifying enzymes and proteins involved in post-translational modification. Our results demonstrate that NO plays a key role in cell wall remodelling in trichoblasts and suggest that it also participates in chromatin modification in root cells of A. thaliana.

  5. Isolation and characterization of the Arabidopsis heat-intolerant 2 (hit2) mutant reveal the essential role of the nuclear export receptor EXPORTIN1A (XPO1A) in plant heat tolerance.

    PubMed

    Wu, Shin-Jye; Wang, Lian-Chin; Yeh, Ching-Hui; Lu, Chun-An; Wu, Shaw-Jye

    2010-06-01

    *The Arabidopsis heat-intolerant 2 (hit2) mutant was isolated on the basis of its impaired ability to withstand moderate heat stress (37 degrees C). Determination of the genetic mutation that underlies the hit2 thermosensitive phenotype allowed better understanding of the mechanisms by which plants cope with heat stress. *Genetic analysis revealed that hit2 is a single recessive mutation. Map-based cloning was used to identify the hit2 locus. The response of hit2 to other types of heat stress was also investigated to characterize the protective role of HIT2. *hit2 was defective in basal but not in acquired thermotolerance. hit2 was sensitive to methyl viologen-induced oxidative stress, and the survival of hit2 seedlings in response to heat stress was affected by light conditions. The mutated locus was located at the EXPORTIN1A (XPO1A) gene, which encodes a nuclear transport receptor. Two T-DNA insertion lines, xpo1a-1 and xpo1a-3, exhibited the same phenotypes as hit2. *The results provide evidence that Arabidopsis XPO1A is dispensable for normal plant growth and development but is essential for thermotolerance, in part by mediating the protection of plants against heat-induced oxidative stress.

  6. CYP77A19 and CYP77A20 characterized from Solanum tuberosum oxidize fatty acids in vitro and partially restore the wild phenotype in an Arabidopsis thaliana cutin mutant.

    PubMed

    Grausem, B; Widemann, E; Verdier, G; Nosbüsch, D; Aubert, Y; Beisson, F; Schreiber, L; Franke, R; Pinot, F

    2014-09-01

    Cutin and suberin represent lipophilic polymers forming plant/environment interfaces in leaves and roots. Despite recent progress in Arabidopsis, there is still a lack on information concerning cutin and suberin synthesis, especially in crops. Based on sequence homology, we isolated two cDNA clones of new cytochrome P450s, CYP77A19 and CYP77A20 from potato tubers (Solanum tuberosum). Both enzymes hydroxylated lauric acid (C12:0) on position ω-1 to ω-5. They oxidized fatty acids with chain length ranging from C12 to C18 and catalysed hydroxylation of 16-hydroxypalmitic acid leading to dihydroxypalmitic (DHP) acids, the major C16 cutin and suberin monomers. CYP77A19 also produced epoxides from linoleic acid (C18:2). Exploration of expression pattern in potato by RT-qPCR revealed the presence of transcripts in all tissues tested with the highest expression in the seed compared with leaves. Water stress enhanced their expression level in roots but not in leaves. Application of methyl jasmonate specifically induced CYP77A19 expression. Expression of either gene in the Arabidopsis null mutant cyp77a6-1 defective in flower cutin restored petal cuticular impermeability. Nanoridges were also observed in CYP77A20-expressing lines. However, only very low levels of the major flower cutin monomer 10,16-dihydroxypalmitate and no C18 epoxy monomers were found in the cutin of the complemented lines.

  7. SnRK2.6/OST1 from Arabidopsis thaliana: cloning, expression, purification, crystallization and preliminary X-ray analysis of K50N and D160A mutants

    PubMed Central

    Yunta, Cristina; Martinez-Ripoll, Martin; Albert, Armando

    2011-01-01

    The SnRK2.6 (SNF1-related kinase 2.6) gene from Arabidopsis thaliana encodes the serine/threonine protein kinase SnRK2.6/OST1 (OPEN STOMATA 1). It plays a central role in the drought-tolerance mechanism. OST1 is in fact the main positive effector in the hydric stress response. The SnRK2.6 gene was cloned into the pGEX4T1 plasmid, mutated and expressed in Escherichia coli, allowing purification to homogeneity in two chromatographic steps. Various OST1 mutants yielded crystals using vapour-diffusion techniques, but only one mutant showed a good diffraction pattern. Its crystals diffracted to 2.8 Å resolution and belonged to space group P2221, with unit-cell parameters a = 77.7, b = 99.4, c = 108.4 Å. A promising molecular-replacement solution was found using the structure of the kinase domain of the yeast AMP-activated protein kinase SNF1 (PDB entry 3hyh) as the search model. PMID:21393844

  8. Ectopic expression of CsCTR1, a cucumber CTR-like gene, attenuates constitutive ethylene signaling in an Arabidopsis ctr1-1 mutant and expression pattern analysis of CsCTR1 in cucumber (Cucumis sativus).

    PubMed

    Bie, Beibei; Sun, Jin; Pan, Junsong; He, Huanle; Cai, Run

    2014-09-15

    The gaseous plant hormone ethylene regulates many aspects of plant growth, development and responses to the environment. Constitutive triple response 1 (CTR1) is a central regulator involved in the ethylene signal transduction pathway. To obtain a better understanding of this particular pathway in cucumber, the cDNA-encoding CTR1 (designated CsCTR1) was isolated from cucumber. A sequence alignment and phylogenetic analyses revealed that CsCTR1 has a high degree of homology with other plant CTR1 proteins. The ectopic expression of CsCTR1 in the Arabidopsis ctr1-1 mutant attenuates constitutive ethylene signaling of this mutant, suggesting that CsCTR1 indeed performs its function as negative regulator of the ethylene signaling pathway. CsCTR1 is constitutively expressed in all of the examined cucumber organs, including roots, stems, leaves, shoot apices, mature male and female flowers, as well as young fruits. CsCTR1 expression gradually declined during male flower development and increased during female flower development. Additionally, our results indicate that CsCTR1 can be induced in the roots, leaves and shoot apices by external ethylene. In conclusion, this study provides a basis for further studies on the role of CTR1 in the biological processes of cucumber and on the molecular mechanism of the cucumber ethylene signaling pathway.

  9. Effects of natural and synthetic auxins on the gravitropic growth habit of roots in two auxin-resistant mutants of Arabidopsis, axr1 and axr4: evidence for defects in the auxin influx mechanism of axr4

    NASA Technical Reports Server (NTRS)

    Yamamoto, M.; Yamamoto, K. T.

    1999-01-01

    The partially agravitropic growth habit of roots of an auxin-resistant mutant of Arabidopsis thaliana, axr4, was restored by the addition of 30-300 nM 1-naphthaleneacetic acid (NAA) to the growth medium. Neither indole 3-acetic acid (IAA) nor 2,4-dichlorophenoxyacetic acid (2,4-D) showed such an effect. Growth of axr4 roots was resistant to IAA and 2,4-D, but not at all to NAA. The differential effects of the three auxins suggest that the defects of axr4 result from a lower auxin influx into its cells. The partially agravitropic growth habit of axr1 roots, which was less severe than that of axr4 roots, was only slightly affected by the three auxins in the growth medium at concentrations up to 300 nM; growth of axr1 roots was resistant to all three of the auxins. These results suggest that the lesion of axrl mutants is different from that of axr4.

  10. Ectopic Expression of CsCTR1, a Cucumber CTR-Like Gene, Attenuates Constitutive Ethylene Signaling in an Arabidopsis ctr1-1 Mutant and Expression Pattern Analysis of CsCTR1 in Cucumber (Cucumis sativus)

    PubMed Central

    Bie, Beibei; Sun, Jin; Pan, Junsong; He, Huanle; Cai, Run

    2014-01-01

    The gaseous plant hormone ethylene regulates many aspects of plant growth, development and responses to the environment. Constitutive triple response 1 (CTR1) is a central regulator involved in the ethylene signal transduction pathway. To obtain a better understanding of this particular pathway in cucumber, the cDNA-encoding CTR1 (designated CsCTR1) was isolated from cucumber. A sequence alignment and phylogenetic analyses revealed that CsCTR1 has a high degree of homology with other plant CTR1 proteins. The ectopic expression of CsCTR1 in the Arabidopsis ctr1-1 mutant attenuates constitutive ethylene signaling of this mutant, suggesting that CsCTR1 indeed performs its function as negative regulator of the ethylene signaling pathway. CsCTR1 is constitutively expressed in all of the examined cucumber organs, including roots, stems, leaves, shoot apices, mature male and female flowers, as well as young fruits. CsCTR1 expression gradually declined during male flower development and increased during female flower development. Additionally, our results indicate that CsCTR1 can be induced in the roots, leaves and shoot apices by external ethylene. In conclusion, this study provides a basis for further studies on the role of CTR1 in the biological processes of cucumber and on the molecular mechanism of the cucumber ethylene signaling pathway. PMID:25226540

  11. Sequence and characterization of two Arabidopsis thaliana cDNAs isolated by functional complementation of a yeast gln3 gdh1 mutant.

    PubMed

    Truong, H N; Caboche, M; Daniel-Vedele, F

    1997-06-30

    We have isolated two Arabidopsis thaliana cDNAs by complementation of a yeast gln3 gdh1 strain that is affected in the regulation of nitrogen metabolism. The two clones (RGA1 and RGA2) are homologous to each other and to the SCARECROW (SCR) gene that is involved in regulating an asymmetric cell division in plants. RGA1, RGA2 and SCR share several structural features and may define a new family of genes. RGA1 and RGA2 have been mapped, respectively, to chromosome II and I, and their expression in plant is constitutive.

  12. Altered Profile of Secondary Metabolites in the Root Exudates of Arabidopsis ATP-Binding Cassette Transporter Mutants1[C][W][OA

    PubMed Central

    Badri, Dayakar V.; Loyola-Vargas, Victor M.; Broeckling, Corey D.; De-la-Peña, Clelia; Jasinski, Michal; Santelia, Diana; Martinoia, Enrico; Sumner, Lloyd W.; Banta, Lois M.; Stermitz, Frank; Vivanco, Jorge M.

    2008-01-01

    Following recent indirect evidence suggesting a role for ATP-binding cassette (ABC) transporters in root exudation of phytochemicals, we identified 25 ABC transporter genes highly expressed in the root cells most likely to be involved in secretion processes. Of these 25 genes, we also selected six full-length ABC transporters and a half-size transporter for in-depth molecular and biochemical analyses. We compared the exuded root phytochemical profiles of these seven ABC transporter mutants to those of the wild type. There were three nonpolar phytochemicals missing in various ABC transporter mutants compared to the wild type when the samples were analyzed by high-performance liquid chromatography-mass spectrometry. These data suggest that more than one ABC transporter can be involved in the secretion of a given phytochemical and that a transporter can be involved in the secretion of more than one secondary metabolite. The primary and secondary metabolites present in the root exudates of the mutants were also analyzed by gas chromatography-mass spectrometry, which allowed for the identification of groups of compounds differentially found in some of the mutants compared to the wild type. For instance, the mutant Atpdr6 secreted a lower level of organic acids and Atmrp2 secreted a higher level of amino acids as compared to the wild type. We conclude that the release of phytochemicals by roots is partially controlled by ABC transporters. PMID:18065561

  13. Isolation of a mutant Arabidopsis plant that lacks N-aetyl glucosaminyl transferase I and is unable to synthesize Golgi-modified complex N-linked glycans

    SciTech Connect

    Schaewen, A. von; O'Neill, J.; Chrispeels, M.J. ); Sturm, A. )

    1993-08-01

    The complex asparagine-linked glycans of plant glycoproteins, characterized by the presence of [beta]1[yields]2 xylose and [alpha]1[yields]3 fucose residues, are derived from typical mannose[sub 9](N-acetylglucosamine)[sub 2] (Man[sub 9]GlcNAc[sub 2]) N-linked glycans through the activity of a series of glycosidases and glycosyl transferases in the Golgi apparatus. By screening leaf extracts with an antiserum against complex glycans, we isolated a mutant of Arbidopsis thaliana that is blocked in the conversion of high-manne to complex glycans. In callus tissues derived from the mutant plants, all glycans bind to concanavalin A. These glycans can be released by treatment with endoglycosidase H, and the majority has the same size as Man[sub 5]GlcNAc[sub 1] glycans. In the presence of deoxymannojirimycin, an inhibitor of mannosidase I, the mutant cells synthesize Man[sub 9]GlcNAc[sub 2] and Man[sub 8]GlcNAc[sub 2] glycans, suggesting that the biochemical lesion in the mutant is not in the biosynthesis of high-mannose glycans in the endoplasmic reticulum but in their modification in the Golgi. Direct enzyme assays of cell extracts show that the mutant cells lack N-acetyl glucosaminyl transferase I, the first enzyme in the pathway of complex glycan biosynthesis. The mutant plants are able to complete their development normally under several environmental conditions, suggesting that complex glycans are not essential for normal developmental processes. By crossing the complex-glycan-deficient strain of A. thaliana with a transgenic strain that expresses the glycoprotein phytohemagglutinin, a unique strain was obtained that synthesizes phytohemagglutinin with two high-mannose glycans, instead of one high-mannose and one complex glycan. 42 refs., 8 figs., 1 tab.

  14. Elevated salicylic acid levels conferred by increased expression of ISOCHORISMATE SYNTHASE 1 contribute to hyperaccumulation of SUMO1 conjugates in the Arabidopsis mutant early in short days 4.

    PubMed

    Villajuana-Bonequi, Mitzi; Elrouby, Nabil; Nordström, Karl; Griebel, Thomas; Bachmair, Andreas; Coupland, George

    2014-07-01

    Post-translational modification of proteins by attachment of small ubiquitin-like modifier (SUMO) is essential for plant growth and development. Mutations in the SUMO protease early in short days 4 (ESD4) cause hyperaccumulation of conjugates formed between SUMO and its substrates, and phenotypically are associated with extreme early flowering and impaired growth. We performed a suppressor mutagenesis screen of esd4 and identified a series of mutants called suppressor of esd4 (sed), which delay flowering, enhance growth and reduce hyperaccumulation of SUMO conjugates. Genetic mapping and genome sequencing indicated that one of these mutations (sed111) is in the gene salicylic acid induction-deficient 2 (SID2), which encodes ISOCHORISMATE SYNTHASE I, an enzyme required for biosynthesis of salicylic acid (SA). Analyses showed that compared with wild-type plants, esd4 contains higher levels of SID2 mRNA and about threefold more SA, whereas sed111 contains lower SA levels. Other sed mutants also contain lower SA levels but are not mutant for SID2, although most reduce SID2 mRNA levels. Therefore, higher SA levels contribute to the small size, early flowering and elevated SUMO conjugate levels of esd4. Our results support previous data indicating that SUMO homeostasis influences SA biosynthesis in wild-type plants, and also demonstrate that elevated levels of SA strongly increase the abundance of SUMO conjugates.

  15. The turnip Mutant of Arabidopsis Reveals That LEAFY COTYLEDON1 Expression Mediates the Effects of Auxin and Sugars to Promote Embryonic Cell Identity1[W

    PubMed Central

    Casson, Stuart A.; Lindsey, Keith

    2006-01-01

    The transition from embryonic to vegetative growth marks an important developmental stage in the plant life cycle. The turnip (tnp) mutant was identified in a screen for modifiers of POLARIS expression, a gene required for normal root growth. Mapping and molecular characterization of tnp shows that it represents a gain-of-function mutant of LEAFY COTYLEDON1 (LEC1), due to a promoter mutation. This results in the ectopic expression of LEC1, but not of other LEC genes, in vegetative tissues. The LEC class of genes are known regulators of embryogenesis, involved in the control of embryonic cell identity by currently unknown mechanisms. Activation of the LEC-dependent pathway in tnp leads to the loss of hypocotyl epidermal cell marker expression and loss of SCARECROW expression in the endodermis, the ectopic accumulation of starch and lipids, and the up-regulation of early and late embryonic genes. tnp also shows partial deetiolation during dark growth. Penetrance of the mutant phenotype is strongly enhanced in the presence of exogenous auxin and sugars, but not by gibberellin or abscisic acid, and is antagonized by cytokinin. We propose that the role of LEC1 in embryonic cell fate control requires auxin and sucrose to promote cell division and embryonic differentiation. PMID:16935993

  16. Transcriptional profiling of an Fd-GOGAT1/GLU1 mutant in Arabidopsis thaliana reveals a multiple stress response and extensive reprogramming of the transcriptome

    PubMed Central

    2010-01-01

    Background Glutamate plays a central position in the synthesis of a variety of organic molecules in plants and is synthesised from nitrate through a series of enzymatic reactions. Glutamate synthases catalyse the last step in this pathway and two types are present in plants: NADH- or ferredoxin-dependent. Here we report a genome wide microarray analysis of the transcriptional reprogramming that occurs in leaves and roots of the A. thaliana mutant glu1-2 knocked-down in the expression of Fd-GOGAT1 (GLU1; At5g04140), one of the two genes of A. thaliana encoding ferredoxin-dependent glutamate synthase. Results Transcriptional profiling of glu1-2 revealed extensive changes with the expression of more than 5500 genes significantly affected in leaves and nearly 700 in roots. Both genes involved in glutamate biosynthesis and transformation are affected, leading to changes in amino acid compositions as revealed by NMR metabolome analysis. An elevated glutamine level in the glu1-2 mutant was the most prominent of these changes. An unbiased analysis of the gene expression datasets allowed us to identify the pathways that constitute the secondary response of an FdGOGAT1/GLU1 knock-down. Among the most significantly affected pathways, photosynthesis, photorespiratory cycle and chlorophyll biosynthesis show an overall downregulation in glu1-2 leaves. This is in accordance with their slight chlorotic phenotype. Another characteristic of the glu1-2 transcriptional profile is the activation of multiple stress responses, mimicking cold, heat, drought and oxidative stress. The change in expression of genes involved in flavonoid biosynthesis is also revealed. The expression of a substantial number of genes encoding stress-related transcription factors, cytochrome P450 monooxygenases, glutathione S-transferases and UDP-glycosyltransferases is affected in the glu1-2 mutant. This may indicate an induction of the detoxification of secondary metabolites in the mutant. Conclusions Analysis

  17. A nuclear mutant of Arabidopsis with impaired stability on distinct transcripts of the plastid psbB, psbD/C, ndhH, and ndhC operons.

    PubMed Central

    Meurer, J; Berger, A; Westhoff, P

    1996-01-01

    The high-chlorophyll fluorescence photosynthesis mutant hcf109 of Arabidopsis was characterized in detail to gain insights into the regulatory mechanism of RNA processing in higher plants. By using electron transport, chlorophyll fluorescence, and immunoblot studies, we assigned the mutational lesion to photosystems I and II and the plastid NAD(P)H dehydrogenase complex. The functional pleiotropy was reflected in RNA deficiencies. Although all nuclear-encoded photosynthetic RNAs analyzed revealed no difference in size or steady state level between mutant and wild type, the RNA patterns of the plastome-encoded psbB-psbT-psbH-petB-petD, psbD-psbC-ycf9, ndhC-ndhK-ndhJ, and ndhH-ndhA-ndhI-ndhG-ndhE-psaC-ndh D transcription units were severely disturbed. These operons encode subunits of photosystems I (psa) and II (psb), the cytochrome bGf complex (pet), the plastid NAD(P)H dehydrogenase (ndh), and the unidentified open reading frame ycf9. With the exception of the ndhC operon, the RNA deficiencies observed were specific and restricted to particular segments of the psbB, psbD/C, and ndhH operons, that is, the psbB-psbT, ycf9, and psaC regions. Run-on transcription studies with isolated chloroplasts showed that the failure of these transcripts to accumulate was due to RNA stability and not transcription. Other polycistronic transcription units analyzed were not affected by the mutation. This result indicates that the trans-regulatory factor encoded by the hcf109 gene is not a general RNA stability factor but that it specifically controls the stability of only these distinct transcripts. Because the hcf109 locus was mapped at a distance < 0.1 centimorgans from the phytochrome C gene, its molecular characterization by positional cloning is possible. PMID:8768377

  18. Arabidopsis thaliana ggt1 photorespiratory mutants maintain leaf carbon/nitrogen balance by reducing RuBisCO content and plant growth.

    PubMed

    Dellero, Younès; Lamothe-Sibold, Marlène; Jossier, Mathieu; Hodges, Michael

    2015-09-01

    Metabolic and physiological analyses of glutamate:glyoxylate aminotransferase 1 (GGT1) mutants were performed at the global leaf scale to elucidate the mechanisms involved in their photorespiratory growth phenotype. Air-grown ggt1 mutants showed retarded growth and development, that was not observed at high CO2 (3000 μL L(-1) ). When compared to wild-type (WT) plants, air-grown ggt1 plants exhibited glyoxylate accumulation, global changes in amino acid amounts including a decrease in serine content, lower organic acid levels, and modified ATP/ADP and NADP(+) /NADPH ratios. When compared to WT plants, their net CO2 assimilation rates (An ) were 50% lower and this mirrored decreases in ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) contents. High CO2 -grown ggt1 plants transferred to air revealed a rapid decrease of An and photosynthetic electron transfer rate while maintaining a high energetic state. Short-term (a night period and 4 h of light) transferred ggt1 leaves accumulated glyoxylate and exhibited low serine contents, while other amino acid levels were not modified. RuBisCO content, activity and activation state were not altered after a short-term transfer while the ATP/ADP ratio was lowered in ggt1 rosettes. However, plant growth and RuBisCO levels were both reduced in ggt1 leaves after a long-term (12 days) acclimation to air from high CO2 when compared to WT plants. The data are discussed with respect to a reduced photorespiratory carbon recycling in the mutants. It is proposed that the low An limits nitrogen-assimilation, this decreases leaf RuBisCO content until plants attain a new homeostatic state that maintains a constant C/N balance and leads to smaller, slower growing plants.

  19. Studies on Arabidopsis athak5, atakt1 double mutants disclose the range of concentrations at which AtHAK5, AtAKT1 and unknown systems mediate K uptake.

    PubMed

    Rubio, Francisco; Alemán, Fernando; Nieves-Cordones, Manuel; Martínez, Vicente

    2010-06-01

    The high-affinity K(+) transporter AtHAK5 and the inward-rectifier K(+) channel AtAKT1 have been described to contribute to K(+) uptake in Arabidopsis thaliana. Studies with T-DNA insertion lines showed that both systems participate in the high-affinity range of concentrations and only AtAKT1 in the low-affinity range. However the contribution of other systems could not be excluded with the information and plant material available. The results presented here with a double knock-out athak5, atakt1 mutant show that AtHAK5 is the only system mediating K(+) uptake at concentrations below 0.01 mM. In the range between 0.01 and 0.05 mM K(+) AtHAK5 and AtAKT1 are the only contributors to K(+) acquisition. At higher K(+) concentrations, unknown systems come into operation and participate together with AtAKT1 in low-affinity K(+) uptake. These systems can supply sufficient K(+) to promote plant growth even in the absence of AtAKT1 or in the presence of 10 mM K(+) where AtAKT1 is not essential.

  20. Transcript Profiling in the chl1-5 Mutant of Arabidopsis Reveals a Role of the Nitrate Transporter NRT1.1 in the Regulation of Another Nitrate Transporter, NRT2.1W⃞

    PubMed Central

    Muños, Stéphane; Cazettes, Céline; Fizames, Cécile; Gaymard, Frédéric; Tillard, Pascal; Lepetit, Marc; Lejay, Laurence; Gojon, Alain

    2004-01-01

    Arabidopsis thaliana mutants deficient for the NRT1.1 NO3− transporter display complex phenotypes, including lowered NO3− uptake, altered development of nascent organs, and reduced stomatal opening. To obtain further insight at the molecular level on the multiple physiological functions of NRT1.1, we performed large-scale transcript profiling by serial analysis of gene expression in the roots of the chl1-5 deletion mutant of NRT1.1 and of the Columbia wild type. Several hundred genes were differentially expressed between the two genotypes, when plants were grown on NH4NO3 as N source. Among these genes, the N satiety-repressed NRT2.1 gene, encoding a major component of the root high-affinity NO3− transport system (HATS), was found to be strongly derepressed in the chl1-5 mutant (as well as in other NRT1.1 mutants). This was associated with a marked stimulation of the NO3− HATS activity in the mutant, suggesting adaptive response to a possible N limitation resulting from NRT1.1 mutation. However, derepression of NRT2.1 in NH4NO3-fed chl1-5 plants could not be attributed to lowered production of N metabolites. Rather, the results show that normal regulation of NRT2.1 expression is strongly altered in the chl1-5 mutant, where this gene is no more repressible by high N provision to the plant. This indicates that NRT1.1 plays an unexpected but important role in the regulation of both NRT2.1 expression and NO3− HATS activity. Overexpression of NRT2.1 was also found in wild-type plants supplied with 1 mM NH4+ plus 0.1 mM NO3−, a situation where NRT1.1 is likely to mediate very low NO3− transport. Thus, we suggest that it is the lack of NRT1.1 activity, rather than the absence of this transporter, that derepresses NRT2.1 expression in the presence of NH4+. Two hypotheses are discussed to explain these results: (1) NRT2.1 is upregulated by a NO3− demand signaling, indirectly triggered by lack of NRT1.1-mediated uptake, which overrides feedback repression by

  1. Light-induced movement of the LOV2 domain in an Asp720Asn mutant LOV2-kinase fragment of Arabidopsis phototropin 2.

    PubMed

    Takayama, Yuki; Nakasako, Masayoshi; Okajima, Koji; Iwata, Aya; Kashojiya, Sachiko; Matsui, Yuka; Tokutomi, Satoru

    2011-02-22

    Phototropin, a blue-light receptor protein of plants, triggers phototropic responses, chloroplast relocation, and opening of stomata to maximize the efficiency of photosynthesis. Phototropin is composed of two light-oxygen-voltage sensing domains (LOV1 and LOV2) that absorb blue light and a serine/theroine kinase domain responsible for light-dependent autophosphorylation leading to cellular signaling cascades. Although the light-activated LOV2 domain is primarily responsible for subsequent activation of the kinase domain, it is unclear how conformational changes in the former transmit to the latter. To understand this molecular mechanism in Arabidopsis phototropin 2, we performed small-angle X-ray scattering analysis on a fragment composed of the LOV2 and kinase domains, which contained an Asp720Asn mutation that led to an absence of ATP binding activity. The scattering data were collected up to a resolution of 25 Å. The apparent molecular weight of the fragment estimated from scattering intensities demonstrated that the fragment existed in a monomeric form in solution. The fragment exhibited photoreversible changes in the scattering profiles, and the radii of gyration under dark and blue-light irradiation conditions were 32.4 and 34.8 Å, respectively. In the dark, the molecular shape restored from the scattering profile appeared as an elongated shape of 110 Å in length and 45 Å in width. The homology modeled LOV2 and kinase domains could be fitted to the molecular shape and appeared to make slight contact. However, under blue-light irradiation, a more extended molecular shape was observed. The changes in the molecular shape and radius of gyration were interpreted as a light-dependent positional shift of the LOV2 domain of approximately 13 Å from the kinase domain. Because the region connecting the LOV2 and kinase domains was categorized as a naturally unfolded polypeptide, we propose that the light-activated LOV2 domain triggers conformational changes in the

  2. Defective etioplasts observed in variegation mutants may reveal the light-independent regulation of white/yellow sectors of Arabidopsis leaves.

    PubMed

    Wu, Wenjuan; Elsheery, Nabil; Wei, Qing; Zhang, Lingang; Huang, Jirong

    2011-11-01

    Leaf variegation resulting from nuclear gene mutations has been used as a model system to elucidate the molecular mechanisms of chloroplast development. Since most variegation genes also function in photosynthesis, it remains unknown whether their roles in photosynthesis and chloroplast development are distinct. Here, using the variegation mutant thylakoid formation1 (thf1) we show that variegation formation is light independent. It was found that slow and uneven chloroplast development in thf1 can be attributed to defects in etioplast development in darkness. Ultrastructural analysis showed the coexistence of plastids with or without prolamellar bodies (PLB) in cells of thf1, but not of WT. Although THF1 mutation leads to significant decreases in the levels of Pchlide and Pchllide oxidoreductase (POR) expression, genetic and 5-aminolevulinic acid (ALA)-feeding analysis did not reveal Pchlide or POR to be critical factors for etioplast formation in thf1. Northern blot analysis showed that plastid gene expression is dramatically reduced in thf1 compared with that in WT, particularly in the dark. Our results also indicate that chlorophyll biosynthesis and expression of plastidic genes are coordinately suppressed in thf1. Based on these results, we propose a model to explain leaf variegation formation from the plastid development perspective.

  3. Functional expression in yeast of an N-deleted form of At-ACA8, a plasma membrane Ca(2+)-ATPase of Arabidopsis thaliana, and characterization of a hyperactive mutant.

    PubMed

    Bonza, Maria Cristina; Luoni, Laura; De Michelis, Maria Ida

    2004-03-01

    A constitutively active form of At-ACA8, a plasma membrane Ca(2+)-ATPase from Arabidopsis thaliana (L.) Heynh., from which the first 74 amino acids containing the calmodulin-binding domain (delta74- At-ACA8) had been deleted, was expressed in Saccharomyces cerevisiae strain K616, which lacks the main endogenous active Ca(2+) transport systems. Delta74- At-ACA8 complemented the K616 phenotype, making it able to grow in a calcium-depleted medium. Delta74- At-ACA8 protein, which co-migrated with the endoplasmic reticulum marker BiP in a sucrose-density gradient, catalyzed MgATP-dependent Ca(2+) uptake and Ca(2+)-dependent MgATP hydrolysis, and retained the biochemical characteristics of the native plant plasma membrane Ca(2+)-ATPase (low specificity for nucleoside triphosphate, high sensitivity to inhibition by the fluorescein derivatives erythrosin B and eosin Y), thus confirming that it is correctly folded and functional. Substitution of the (794)HE residues (numbers refer to full-length At-ACA8) following the highly conserved TGDG(TV)NDP(AS)L motif in the cytoplasmic headpiece with two lysine residues generated an hyperactive protein, with a catalytic activity 2-fold higher than that of delta74- At-ACA8. The (794)HE-->KK mutant was also about 6-fold more sensitive than delta74- At-ACA8 to inhibition by vanadate, indicating that the mutation determines an increase in the proportion of enzyme in the E(2) state during the catalytic cycle.

  4. Characterization of Arabidopsis 6-phosphogluconolactonase T-DNA insertion mutants reveals an essential role for the oxidative section of the plastidic pentose phosphate pathway in plant growth and development.

    PubMed

    Xiong, Yuqing; DeFraia, Christopher; Williams, Donna; Zhang, Xudong; Mou, Zhonglin

    2009-07-01

    Arabidopsis PGL1, PGL2, PGL4 and PGL5 are predicted to encode cytosolic isoforms of 6-phosphogluconolactonase (6PGL), whereas PGL3 is predicted to encode a 6PGL that has been shown to localize in both plastids and peroxisomes. Therefore, 6PGL may exist in the cytosol, plastids and peroxisomes. However, the function of 6PGL in these three subcellular locations has not been well defined. Here we show that PGL3 is essential, whereas PGL1, PGL2 and PGL5 are individually dispensable for plant growth and development. Knockdown of PGL3 in the pgl3 mutant leads to a dramatic decrease in plant size, a significant increase in total glucose-6-phosphate dehydrogenase activity and a marked decrease in cellular redox potential. Interestingly, the pgl3 plants exhibit constitutive pathogenesis-related gene expression and enhanced resistance to Pseudomonas syringae pv. maculicola ES4326 and Hyaloperonospora arabidopsidis Noco2. We found that although pgl3 does not spontaneously accumulate elevated levels of free salicylic acid (SA), the constitutive defense responses in pgl3 plants are almost completely suppressed by the npr1 and sid2/eds16/ics1 mutations, suggesting that the pgl3 mutation activates NPR1- and SID2/EDS16/ICS1-dependent defense responses. We demonstrate that plastidic (not peroxisomal) localization and 6PGL activity of the PGL3 protein are essential for complementing all pgl3 phenotypes, indicating that the oxidative section of the plastidic pentose phosphate pathway (PPP) is required for plant normal growth and development. Thus, pgl3 provides a useful tool not only for defining the role of the PPP in different subcellular compartments, but also for dissecting the SA/NPR1-mediated signaling pathway.

  5. Starch Metabolism in Arabidopsis

    PubMed Central

    Streb, Sebastian; Zeeman, Samuel C.

    2012-01-01

    Starch is the major non-structural carbohydrate in plants. It serves as an important store of carbon that fuels plant metabolism and growth when they are unable to photosynthesise. This storage can be in leaves and other green tissues, where it is degraded during the night, or in heterotrophic tissues such as roots, seeds and tubers, where it is stored over longer time periods. Arabidopsis accumulates starch in many of its tissues, but mostly in its leaves during the day. It has proven to be a powerful genetic system for discovering how starch is synthesised and degraded, and new proteins and processes have been discovered. Such work has major significance for our starch crops, whose yield and quality could be improved by the application of this knowledge. Research into Arabidopsis starch metabolism has begun to reveal how its daily turnover is integrated into the rest of metabolism and adapted to the environmental conditions. Furthermore, Arabidopsis mutant lines deficient in starch metabolism have been employed as tools to study other biological processes ranging from sugar sensing to gravitropism and flowering time control. This review gives a detailed account of the use of Arabidopsis to study starch metabolism. It describes the major discoveries made and presents an overview of our understanding today, together with some as-yet unresolved questions. PMID:23393426

  6. AtCCX3 is an Arabidopsis endomembrane H(+)-dependent K(+) transporter

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Arabidopsis ("Arabidopsis thaliana") cation calcium exchangers (CCXs) were recently identified as a subfamily of cation transporters; however, no plant "CCXs" have been functionally characterized. Here, we show that Arabidopsis AtCCX3 (At3g14070) and AtCCX4 (At1g54115) can suppress yeast mutants...

  7. The Arabidopsis TAC Position Viewer: a high-resolution map of transformation-competent artificial chromosome (TAC) clones aligned with the Arabidopsis thaliana Columbia-0 genome.

    PubMed

    Hirose, Yoshitsugu; Suda, Kunihiro; Liu, Yao-Guang; Sato, Shusei; Nakamura, Yukino; Yokoyama, Koji; Yamamoto, Naoki; Hanano, Shigeru; Takita, Eiji; Sakurai, Nozomu; Suzuki, Hideyuki; Nakamura, Yasukazu; Kaneko, Takakazu; Yano, Kentaro; Tabata, Satoshi; Shibata, Daisuke

    2015-09-01

    We present a high-resolution map of genomic transformation-competent artificial chromosome (TAC) clones extending over all Arabidopsis thaliana (Arabidopsis) chromosomes. The Arabidopsis genomic TAC clones have been valuable genetic tools. Previously, we constructed an Arabidopsis genomic TAC library consisting of more than 10,000 TAC clones harboring large genomic DNA fragments extending over the whole Arabidopsis genome. Here, we determined 13,577 end sequences from 6987 Arabidopsis TAC clones and mapped 5937 TAC clones to precise locations, covering approximately 90% of the Arabidopsis chromosomes. We present the large-scale data set of TAC clones with high-resolution mapping information as a Java application tool, the Arabidopsis TAC Position Viewer, which provides ready-to-go transformable genomic DNA clones corresponding to certain loci on Arabidopsis chromosomes. The TAC clone resources will accelerate genomic DNA cloning, positional walking, complementation of mutants and DNA transformation for heterologous gene expression.

  8. Genetic analysis of photoreceptor action pathways in Arabidopsis thaliana

    SciTech Connect

    Not Available

    1991-01-01

    The specific strategies and long-term goals of this proposal remain intact relative to the original proposal. We continue to isolate and characterize photomorphogenic mutants of Arabidopsis thaliana. The molecular and biochemical characterization of one of these mutants, det1, has led to one publication of original data and to one Society for Experimental Biology Symposium paper (see below). The phenotype of a second mutant, det2, has also been studied during this funding period. In addition, we have continued work on a general strategy to isolate mutations in trans-acting regulatory factors that mediate light-regulated gene expression, and have identified several potentially interesting regulatory mutants. In the third funding period, we will concentrate on the genetical, biochemical, and molecular characterization of these new mutants. Construction of double mutants between the new mutants and the previously characterized morphological mutants should allow us to construct a pathway for light-regulated seedling development in Arabidopsis.

  9. Arabidopsis thaliana life without phytochromes

    PubMed Central

    Strasser, Bárbara; Sánchez-Lamas, Maximiliano; Yanovsky, Marcelo J.; Casal, Jorge J.; Cerdán, Pablo D.

    2010-01-01

    Plants use light as a source of energy for photosynthesis and as a source of environmental information perceived by photoreceptors. Testing whether plants can complete their cycle if light provides energy but no information about the environment requires a plant devoid of phytochromes because all photosynthetically active wavelengths activate phytochromes. Producing such a quintuple mutant of Arabidopsis thaliana has been challenging, but we were able to obtain it in the flowering locus T (ft) mutant background. The quintuple phytochrome mutant does not germinate in the FT background, but it germinates to some extent in the ft background. If germination problems are bypassed by the addition of gibberellins, the seedlings of the quintuple phytochrome mutant exposed to red light produce chlorophyll, indicating that phytochromes are not the sole red-light photoreceptors, but they become developmentally arrested shortly after the cotyledon stage. Blue light bypasses this blockage, rejecting the long-standing idea that the blue-light receptors cryptochromes cannot operate without phytochromes. After growth under white light, returning the quintuple phytochrome mutant to red light resulted in rapid senescence of already expanded leaves and severely impaired expansion of new leaves. We conclude that Arabidopsis development is stalled at several points in the presence of light suitable for photosynthesis but providing no photomorphogenic signal. PMID:20176939

  10. Transgenic evaluation of activated mutant alleles of SOS2 reveals a critical requirement for its kinase activity and C-terminal regulatory domain for salt tolerance in Arabidopsis thaliana

    DOEpatents

    Zhu, Jian-Kang; Quintero-Toscano, Francisco Javier; Pardo-Prieto, Jose Manuel; Qiu, Quansheng; Schumaker, Karen Sue; Ohta, Masaru; Zhang, Changqing; Guo, Yan

    2007-09-04

    The present invention provides a method of increasing salt tolerance in a plant by overexpressing a gene encoding a mutant SOS2 protein in at least one cell type in the plant. The present invention also provides for transgenic plants expressing the mutant SOS2 proteins.

  11. A survey of dominant mutations in Arabidopsis thaliana.

    PubMed

    Meinke, David W

    2013-02-01

    Following the recent publication of a comprehensive dataset of 2400 genes with a loss-of-function mutant phenotype in Arabidopsis (Arabidopsis thaliana), questions remain concerning the diversity of dominant mutations in Arabidopsis. Most of these dominant phenotypes are expected to result from inappropriate gene expression, novel protein function, or disrupted protein complexes. This review highlights the major classes of dominant mutations observed in model organisms and presents a collection of 200 Arabidopsis genes associated with a dominant or semidominant phenotype. Emphasis is placed on mutants identified through forward genetic screens of mutagenized or activation-tagged populations. These datasets illustrate the variety of genetic changes and protein functions that underlie dominance in Arabidopsis and may ultimately contribute to phenotypic variation in flowering plants.

  12. Molecular foundations of reproductive lethality in Arabidopsis thaliana.

    PubMed

    Muralla, Rosanna; Lloyd, Johnny; Meinke, David

    2011-01-01

    The SeedGenes database (www.seedgenes.org) contains information on more than 400 genes required for embryo development in Arabidopsis. Many of these EMBRYO-DEFECTIVE (EMB) genes encode proteins with an essential function required throughout the life cycle. This raises a fundamental question. Why does elimination of an essential gene in Arabidopsis often result in embryo lethality rather than gametophyte lethality? In other words, how do mutant (emb) gametophytes survive and participate in fertilization when an essential cellular function is disrupted? Furthermore, why do some mutant embryos proceed further in development than others? To address these questions, we first established a curated dataset of genes required for gametophyte development in Arabidopsis based on information extracted from the literature. This provided a basis for comparison with EMB genes obtained from the SeedGenes dataset. We also identified genes that exhibited both embryo and gametophyte defects when disrupted by a loss-of-function mutation. We then evaluated the relationship between mutant phenotype, gene redundancy, mutant allele strength, gene expression pattern, protein function, and intracellular protein localization to determine what factors influence the phenotypes of lethal mutants in Arabidopsis. After removing cases where continued development potentially resulted from gene redundancy or residual function of a weak mutant allele, we identified numerous examples of viable mutant (emb) gametophytes that required further explanation. We propose that the presence of gene products derived from transcription in diploid (heterozygous) sporocytes often enables mutant gametophytes to survive the loss of an essential gene in Arabidopsis. Whether gene disruption results in embryo or gametophyte lethality therefore depends in part on the ability of residual, parental gene products to support gametophyte development. We also highlight here 70 preglobular embryo mutants with a zygotic pattern

  13. Drought and Salt Stress Tolerance of an Arabidopsis Glutathione S-Transferase U17 Knockout Mutant Are Attributed to the Combined Effect of Glutathione and Abscisic Acid1[C][W][OA

    PubMed Central

    Chen, Jui-Hung; Jiang, Han-Wei; Hsieh, En-Jung; Chen, Hsing-Yu; Chien, Ching-Te; Hsieh, Hsu-Liang; Lin, Tsan-Piao

    2012-01-01

    Although glutathione S-transferases (GSTs) are thought to play major roles in oxidative stress metabolism, little is known about the regulatory functions of GSTs. We have reported that Arabidopsis (Arabidopsis thaliana) GLUTATHIONE S-TRANSFERASE U17 (AtGSTU17; At1g10370) participates in light signaling and might modulate various aspects of development by affecting glutathione (GSH) pools via a coordinated regulation with phytochrome A. Here, we provide further evidence to support a negative role of AtGSTU17 in drought and salt stress tolerance. When AtGSTU17 was mutated, plants were more tolerant to drought and salt stresses compared with wild-type plants. In addition, atgstu17 accumulated higher levels of GSH and abscisic acid (ABA) and exhibited hyposensitivity to ABA during seed germination, smaller stomatal apertures, a lower transpiration rate, better development of primary and lateral root systems, and longer vegetative growth. To explore how atgstu17 accumulated higher ABA content, we grew wild-type plants in the solution containing GSH and found that they accumulated ABA to a higher extent than plants grown in the absence of GSH, and they also exhibited the atgstu17 phenotypes. Wild-type plants treated with GSH also demonstrated more tolerance to drought and salt stresses. Furthermore, the effect of GSH on root patterning and drought tolerance was confirmed by growing the atgstu17 in solution containing l-buthionine-(S,R)-sulfoximine, a specific inhibitor of GSH biosynthesis. In conclusion, the atgstu17 phenotype can be explained by the combined effect of GSH and ABA. We propose a role of AtGSTU17 in adaptive responses to drought and salt stresses by functioning as a negative component of stress-mediated signal transduction pathways. PMID:22095046

  14. Forward genetic screen for auxin-deficient mutants by cytokinin.

    PubMed

    Wu, Lei; Luo, Pan; Di, Dong-Wei; Wang, Li; Wang, Ming; Lu, Cheng-Kai; Wei, Shao-Dong; Zhang, Li; Zhang, Tian-Zi; Amakorová, Petra; Strnad, Miroslav; Novák, Ondřej; Guo, Guang-Qin

    2015-07-06

    Identification of mutants with impairments in auxin biosynthesis and dynamics by forward genetic screening is hindered by the complexity, redundancy and necessity of the pathways involved. Furthermore, although a few auxin-deficient mutants have been recently identified by screening for altered responses to shade, ethylene, N-1-naphthylphthalamic acid (NPA) or cytokinin (CK), there is still a lack of robust markers for systematically isolating such mutants. We hypothesized that a potentially suitable phenotypic marker is root curling induced by CK, as observed in the auxin biosynthesis mutant CK-induced root curling 1 / tryptophan aminotransferase of Arabidopsis 1 (ckrc1/taa1). Phenotypic observations, genetic analyses and biochemical complementation tests of Arabidopsis seedlings displaying the trait in large-scale genetic screens showed that it can facilitate isolation of mutants with perturbations in auxin biosynthesis, transport and signaling. However, unlike transport/signaling mutants, the curled (or wavy) root phenotypes of auxin-deficient mutants were significantly induced by CKs and could be rescued by exogenous auxins. Mutants allelic to several known auxin biosynthesis mutants were re-isolated, but several new classes of auxin-deficient mutants were also isolated. The findings show that CK-induced root curling provides an effective marker for discovering genes involved in auxin biosynthesis or homeostasis.

  15. Photoperiod Affects the Phenotype of Mitochondrial Complex I Mutants.

    PubMed

    Pétriacq, Pierre; de Bont, Linda; Genestout, Lucie; Hao, Jingfang; Laureau, Constance; Florez-Sarasa, Igor; Rzigui, Touhami; Queval, Guillaume; Gilard, Françoise; Mauve, Caroline; Guérard, Florence; Lamothe-Sibold, Marlène; Marion, Jessica; Fresneau, Chantal; Brown, Spencer; Danon, Antoine; Krieger-Liszkay, Anja; Berthomé, Richard; Ribas-Carbo, Miquel; Tcherkez, Guillaume; Cornic, Gabriel; Pineau, Bernard; Gakière, Bertrand; De Paepe, Rosine

    2017-01-01

    Plant mutants for genes encoding subunits of mitochondrial complex I (CI; NADH:ubiquinone oxidoreductase), the first enzyme of the respiratory chain, display various phenotypes depending on growth conditions. Here, we examined the impact of photoperiod, a major environmental factor controlling plant development, on two Arabidopsis (Arabidopsis thaliana) CI mutants: a new insertion mutant interrupted in both ndufs8.1 and ndufs8.2 genes encoding the NDUFS8 subunit and the previously characterized ndufs4 CI mutant. In the long day (LD) condition, both ndufs8.1 and ndufs8.2 single mutants were indistinguishable from Columbia-0 at phenotypic and biochemical levels, whereas the ndufs8.1 ndufs8.2 double mutant was devoid of detectable holo-CI assembly/activity, showed higher alternative oxidase content/activity, and displayed a growth retardation phenotype similar to that of the ndufs4 mutant. Although growth was more affected in ndufs4 than in ndufs8.1 ndufs8.2 under the short day (SD) condition, both mutants displayed a similar impairment of growth acceleration after transfer to LD compared with the wild type. Untargeted and targeted metabolomics showed that overall metabolism was less responsive to the SD-to-LD transition in mutants than in the wild type. The typical LD acclimation of carbon and nitrogen assimilation as well as redox-related parameters was not observed in ndufs8.1 ndufs8 Similarly, NAD(H) content, which was higher in the SD condition in both mutants than in Columbia-0, did not adjust under LD We propose that altered redox homeostasis and NAD(H) content/redox state control the phenotype of CI mutants and photoperiod acclimation in Arabidopsis.

  16. Genetic analysis of photoreceptor action pathways in Arabidopsis thaliana. Progress report

    SciTech Connect

    Not Available

    1991-12-31

    The specific strategies and long-term goals of this proposal remain intact relative to the original proposal. We continue to isolate and characterize photomorphogenic mutants of Arabidopsis thaliana. The molecular and biochemical characterization of one of these mutants, det1, has led to one publication of original data and to one Society for Experimental Biology Symposium paper (see below). The phenotype of a second mutant, det2, has also been studied during this funding period. In addition, we have continued work on a general strategy to isolate mutations in trans-acting regulatory factors that mediate light-regulated gene expression, and have identified several potentially interesting regulatory mutants. In the third funding period, we will concentrate on the genetical, biochemical, and molecular characterization of these new mutants. Construction of double mutants between the new mutants and the previously characterized morphological mutants should allow us to construct a pathway for light-regulated seedling development in Arabidopsis.

  17. Mining the plant-herbivore interface with a leafmining Drosophila of Arabidopsis

    PubMed Central

    Whiteman, Noah K.; Groen, Simon C.; Chevasco, Daniela; Bear, Ashley; Beckwith, Noor; Gregory, T. Ryan; Denoux, Carine; Mammarella, Nicole; Ausubel, Frederick M.; Pierce, Naomi E.

    2010-01-01

    Experimental infections of Arabidopsis thaliana (Arabidopsis) with genomically characterized plant pathogens such as Pseudomonas syringae have facilitated dissection of canonical eukaryotic defense pathways and parasite virulence factors. Plants are also attacked by herbivorous insects, and the development of an ecologically relevant genetic model herbivore that feeds on Arabidopsis will enable the parallel dissection of host defense and reciprocal resistance pathways such as those involved in xenobiotic metabolism. An ideal candidate is Scaptomyza flava, a drosophilid fly whose leafmining larvae are true herbivores that can be found in nature feeding on Arabidopsis and other crucifers. Here we describe the eukaryotic life cycle of S. flava on Arabidopsis, and use multiple approaches to characterize the response of Arabidopsis to S. flava attack. Oviposition choice tests and growth performance assays on different Arabidopsis ecotypes, defense-related mutants, and hormone and chitin-treated plants revealed significant differences in host preference and variation in larval performance across Arabidopsis accessions. The jasmonate (JA) and glucosinolate pathways in Arabidopsis are important in mediating quantitative resistance against S. flava, and priming with JA or chitin resulted in increased resistance. Expression of xenobiotic detoxification genes was reduced in S. flava larvae reared on Arabidopsis JA signaling mutants, and increased in plants pre-treated with chitin. These results and future research directions are discussed in the context of developing a genetic model system to analyze insect/plant interactions. PMID:21073583

  18. Meta-analyses of microarrays of Arabidopsis asymmetric leaves1 (as1), as2 and their modifying mutants reveal a critical role for the ETT pathway in stabilization of adaxial-abaxial patterning and cell division during leaf development.

    PubMed

    Takahashi, Hiro; Iwakawa, Hidekazu; Ishibashi, Nanako; Kojima, Shoko; Matsumura, Yoko; Prananingrum, Pratiwi; Iwasaki, Mayumi; Takahashi, Anna; Ikezaki, Masaya; Luo, Lilan; Kobayashi, Takeshi; Machida, Yasunori; Machida, Chiyoko

    2013-03-01

    It is necessary to use algorithms to analyze gene expression data from DNA microarrays, such as in clustering and machine learning. Previously, we developed the knowledge-based fuzzy adaptive resonance theory (KB-FuzzyART), a clustering algorithm suitable for analyzing gene expression data, to find clues for identifying gene networks. Leaf primordia form around the shoot apical meristem (SAM), which consists of indeterminate stem cells. Upon initiation of leaf development, adaxial-abaxial patterning is crucial for lateral expansion, via cellular proliferation, and the formation of flat symmetric leaves. Many regulatory genes that specify such patterning have been identified. Analysis by the KB-FuzzyART and subsequent molecular and genetic analyses previously showed that ASYMMETRIC LEAVES1 (AS1) and AS2 repress the expression of some abaxial-determinant genes, such as AUXIN RESPONSE FACTOR3 (ARF3)/ETTIN (ETT) and ARF4, which are responsible for defects in leaf adaxial-abaxial polarity in as1 and as2. In the present study, genetic analysis revealed that ARF3/ETT and ARF4 were regulated by modifier genes, BOBBER1 (BOB1) and ELONGATA3 (ELO3), together with AS1-AS2. We analyzed expression arrays with as2 elo3 and as2 bob1, and extracted genes downstream of ARF3/ETT by using KB-FuzzyART and molecular analyses. The results showed that expression of Kip-related protein (KRP) (for inhibitors of cyclin-dependent protein kinases) and Isopentenyltransferase (IPT) (for biosynthesis of cytokinin) genes were controlled by AS1-AS2 through ARF3/ETT and ARF4 functions, which suggests that the AS1-AS2-ETT pathway plays a critical role in controlling the cell division cycle and the biosynthesis of cytokinin around SAM to stabilize leaf development in Arabidopsis thaliana.

  19. Thermoinhibition uncovers a role for strigolactones in Arabidopsis seed germination.

    PubMed

    Toh, Shigeo; Kamiya, Yuji; Kawakami, Naoto; Nambara, Eiji; McCourt, Peter; Tsuchiya, Yuichiro

    2012-01-01

    Strigolactones are host factors that stimulate seed germination of parasitic plant species such as Striga and Orobanche. This hormone is also important in shoot branching architecture and photomorphogenic development. Strigolactone biosynthetic and signaling mutants in model systems, unlike parasitic plants, only show seed germination phenotypes under limited growth condition. To understand the roles of strigolactones in seed germination, it is necessary to develop a tractable experimental system using model plants such as Arabidopsis. Here, we report that thermoinhibition, which involves exposing seeds to high temperatures, uncovers a clear role for strigolactones in promoting Arabidopsis seed germination. Both strigolactone biosynthetic and signaling mutants showed increased sensitivity to seed thermoinhibition. The synthetic strigolactone GR24 rescued germination of thermoinbibited biosynthetic mutant seeds but not a signaling mutant. Hormone analysis revealed that strigolactones alleviate thermoinhibition by modulating levels of the two plant hormones, GA and ABA. We also showed that GR24 was able to counteract secondary dormancy in Arabidopsis ecotype Columbia (Col) and Cape Verde island (Cvi). Systematic hormone analysis of germinating Striga helmonthica seeds suggested a common mechanism between the parasitic and non-parasitic seeds with respect to how hormones regulate germination. Thus, our simple assay system using Arabidopsis thermoinhibition allows comparisons to determine similarities and differences between parasitic plants and model experimental systems for the use of strigolactones.

  20. Arabidopsis ECERIFERUM9 involvement in cuticle formation and maintenance of plant water status

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A unique set of allelic Arabidopsis mutants are described that exhibit either suppressed or completely inhibited expression of a gene designated ECERIFERUM9 (CER9). These mutants exhibit a dramatic elevation in the total amount of leaf cutin monomers, and a dramatic shift in the leaf cuticular wax p...

  1. Sodium Influx and Accumulation in Arabidopsis1

    PubMed Central

    Essah, Pauline A.; Davenport, Romola; Tester, Mark

    2003-01-01

    Arabidopsis is frequently used as a genetic model in plant salt tolerance studies, however, its physiological responses to salinity remain poorly characterized. This study presents a characterization of initial Na+ entry and the effects of Ca2+ on plant growth and net Na+ accumulation in saline conditions. Unidirectional Na+ influx was measured carefully using very short influx times in roots of 12-d-old seedlings. Influx showed three components with distinct sensitivities to Ca2+, diethylpyrocarbonate, and osmotic pretreatment. Pharmacological agents and known mutants were used to test the contribution of different transport pathways to Na+ uptake. Influx was stimulated by 4-aminobutyric acid and glutamic acid; was inhibited by flufenamate, quinine, and cGMP; and was insensitive to modulators of K+ and Ca2+ channels. Influx did not differ from wild type in akt1 and hkt1 insertional mutants. These data suggested that influx was mediated by several different types of nonselective cation channels. Na+ accumulation in plants grown in 50 mm NaCl was strongly reduced by increasing Ca2+ activity (from 0.05-3.0 mm), and plant survival was improved. However, plant biomass was not affected by shoot Na+ concentration, suggesting that in Arabidopsis Na+ toxicity is not dependent on shoot Na+ accumulation. These data suggest that Arabidopsis is a good model for investigation of Na+ transport, but may be of limited utility as a model for the study of Na+ toxicity. PMID:12970496

  2. Arabidopsis hybrid speciation processes

    PubMed Central

    Schmickl, Roswitha; Koch, Marcus A.

    2011-01-01

    The genus Arabidopsis provides a unique opportunity to study fundamental biological questions in plant sciences using the diploid model species Arabidopsis thaliana and Arabidopsis lyrata. However, only a few studies have focused on introgression and hybrid speciation in Arabidopsis, although polyploidy is a common phenomenon within this genus. More recently, there is growing evidence of significant gene flow between the various Arabidopsis species. So far, we know Arabidopsis suecica and Arabidopsis kamchatica as fully stabilized allopolyploid species. Both species evolved during Pleistocene glaciation and deglaciation cycles in Fennoscandinavia and the amphi-Beringian region, respectively. These hybrid studies were conducted either on a phylogeographic scale or reconstructed experimentally in the laboratory. In our study we focus at a regional and population level. Our research area is located in the foothills of the eastern Austrian Alps, where two Arabidopsis species, Arabidopsis arenosa and A. lyrata ssp. petraea, are sympatrically distributed. Our hypothesis of genetic introgression, migration, and adaptation to the changing environment during the Pleistocene has been confirmed: We observed significant, mainly unidirectional gene flow between the two species, which has given rise to the tetraploid A. lyrata. This cytotype was able to escape from the narrow ecological niche occupied by diploid A. lyrata ssp. petraea on limestone outcrops by migrating northward into siliceous areas, leaving behind a trail of genetic differentiation. PMID:21825128

  3. Strigolactones suppress adventitious rooting in Arabidopsis and pea.

    PubMed

    Rasmussen, Amanda; Mason, Michael Glenn; De Cuyper, Carolien; Brewer, Philip B; Herold, Silvia; Agusti, Javier; Geelen, Danny; Greb, Thomas; Goormachtig, Sofie; Beeckman, Tom; Beveridge, Christine Anne

    2012-04-01

    Adventitious root formation is essential for the propagation of many commercially important plant species and involves the formation of roots from nonroot tissues such as stems or leaves. Here, we demonstrate that the plant hormone strigolactone suppresses adventitious root formation in Arabidopsis (Arabidopsis thaliana) and pea (Pisum sativum). Strigolactone-deficient and response mutants of both species have enhanced adventitious rooting. CYCLIN B1 expression, an early marker for the initiation of adventitious root primordia in Arabidopsis, is enhanced in more axillary growth2 (max2), a strigolactone response mutant, suggesting that strigolactones restrain the number of adventitious roots by inhibiting the very first formative divisions of the founder cells. Strigolactones and cytokinins appear to act independently to suppress adventitious rooting, as cytokinin mutants are strigolactone responsive and strigolactone mutants are cytokinin responsive. In contrast, the interaction between the strigolactone and auxin signaling pathways in regulating adventitious rooting appears to be more complex. Strigolactone can at least partially revert the stimulatory effect of auxin on adventitious rooting, and auxin can further increase the number of adventitious roots in max mutants. We present a model depicting the interaction of strigolactones, cytokinins, and auxin in regulating adventitious root formation.

  4. Root gravitropism in maize and Arabidopsis

    NASA Technical Reports Server (NTRS)

    Evans, Michael L.

    1993-01-01

    Research during the period 1 March 1992 to 30 November 1993 focused on improvements in a video digitizer system designed to automate the recording of surface extension in plants responding to gravistimulation. The improvements included modification of software to allow detailed analysis of localized extension patterns in roots of Arabidopsis. We used the system to analyze the role of the postmitotic isodiametric growth zone (a region between the meristem and the elongation zone) in the response of maize roots to auxin, calcium, touch and gravity. We also used the system to analyze short-term auxin and gravitropic responses in mutants of Arabidopsis with reduced auxin sensitivity. In a related project, we studied the relationship between growth rate and surface electrical currents in roots by examining the effects of gravity and thigmostimulation on surface potentials in maize roots.

  5. Isolation of New Gravitropic Mutants under Hypergravity Conditions

    PubMed Central

    Mori, Akiko; Toyota, Masatsugu; Shimada, Masayoshi; Mekata, Mika; Kurata, Tetsuya; Tasaka, Masao; Morita, Miyo T.

    2016-01-01

    Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upward. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes). In the present study, we report a new screening system using hypergravity conditions to isolate enhancers of gravitropism mutants, and we also describe a rapid and efficient genome mapping method, using next-generation sequencing (NGS) and single nucleotide polymorphism (SNP)-based markers. Using the endodermal-amyloplast less 1 (eal1) mutant, which exhibits defective development of endodermal cells and gravitropism, we found that hypergravity (10 g) restored the reduced gravity responsiveness in eal1 hypocotyls and could, therefore, be used to obtain mutants with further reduction in gravitropism in the eal1 background. Using the new screening system, we successfully isolated six ene (enhancer of eal1) mutants that exhibited little or no gravitropism under hypergravity conditions, and using NGS and map-based cloning with SNP markers, we narrowed down the potential causative genes, which revealed a new genetic network for shoot gravitropism in Arabidopsis. PMID:27746791

  6. Role of a chalcone isomerase-like protein in flavonoid biosynthesis in Arabidopsis thaliana.

    PubMed

    Jiang, Wenbo; Yin, Qinggang; Wu, Ranran; Zheng, Guangshun; Liu, Jinyue; Dixon, Richard A; Pang, Yongzhen

    2015-12-01

    Flavonoids are important natural products for plant defence and human health. Although almost all the flavonoid pathway genes have been well-documented by biochemical and/or genetic approaches, the role of the Arabidopsis chalcone isomerase-like (CHIL) gene remains unclear. Two chil mutants with a seed colour similar to that of wild-type Arabidopsis have been identified here, but in sharp contrast to the characteristic transparent testa seed phenotype associated with other known flavonoid pathway genes. CHIL loss-of-function mutations led to a strong reduction in the proanthocyanidin and flavonol levels in seeds, but not in the anthocyanin levels in leaves. CHIL over-expression could partially recover the mutant phenotype of the chil mutant and increased both proanthocyanidin and flavonol accumulation in wild-type Arabidopsis. However, the CHIL gene could not rescue the mutant phenotype of TT5 that encodes the intrinsic chalcone isomerase in Arabidopsis. Parallel phenotypical and metabolic analyses of the chil, tt5, chs, and f3h mutants revealed that, genetically, CHIL functions at the same step as TT5. Moreover, it is demonstrated that CHIL co-expresses, co-localizes, and interacts with TT5 in Arabidopsis for flavonoid production. Based on these genetic and metabolic studies, it is concluded that CHIL functions with TT5 to promote flavonoid production, which is a unique enhancer in the flavonoid pathway.

  7. The Arabidopsis Circadian System

    PubMed Central

    McClung, C. Robertson; Salomé, Patrice A.; Michael, Todd P.

    2002-01-01

    Rhythms with periods of approximately 24 hr are widespread in nature. Those that persist in constant conditions are termed circadian rhythms and reflect the activity of an endogenous biological clock. Plants, including Arabidopsis, are richly rhythmic. Expression analysis, most recently on a genomic scale, indicates that the Arabidopsis circadian clock regulates a number of key metabolic pathways and stress responses. A number of sensitive and high-throughput assays have been developed to monitor the Arabidopsis clock. These assays have facilitated the identification of components of plant circadian systems through genetic and molecular biological studies. Although much remains to be learned, the framework of the Arabidopsis circadian system is coming into focus. Dedication This review is dedicated to the memory of DeLill Nasser, a wonderful mentor and an unwavering advocate of both Arabidopsis and circadian rhythms research. PMID:22303209

  8. Using Arabidopsis to study shoot branching in biomass willow.

    PubMed

    Ward, Sally P; Salmon, Jemma; Hanley, Steven J; Karp, Angela; Leyser, Ottoline

    2013-06-01

    The success of the short-rotation coppice system in biomass willow (Salix spp.) relies on the activity of the shoot-producing meristems found on the coppice stool. However, the regulation of the activity of these meristems is poorly understood. In contrast, our knowledge of the mechanisms behind axillary meristem regulation in Arabidopsis (Arabidopsis thaliana) has grown rapidly in the past few years through the exploitation of integrated physiological, genetic, and molecular assays. Here, we demonstrate that these assays can be directly transferred to study the control of bud activation in biomass willow and to assess similarities with the known hormone regulatory system in Arabidopsis. Bud hormone response was found to be qualitatively remarkably similar in Salix spp. and Arabidopsis. These similarities led us to test whether Arabidopsis hormone mutants could be used to assess allelic variation in the cognate Salix spp. hormone genes. Allelic differences in Salix spp. strigolactone genes were observed using this approach. These results demonstrate that both knowledge and assays from Arabidopsis axillary meristem biology can be successfully applied to Salix spp. and can increase our understanding of a fundamental aspect of short-rotation coppice biomass production, allowing more targeted breeding.

  9. Testing Xylella fastidiosa pathogenesis mutants in Arabidopsis thaliana

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The bacterium Xylella fastidiosa (Xf) causes Pierce's disease and a number of other plant diseases of significant economic impact. To date, progress determining mechanisms of host plant susceptibility, tolerance or resistance has been slow, due in large part to the long generation time and limited ...

  10. IDENTIFICATION OF ARABIDOPSIS RAT MUTANTS. (R829479C001)

    EPA Science Inventory

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  11. [Imprinting genes and it's expression in Arabidopsis].

    PubMed

    Zhang, Hong-Yu; Xu, Pei-Zhou; Yang, Hua; Wu, Xian-Jun

    2010-07-01

    Genomic imprinting refers to the phenomenon that the expression of a gene copy depends on its parent of origin. The Arabidopsis imprinted FIS (Fertilisation-independent seed) genes, mea, fis2, and fie, play essential roles in the repression of central cell and the regulation of early endosperm development. fis mutants display two phenotypes: autonomous diploid endosperm development when fertilization is absent and un-cellularised endosperm formation when fertilization occurs. The FIS Polycomb protein complex including the above three FIS proteins catalyzes histone H3 K27 tri-methylation on target loci. DME (DEMETER), a DNA glycosylase, and AtMET1 (Methyltransferase1), a DNA methyltransferase, are involved in the regulation of imprinted expression of both mea and fis2. This review summarizes the studies on the Arabidopsis imprinted FIS genes and other related genes. Recent works have shown that the insertion of transposons may affect nearby gene expression, which may be the main driving force behind the evolution of genomic imprinting. This summary covers the achievements on Arabidopsis imprinted genes will provide important information for studies on genomic imprinting in the important crops such as rice and maize.

  12. Photoperiod Affects the Phenotype of Mitochondrial Complex I Mutants1[OPEN

    PubMed Central

    de Bont, Linda; Hao, Jingfang; Laureau, Constance; Rzigui, Touhami; Queval, Guillaume; Gilard, Françoise; Mauve, Caroline; Guérard, Florence; Lamothe-Sibold, Marlène; Marion, Jessica; Fresneau, Chantal; Tcherkez, Guillaume; Pineau, Bernard; De Paepe, Rosine

    2017-01-01

    Plant mutants for genes encoding subunits of mitochondrial complex I (CI; NADH:ubiquinone oxidoreductase), the first enzyme of the respiratory chain, display various phenotypes depending on growth conditions. Here, we examined the impact of photoperiod, a major environmental factor controlling plant development, on two Arabidopsis (Arabidopsis thaliana) CI mutants: a new insertion mutant interrupted in both ndufs8.1 and ndufs8.2 genes encoding the NDUFS8 subunit and the previously characterized ndufs4 CI mutant. In the long day (LD) condition, both ndufs8.1 and ndufs8.2 single mutants were indistinguishable from Columbia-0 at phenotypic and biochemical levels, whereas the ndufs8.1 ndufs8.2 double mutant was devoid of detectable holo-CI assembly/activity, showed higher alternative oxidase content/activity, and displayed a growth retardation phenotype similar to that of the ndufs4 mutant. Although growth was more affected in ndufs4 than in ndufs8.1 ndufs8.2 under the short day (SD) condition, both mutants displayed a similar impairment of growth acceleration after transfer to LD compared with the wild type. Untargeted and targeted metabolomics showed that overall metabolism was less responsive to the SD-to-LD transition in mutants than in the wild type. The typical LD acclimation of carbon and nitrogen assimilation as well as redox-related parameters was not observed in ndufs8.1 ndufs8. Similarly, NAD(H) content, which was higher in the SD condition in both mutants than in Columbia-0, did not adjust under LD. We propose that altered redox homeostasis and NAD(H) content/redox state control the phenotype of CI mutants and photoperiod acclimation in Arabidopsis. PMID:27852950

  13. Light responses in Photoperiodism in Arabidopsis thaliana

    SciTech Connect

    Anthony R. Cashmore

    2006-08-01

    ADO1: An Arabidopsis blue light photoreceptor We have reported the characterization of an Arabidopsis gene encoding the ADAGIO 1 (ADO1) protein (Jarillo et al., 2001a). ADO1 contains a LOV domain, similar to WHITE COLLAR 1 (WC1), a photoreceptor for entrainment of Neurospora circadian rhythms (Froehlich et al., 2002), as well as PHOT1 and PHOT2, the blue light photoreceptors for phototropism (Briggs et al., 2001; Christie et al., 1998; Jarillo et al., 2001b; Kinoshita et al., 2001). Loss of function ado1 mutants show an unusually long periodicity for their free running circadian rhythm (Jarillo et al., 2001a). This observation holds for plants grown under white light as well as blue light and surprisingly, plants grown under red light also show altered circadian properties. The similarity of the LOV domain of ADO1 to those of PHOT1, PHOT2 and WC1 (known flavoprotein photoreceptors) as well as the genetic and molecular properties of ADO1, indicate that ADO1 is likely a new class of blue light photoreceptor. Indeed, the LOV domain of the related FKF1/ADO3 has been shown to bind FMN, and exhibit the in vitro photochemistry characteristic of PHOT1 (Imaizumi et al., 2003). Furthermore, ZTL/ADO1 has been shown to participate in the circadian and proteasome mediated degradation of the Arabidopsis clock protein, TOC1 (Mas et al., 2003). We also showed that the ado1 mutation selectively confers hypersensitivity to red light — when grown under red light (but not blue light) the ado1 mutant possesses an unusually short hypocotyl. This red light hypersensivity is even more severe in a triple ado1 ado2 ado3 mutant — ADO2 and ADO3 being the two other members of this ADAGIO gene family. This finding of a mutant phenotype under red light is somewhat unexpected for a protein thought to function as a photoreceptor for blue light. We have pursued our studies of ADO1 by preparing a mutant gene for which we have altered the codon for the cysteine residue conserved in all LOV

  14. Adenosine-5'-phosphosulfate kinase is essential for Arabidopsis viability.

    PubMed

    Mugford, Sarah G; Matthewman, Colette A; Hill, Lionel; Kopriva, Stanislav

    2010-01-04

    In Arabidopsis thaliana, adenosine-5'-phosphosulfate kinase (APK) provides activated sulfate for sulfation of secondary metabolites, including the glucosinolates. We have successfully isolated three of the four possible triple homozygous mutant combinations of this family. The APK1 isoform alone was sufficient to maintain WT levels of growth and development. Analysis of apk1 apk2 apk3 and apk1 apk3 apk4 mutants suggests that APK3 and APK4 are functionally redundant, despite being located in cytosol and plastids, respectively. We were, however, unable to isolate apk1 apk3 apk4 mutants, most probably because the apk1 apk3 apk4 triple mutant combination is pollen lethal. Therefore, we conclude that APS kinase is essential for plant reproduction and viability.

  15. Thermospermine suppresses auxin-inducible xylem differentiation in Arabidopsis thaliana.

    PubMed

    Yoshimoto, Kaori; Noutoshi, Yoshiteru; Hayashi, Ken-ichiro; Shirasu, Ken; Takahashi, Taku; Motose, Hiroyasu

    2012-08-01

    Thermospermine, a structural isomer of spermine, is synthesized by a thermospermine synthase designated ACAULIS5 (ACL5). Thermospermine-deficient acl5 mutant of Arabidopsis thaliana shows severe dwarfism and excessive xylem differentiation. By screening for compounds that affect xylem differentiation in the acl5 mutant, we identified auxin analogs that remarkably enhanced xylem vessel differentiation in the acl5 mutant but not in the wild type. The xylem-inducing effect of auxin analogs was clearly suppressed by thermospermine, indicating that auxin-inducible xylem differentiation is normally limited by thermospermine. Here, we further characterized xylem-inducing effect of auxin analogs in various organs. Auxin analogs promoted protoxylem differentiation in roots and cotyledons in the acl5 mutant. Our results indicate that the opposite action between thermospermine and auxin in xylem differentiation is common in different organs and also suggest that thermospermine might be required for the suppression of protoxylem differentiation.

  16. A mutational analysis of leaf morphogenesis in Arabidopsis thaliana.

    PubMed Central

    Berná, G; Robles, P; Micol, J L

    1999-01-01

    As a contribution to a better understanding of the developmental processes that are specific to plants, we have begun a genetic analysis of leaf ontogeny in the model system Arabidopsis thaliana by performing a large-scale screening for mutants with abnormal leaves. After screening 46,159 M2 individuals, arising from 5770 M1 parental seeds exposed to EMS, we isolated 1926 M2 putative leaf mutants, 853 of which yielded viable M3 inbred progeny. Mutant phenotypes were transmitted with complete penetrance and small variations in expressivity in 255 lines. Most of them were inherited as recessive monogenic traits, belonging to 94 complementation groups, which suggests that we did not reach saturation of the genome. We discuss the nature of the processes presumably perturbed in the phenotypic classes defined among our mutants. PMID:10353913

  17. Insertional mutagenesis of genes required for seed development in Arabidopsis thaliana.

    PubMed Central

    McElver, J; Tzafrir, I; Aux, G; Rogers, R; Ashby, C; Smith, K; Thomas, C; Schetter, A; Zhou, Q; Cushman, M A; Tossberg, J; Nickle, T; Levin, J Z; Law, M; Meinke, D; Patton, D

    2001-01-01

    The purpose of this project was to identify large numbers of Arabidopsis genes with essential functions during seed development. More than 120,000 T-DNA insertion lines were generated following Agrobacterium-mediated transformation. Transgenic plants were screened for defective seeds and putative mutants were subjected to detailed analysis in subsequent generations. Plasmid rescue and TAIL-PCR were used to recover plant sequences flanking insertion sites in tagged mutants. More than 4200 mutants with a wide range of seed phenotypes were identified. Over 1700 of these mutants were analyzed in detail. The 350 tagged embryo-defective (emb) mutants identified to date represent a significant advance toward saturation mutagenesis of EMB genes in Arabidopsis. Plant sequences adjacent to T-DNA borders in mutants with confirmed insertion sites were used to map genome locations and establish tentative identities for 167 EMB genes with diverse biological functions. The frequency of duplicate mutant alleles recovered is consistent with a relatively small number of essential (EMB) genes with nonredundant functions during seed development. Other functions critical to seed development in Arabidopsis may be protected from deleterious mutations by extensive genome duplications. PMID:11779812

  18. Rapid Analysis of Circadian Phenotypes in Arabidopsis Protoplasts Transfected with a Luminescent Clock Reporter

    PubMed Central

    Hansen, Louise L.; van Ooijen, Gerben

    2016-01-01

    The plant circadian clock allows the anticipation of daily changes to the environment. This anticipation aids the responses to temporally predictable biotic and abiotic stress. Conversely, disruption of circadian timekeeping severely compromises plant health and reduces agricultural crop yields. It is therefore imperative that we understand the intricate regulation of circadian rhythms in plants, including the factors that affect motion of the transcriptional clockwork itself. Testing circadian defects in the model plant Arabidopsis thaliana (Arabidopsis) traditionally involves crossing specific mutant lines to a line rhythmically expressing firefly luciferase from a circadian clock gene promoter. This approach is laborious, time-consuming, and could be fruitless if a mutant has no circadian phenotype. The methodology presented here allows a rapid initial assessment of circadian phenotypes. Protoplasts derived from mutant and wild-type Arabidopsis are isolated, transfected with a rhythmically expressed luminescent reporter, and imaged under constant light conditions for 5 days. Luminescent traces will directly reveal whether the free-running period of mutant plants is different from wild-type plants. The advantage of the method is that any Arabidopsis line can efficiently be screened, without the need for generating a stably transgenic luminescent clock marker line in that mutant background. PMID:27684315

  19. Ethylene signaling in rice and Arabidopsis: conserved and diverged aspects.

    PubMed

    Yang, Chao; Lu, Xiang; Ma, Biao; Chen, Shou-Yi; Zhang, Jin-Song

    2015-04-01

    Ethylene as a gas phytohormone plays significant roles in the whole life cycle of plants, ranging from growth and development to stress responses. A linear ethylene signaling pathway has been established in the dicotyledonous model plant Arabidopsis. However, the ethylene signaling mechanism in monocotyledonous plants such as rice is largely unclear. In this review, we compare the ethylene response phenotypes of dark-grown seedlings of Arabidopsis, rice, and other monocotyledonous plants (maize, wheat, sorghum, and Brachypodium distachyon) and pinpoint that rice has a distinct phenotype of root inhibition but coleoptile promotion in etiolated seedlings upon ethylene treatment. We further summarize the homologous genes of Arabidopsis ethylene signaling components in these monocotyledonous plants and discuss recent progress. Although conserved in most aspects, ethylene signaling in rice has evolved new features compared with that in Arabidopsis. These analyses provide novel insights into the understanding of ethylene signaling in the dicotyledonous Arabidopsis and monocotyledonous plants, particularly rice. Further characterization of rice ethylene-responsive mutants and their corresponding genes will help us better understand the whole picture of ethylene signaling mechanisms in plants.

  20. A role for the TOC complex in Arabidopsis root gravitropism.

    PubMed

    Stanga, John P; Boonsirichai, Kanokporn; Sedbrook, John C; Otegui, Marisa S; Masson, Patrick H

    2009-04-01

    Arabidopsis (Arabidopsis thaliana) roots perceive gravity and reorient their growth accordingly. Starch-dense amyloplasts within the columella cells of the root cap are important for gravitropism, and starchless mutants such as pgm1 display an attenuated response to gravistimulation. The altered response to gravity1 (arg1) mutant is known to be involved with the early phases of gravity signal transduction. arg1 responds slowly to gravistimulation and is in a genetically distinct pathway from pgm1, as pgm1 mutants enhance the gravitropic defect of arg1. arg1 seeds were mutagenized with ethylmethane sulfonate to identify new mutants that enhance the gravitropic defect of arg1. Two modifier of arg1 mutants (mar1 and mar2) grow in random directions only when arg1 is present, do not affect phototropism, and respond like the wild type to application of phytohormones. Both have mutations affecting different components of the Translocon of Outer Membrane of Chloroplasts (TOC) complex. mar1 possesses a mutation in the TOC75-III gene; mar2 possesses a mutation in the TOC132 gene. Overexpression of TOC132 rescues the random growth phenotype of mar2 arg1 roots. Root cap amyloplasts in mar2 arg1 appear ultrastructurally normal. They saltate like the wild type and sediment at wild-type rates upon gravistimulation. These data point to a role for the plastidic TOC complex in gravity signal transduction within the statocytes.

  1. Methylation of Gibberellins by Arabidopsis GAMT1 and GAMT2

    SciTech Connect

    Varbanova,M.; Yamaguchi, S.; Yang, Y.; McKelvey, K.; Hanada, A.; Borochov, R.; Yu, F.; Jikumaru, Y.; Ross, J.; et al

    2007-01-01

    Arabidopsis thaliana GAMT1 and GAMT2 encode enzymes that catalyze formation of the methyl esters of gibberellins (GAs). Ectopic expression of GAMT1 or GAMT2 in Arabidopsis, tobacco (Nicotiana tabacum), and petunia (Petunia hybrida) resulted in plants with GA deficiency and typical GA deficiency phenotypes, such as dwarfism and reduced fertility. GAMT1 and GAMT2 are both expressed mainly in whole siliques (including seeds), with peak transcript levels from the middle until the end of silique development. Within whole siliques, GAMT2 was previously shown to be expressed mostly in developing seeds, and we show here that GAMT1 expression is also localized mostly to seed, suggesting a role in seed development. Siliques of null single GAMT1 and GAMT2 mutants accumulated high levels of various GAs, with particularly high levels of GA1 in the double mutant. Methylated GAs were not detected in wild-type siliques, suggesting that methylation of GAs by GAMT1 and GAMT2 serves to deactivate GAs and initiate their degradation as the seeds mature. Seeds of homozygous GAMT1 and GAMT2 null mutants showed reduced inhibition of germination, compared with the wild type, when placed on plates containing the GA biosynthesis inhibitor ancymidol, with the double mutant showing the least inhibition. These results suggest that the mature mutant seeds contained higher levels of active GAs than wild-type seeds.

  2. BODYGUARD is required for the biosynthesis of cutin in Arabidopsis.

    PubMed

    Jakobson, Liina; Lindgren, Leif Ove; Verdier, Gaëtan; Laanemets, Kristiina; Brosché, Mikael; Beisson, Fred; Kollist, Hannes

    2016-07-01

    The cuticle plays a critical role in plant survival during extreme drought conditions. There are, however, surprisingly, many gaps in our understanding of cuticle biosynthesis. An Arabidopsis thaliana T-DNA mutant library was screened for mutants with enhanced transpiration using a simple condensation spot method. Five mutants, named cool breath (cb), were isolated. The cb5 mutant was found to be allelic to bodyguard (bdg), which is affected in an α/β-hydrolase fold protein important for cuticle structure. The analysis of cuticle components in cb5 (renamed as bdg-6) and another T-DNA mutant allele (bdg-7) revealed no impairment in wax synthesis, but a strong decrease in total cutin monomer load in young leaves and flowers. Root suberin content was also reduced. Overexpression of BDG increased total leaf cutin monomer content nearly four times by affecting preferentially C18 polyunsaturated ω-OH fatty acids and dicarboxylic acids. Whole-plant gas exchange analysis showed that bdg-6 had higher cuticular conductance and rate of transpiration; however, plant lines overexpressing BDG resembled the wild-type with regard to these characteristics. This study identifies BDG as an important component of the cutin biosynthesis machinery in Arabidopsis. We also show that, using BDG, cutin can be greatly modified without altering the cuticular water barrier properties and transpiration.

  3. PAUSED Encodes the Arabidopsis Exportin-t Ortholog1

    PubMed Central

    Hunter, Christine A.; Aukerman, Milo J.; Sun, Hui; Fokina, Maria; Poethig, R. Scott

    2003-01-01

    Los1p/exportin-t (XPOT) mediates the nuclear export of tRNAs in yeast and mammals. The requirements for this transport pathway are unclear, however, because los1 mutations do not affect yeast growth, and the phenotype of XPOT mutations in mammals is unknown. Here, we show that PAUSED (PSD) is the Arabidopsis ortholog of LOS1/XPOT and is capable of rescuing the tRNA export defect of los1 in Brewer's yeast (Saccharomyces cerevisiae), suggesting that its function has been conserved. Putative null alleles of PSD disrupt the initiation of the shoot apical meristem and delay leaf initiation after germination, the emergence of the radicle and lateral roots, and the transition to flowering. Plants doubly mutant for psd and hasty, the Arabidopsis ortholog of exportin 5, are viable but have a more severe phenotype than either single mutant. These results suggest that PSD plays a role in tRNA export in Arabidopsis, but that at least one—and perhaps several—additional tRNA export pathways also exist. The PSD transcript is broadly expressed during development and is alternatively spliced in the 3′-untranslated region. No temporal or spatial difference in the abundance of different splice forms was observed. We propose that the mutant phenotype of psd reflects defects in developmental events and cell/tissue types that require elevated levels of protein synthesis and are therefore acutely sensitive to a reduction in tRNA export. PMID:12913168

  4. PAUSED encodes the Arabidopsis exportin-t ortholog.

    PubMed

    Hunter, Christine A; Aukerman, Milo J; Sun, Hui; Fokina, Maria; Poethig, R Scott

    2003-08-01

    Los1p/exportin-t (XPOT) mediates the nuclear export of tRNAs in yeast and mammals. The requirements for this transport pathway are unclear, however, because los1 mutations do not affect yeast growth, and the phenotype of XPOT mutations in mammals is unknown. Here, we show that PAUSED (PSD) is the Arabidopsis ortholog of LOS1/XPOT and is capable of rescuing the tRNA export defect of los1 in Brewer's yeast (Saccharomyces cerevisiae), suggesting that its function has been conserved. Putative null alleles of PSD disrupt the initiation of the shoot apical meristem and delay leaf initiation after germination, the emergence of the radicle and lateral roots, and the transition to flowering. Plants doubly mutant for psd and hasty, the Arabidopsis ortholog of exportin 5, are viable but have a more severe phenotype than either single mutant. These results suggest that PSD plays a role in tRNA export in Arabidopsis, but that at least one-and perhaps several-additional tRNA export pathways also exist. The PSD transcript is broadly expressed during development and is alternatively spliced in the 3'-untranslated region. No temporal or spatial difference in the abundance of different splice forms was observed. We propose that the mutant phenotype of psd reflects defects in developmental events and cell/tissue types that require elevated levels of protein synthesis and are therefore acutely sensitive to a reduction in tRNA export.

  5. Saccharomyces cerevisiae aldolase mutants.

    PubMed Central

    Lobo, Z

    1984-01-01

    Six mutants lacking the glycolytic enzyme fructose 1,6-bisphosphate aldolase have been isolated in the yeast Saccharomyces cerevisiae by inositol starvation. The mutants grown on gluconeogenic substrates, such as glycerol or alcohol, and show growth inhibition by glucose and related sugars. The mutations are recessive, segregate as one gene in crosses, and fall in a single complementation group. All of the mutants synthesize an antigen cross-reacting to the antibody raised against yeast aldolase. The aldolase activity in various mutant alleles measured as fructose 1,6-bisphosphate cleavage is between 1 to 2% and as condensation of triose phosphates to fructose 1,6-bisphosphate is 2 to 5% that of the wild-type. The mutants accumulate fructose 1,6-bisphosphate from glucose during glycolysis and dihydroxyacetone phosphate during gluconeogenesis. This suggests that the aldolase activity is absent in vivo. PMID:6384192

  6. Arabidopsis cell-free extract, ACE, a new in vitro translation system derived from Arabidopsis callus cultures.

    PubMed

    Murota, Katsunori; Hagiwara-Komoda, Yuka; Komoda, Keisuke; Onouchi, Hitoshi; Ishikawa, Masayuki; Naito, Satoshi

    2011-08-01

    The analysis of post-transcriptional regulatory mechanisms in plants has benefited greatly from the use of cell-free extract systems. Arabidopsis as a model system provides extensive genetic resources; however, to date a suitable cell-free translation system from Arabidopsis has not been available. In this study, we devised an Arabidopsis cell-free extract (ACE) to be used for in vitro translation studies. Protoplasts were prepared from callus cultures derived from Arabidopsis seedlings, and cell-free extracts were prepared after evacuolation of the protoplasts by Percoll gradient centrifugation. The new ACE system exhibits translation activity comparable with that of the wheat germ extract system. We demonstrated that ACE prepared from the 5'-3' exoribonuclease-deficient mutant of Arabidopsis, xrn4-5, exhibited increased stability of an uncapped mRNA as compared with that from wild-type Arabidopsis. We applied the ACE system to study post-transcriptional regulation of AtCGS1. AtCGS1 codes for cystathionine γ-synthase (CGS) that catalyzes the first committed step of methionine and S-adenosyl-l-methionine (AdoMet) biosynthesis in plants, and is feedback regulated by mRNA degradation coupled with translation elongation arrest. The ACE system was capable of reproducing translation elongation arrest and subsequent AtCGS1 mRNA degradation that are induced by AdoMet. The ACE system described here can be prepared in a month after seed sowing and will make it possible to study post-transcriptional regulation of plant genes while taking advantage of the genetics of Arabidopsis.

  7. Proteomics and transcriptomics analyses of Arabidopsis floral buds uncover important functions of ARABIDOPSIS SKP1-LIKE1

    SciTech Connect

    Lu, Dihong; Ni, Weimin; Stanley, Bruce A.; Ma, Hong

    2016-03-03

    The ARABIDOPSIS SKP1-LIKE1 (ASK1) protein functions as a subunit of SKP1-CUL1-F-box (SCF) E3 ubiquitin ligases. Previous genetic studies showed that ASK1 plays important roles in Arabidopsis flower development and male meiosis. However, the molecular impact of ASK1-containing SCF E3 ubiquitin ligases (ASK1-E3s) on the floral proteome and transcriptome is unknown. Here we identified proteins that are potentially regulated by ASK1-E3s by comparing floral bud proteomes of wild-type and the ask1 mutant plants. More than 200 proteins were detected in the ask1 mutant but not in wild-type and >300 were detected at higher levels in the ask1 mutant than in wild-type, but their RNA levels were not significantly different between wild-type and ask1 floral buds as shown by transcriptomics analysis, suggesting that they are likely regulated at the protein level by ASK1-E3s. Integrated analyses of floral proteomics and transcriptomics of ask1 and wild-type uncovered several potential aspects of ASK1-E3 functions, including regulation of transcription regulators, kinases, peptidases, and ribosomal proteins, with implications on possible mechanisms of ASK1-E3 functions in floral development. In conclusion, our results suggested that ASK1-E3s play important roles in Arabidopsis protein degradation during flower development. This study opens up new possibilities for further functional studies of these candidate E3 substrates.

  8. Verticillium dahliae toxins-induced nitric oxide production in Arabidopsis is major dependent on nitrate reductase.

    PubMed

    Shi, Fu-Mei; Li, Ying-Zhang

    2008-01-31

    The source of nitric oxide (NO) in plants is unclear and it has been reported NO can be produced by nitric oxide synthase (NOS) like enzymes and by nitrate reductase (NR). Here we used wild-type, Atnos1 mutant and nia1, nia2 NR-deficient mutant plants of Arabidopsis thaliana to investigate the potential source of NO production in response to Verticillium dahliae toxins (VD-toxins). The results revealed that NO production is much higher in wild-type and Atnos1 mutant than in nia1, nia2 NR-deficient mutants. The NR inhibitor had a significant effect on VD-toxins-induced NO production; whereas NOS inhibitor had a slight effect. NR activity was significantly implicated in NO production. The results indicated that as NO was induced in response to VD-toxins in Arabidopsis, the major source was the NR pathway. The production of NOS-system appeared to be secondary.

  9. Arabidopsis myrosinases link the glucosinolate-myrosinase system and the cuticle.

    PubMed

    Ahuja, Ishita; de Vos, Ric C H; Rohloff, Jens; Stoopen, Geert M; Halle, Kari K; Ahmad, Samina Jam Nazeer; Hoang, Linh; Hall, Robert D; Bones, Atle M

    2016-12-15

    Both physical barriers and reactive phytochemicals represent two important components of a plant's defence system against environmental stress. However, these two defence systems have generally been studied independently. Here, we have taken an exclusive opportunity to investigate the connection between a chemical-based plant defence system, represented by the glucosinolate-myrosinase system, and a physical barrier, represented by the cuticle, using Arabidopsis myrosinase (thioglucosidase; TGG) mutants. The tgg1, single and tgg1 tgg2 double mutants showed morphological changes compared to wild-type plants visible as changes in pavement cells, stomatal cells and the ultrastructure of the cuticle. Extensive metabolite analyses of leaves from tgg mutants and wild-type Arabidopsis plants showed altered levels of cuticular fatty acids, fatty acid phytyl esters, glucosinolates, and indole compounds in tgg single and double mutants as compared to wild-type plants. These results point to a close and novel association between chemical defence systems and physical defence barriers.

  10. Suppression of Arabidopsis flowering by near-null magnetic field is affected by light.

    PubMed

    Xu, Chunxiao; Li, Yue; Yu, Yang; Zhang, Yuxia; Wei, Shufeng

    2015-09-01

    We previously reported that a near-null magnetic field suppressed Arabidopsis flowering in white light, which might be related to the function modification of cryptochrome (CRY). To further demonstrate that the effect of near-null magnetic field on Arabidopsis flowering is associated with CRY, Arabidopsis wild type and CRY mutant plants were grown in the near-null magnetic field under blue or red light with different light cycle and photosynthetic photon flux density. We found that Arabidopsis flowering was significantly suppressed by near-null magnetic field in blue light with lower intensity (10 µmol/m(2) /s) and shorter cycle (12 h period: 6 h light/6 h dark). However, flowering time of CRY1/CRY2 mutants did not show any difference between plants grown in near-null magnetic field and in local geomagnetic field under detected light conditions. In red light, no significant difference was shown in Arabidopsis flowering between plants in near-null magnetic field and local geomagnetic field under detected light cycles and intensities. These results suggest that changes of blue light cycle and intensity alter the effect of near-null magnetic field on Arabidopsis flowering, which is mediated by CRY.

  11. Handling Arabidopsis plants: growth, preservation of seeds, transformation, and genetic crosses.

    PubMed

    Rivero, Luz; Scholl, Randy; Holomuzki, Nicholas; Crist, Deborah; Grotewold, Erich; Brkljacic, Jelena

    2014-01-01

    Growing healthy plants is essential for the advancement of Arabidopsis thaliana (Arabidopsis) research. Over the last 20 years, the Arabidopsis Biological Resource Center (ABRC) has collected and developed a series of best-practice protocols, some of which are presented in this chapter. Arabidopsis can be grown in a variety of locations, growth media, and environmental conditions. Most laboratory accessions and their mutant or transgenic derivatives flower after 4-5 weeks and set seeds after 7-8 weeks, under standard growth conditions (soil, long day, 23 ºC). Some mutant genotypes, natural accessions, and Arabidopsis relatives require strict control of growth conditions best provided by growth rooms, chambers, or incubators. Other lines can be grown in less-controlled greenhouse settings. Although the majority of lines can be grown in soil, certain experimental purposes require utilization of sterile solid or liquid growth media. These include the selection of primary transformants, identification of homozygous lethal individuals in a segregating population, or bulking of a large amount of plant material. The importance of controlling, observing, and recording growth conditions is emphasized and appropriate equipment required to perform monitoring of these conditions is listed. Proper conditions for seed harvesting and preservation, as well as seed quality control, are also described. Plant transformation and genetic crosses, two of the methods that revolutionized Arabidopsis genetics, are introduced as well.

  12. P-proteins in Arabidopsis are heteromeric structures involved in rapid sieve tube sealing.

    PubMed

    Jekat, Stephan B; Ernst, Antonia M; von Bohl, Andreas; Zielonka, Sascia; Twyman, Richard M; Noll, Gundula A; Prüfer, Dirk

    2013-01-01

    Structural phloem proteins (P-proteins) are characteristic components of the sieve elements in all dicotyledonous and many monocotyledonous angiosperms. Tobacco P-proteins were recently confirmed to be encoded by the widespread sieve element occlusion (SEO) gene family, and tobacco SEO proteins were shown to be directly involved in sieve tube sealing thus preventing the loss of photosynthate. Analysis of the two Arabidopsis SEO proteins (AtSEOa and AtSEOb) indicated that the corresponding P-protein subunits do not act in a redundant manner. However, there are still pending questions regarding the interaction properties and specific functions of AtSEOa and AtSEOb as well as the general function of structural P-proteins in Arabidopsis. In this study, we characterized the Arabidopsis P-proteins in more detail. We used in planta bimolecular fluorescence complementation assays to confirm the predicted heteromeric interactions between AtSEOa and AtSEOb. Arabidopsis mutants depleted for one or both AtSEO proteins lacked the typical P-protein structures normally found in sieve elements, underlining the identity of AtSEO proteins as P-proteins and furthermore providing the means to determine the role of Arabidopsis P-proteins in sieve tube sealing. We therefore developed an assay based on phloem exudation. Mutants with reduced AtSEO expression levels lost twice as much photosynthate following injury as comparable wild-type plants, confirming that Arabidopsis P-proteins are indeed involved in sieve tube sealing.

  13. Identification of the Arabidopsis CHL3 gene as the nitrate reductase structural gene NIA2.

    PubMed Central

    Wilkinson, J Q; Crawford, N M

    1991-01-01

    Chlorate, the chlorine analog of nitrate, is a herbicide that has been used to select mutants impaired in the process of nitrate assimilation. In Arabidopsis thaliana, mutations at any one of eight distinct loci confer resistance to chlorate. The molecular identities of the genes at these loci are not known; however, one of these loci--chl3--maps very near the nitrate reductase structural gene NIA2. Through the isolation, characterization, and genetic analysis of new chlorate-resistant mutants generated by gamma irradiation, we have been able to demonstrate that the CHL3 gene and the NIA2 gene are identical. Three new chlorate-resistant mutants were identified that had deletions of the entire NIA2 gene. These nia2 null mutants were viable and still retained 10% of wild-type nitrate reductase activity in the leaves of the plants. All three deletion mutations were found to be new alleles of chl3. Introduction of the NIA2 gene back into these chl3 mutants by Agrobacterium-mediated transformation partially complemented their mutant phenotype. From these data, we conclude that Arabidopsis has at least two functional nitrate reductase genes and that the NIA2 gene product accounts for the majority of the leaf nitrate reductase activity and chlorate sensitivity of Arabidopsis plants. PMID:1840922

  14. Multi-Element Bioimaging of Arabidopsis thaliana Roots1[OPEN

    PubMed Central

    Salt, David E.

    2016-01-01

    Better understanding of root function is central for the development of plants with more efficient nutrient uptake and translocation. We here present a method for multielement bioimaging at the cellular level in roots of the genetic model system Arabidopsis (Arabidopsis thaliana). Using conventional protocols for microscopy, we observed that diffusible ions such as potassium and sodium were lost during sample dehydration. Thus, we developed a protocol that preserves ions in their native, cellular environment. Briefly, fresh roots are encapsulated in paraffin, cryo-sectioned, and freeze dried. Samples are finally analyzed by laser ablation-inductively coupled plasma-mass spectrometry, utilizing a specially designed internal standard procedure. The method can be further developed to maintain the native composition of proteins, enzymes, RNA, and DNA, making it attractive in combination with other omics techniques. To demonstrate the potential of the method, we analyzed a mutant of Arabidopsis unable to synthesize the metal chelator nicotianamine. The mutant accumulated substantially more zinc and manganese than the wild type in the tissues surrounding the vascular cylinder. For iron, the images looked completely different, with iron bound mainly in the epidermis of the wild-type plants but confined to the cortical cell walls of the mutant. The method offers the power of inductively coupled plasma-mass spectrometry to be fully employed, thereby providing a basis for detailed studies of ion transport in roots. Being applicable to Arabidopsis, the molecular and genetic approaches available in this system can now be fully exploited in order to gain a better mechanistic understanding of these processes. PMID:27566167

  15. Overexpression of several Arabidopsis histone genes increases Agrobacterium-medicated transformation and transgene expression in plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Arabidopsis histone H2A-1 is important for Agrobacterium-mediated plant transformation. Mutation of HTA1, the gene encoding histone H2A-1, in the rat5 mutant results in decreased T-(transferred) DNA integration into the plant genome, whereas over-expression of HTA1 increases transformation freq...

  16. Putrescine regulating by stress-responsive MAPK cascade contributes to bacterial pathogen defense in Arabidopsis.

    PubMed

    Kim, Su-Hyun; Kim, Sun-Hwa; Yoo, Seung-Jin; Min, Kwang-Hyun; Nam, Seung-Hee; Cho, Baik Ho; Yang, Kwang-Yeol

    2013-08-09

    Polyamines in plants are involved in various physiological and developmental processes including abiotic and biotic stress responses. We investigated the expression of ADCs, which are key enzymes in putrescine (Put) biosynthesis, and roles of Put involving defense response in Arabidopsis. The increased expression of ADC1 and ADC2, and the induction of Put were detected in GVG-NtMEK2(DD) transgenic Arabidopsis, whereas, their performance was partially compromised in GVG-NtMEK2(DD)/mpk3 and GVG-NtMEK2(DD)/mpk6 mutant following DEX treatment. The expression of ADC2 was highly induced by Pst DC3000 inoculation, while the transcript levels of ADC1 were slightly up-regulated. Compared to the WT plant, Put content in the adc2 knock-out mutant was reduced after Pst DC3000 inoculation, and showed enhanced susceptibility to pathogen infection. The adc2 mutant exhibited reduced expression of PR-1 after bacterial infection and the growth of the pathogen was about 4-fold more than that in the WT plant. Furthermore, the disease susceptibility of the adc2 mutant was recovered by the addition of exogenous Put. Taken together, these results suggest that Arabidopsis MPK3 and MPK6 play a positive role in the regulation of Put biosynthesis, and that Put contributes to bacterial pathogen defense in Arabidopsis.

  17. Isolation of cDNAs encoding GTP cyclohydrolase II from Arabidopsis thaliana.

    PubMed

    Kobayashi, M; Sugiyama, M; Yamamoto, K

    1995-07-28

    A GTP cyclohydrolase II-encoding gene from Arabidopsis thaliana was isolated through functional complementation of a mutant of Escherichia coli, BSV18, deficient in this protein. The derived amino-acid sequence constitutes a polypeptide of 27 kDa and shows 37-58% identity with previously published sequences of Escherichia coli, Bacillus subtilis, Photobacterium leiognathi and P. phosphoreum.

  18. Carbon monoxide interacts with auxin and nitric oxide to cope with iron deficiency in Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To clarify the roles of CO, NO and auxin in the plant response to iron deficiency and to establish how the signaling molecules interact to enhance Fe acquisition, we conducted physiological, genetic, and molecular analyses that compared the responses of various Arabidopsis mutants, including hy1 (CO...

  19. Arabidopsis Hormone Database: a comprehensive genetic and phenotypic information database for plant hormone research in Arabidopsis.

    PubMed

    Peng, Zhi-yu; Zhou, Xin; Li, Linchuan; Yu, Xiangchun; Li, Hongjiang; Jiang, Zhiqiang; Cao, Guangyu; Bai, Mingyi; Wang, Xingchun; Jiang, Caifu; Lu, Haibin; Hou, Xianhui; Qu, Lijia; Wang, Zhiyong; Zuo, Jianru; Fu, Xiangdong; Su, Zhen; Li, Songgang; Guo, Hongwei

    2009-01-01

    Plant hormones are small organic molecules that influence almost every aspect of plant growth and development. Genetic and molecular studies have revealed a large number of genes that are involved in responses to numerous plant hormones, including auxin, gibberellin, cytokinin, abscisic acid, ethylene, jasmonic acid, salicylic acid, and brassinosteroid. Here, we develop an Arabidopsis hormone database, which aims to provide a systematic and comprehensive view of genes participating in plant hormonal regulation, as well as morphological phenotypes controlled by plant hormones. Based on data from mutant studies, transgenic analysis and gene ontology (GO) annotation, we have identified a total of 1026 genes in the Arabidopsis genome that participate in plant hormone functions. Meanwhile, a phenotype ontology is developed to precisely describe myriad hormone-regulated morphological processes with standardized vocabularies. A web interface (http://ahd.cbi.pku.edu.cn) would allow users to quickly get access to information about these hormone-related genes, including sequences, functional category, mutant information, phenotypic description, microarray data and linked publications. Several applications of this database in studying plant hormonal regulation and hormone cross-talk will be presented and discussed.

  20. NFXL2 modifies cuticle properties in Arabidopsis.

    PubMed

    Lisso, Janina; Schröder, Florian; Schippers, Jos H M; Müssig, Carsten

    2012-05-01

    Loss of the Arabidopsis NFX1-LIKE2 (NFXL2) gene (At5g05660) results in elevated ABA levels, elevated hydrogen peroxide levels, reduced stomatal aperture, and enhanced drought stress tolerance. Introduction of the NFXL2-78 isoform into the nfxl2-1 mutant is largely sufficient for complementation of the phenotype. We show here that cuticular properties are altered in the nfxl2-1 mutant. The NFXL2-78 protein binds to the SHINE1 (SHN1), SHN2, SHN3, and BODYGUARD1 (BDG1) promoters and mediates weaker expression of these genes. The SHN AP2 domain transcription factors influence cuticle properties. Stronger SHN1, SHN2, and SHN3 expression in the nfxl2-1 mutant may cause altered cuticle properties including reduced stomatal density, and partly explain the enhanced drought stress tolerance. The BDG1 protein also controls cuticle development and is essential for osmotic stress regulation of ABA biosynthesis. Stronger BDG1 expression in nfxl2-1 plants may allow elevated ABA accumulation under drought stress. We conclude that the NFXL2-78 protein is part of a regulatory network that integrates the biosynthesis and action of ABA, ROS, and cuticle components.

  1. PDGFRA-mutant syndrome.

    PubMed

    Ricci, Riccardo; Martini, Maurizio; Cenci, Tonia; Carbone, Arnaldo; Lanza, Paola; Biondi, Alberto; Rindi, Guido; Cassano, Alessandra; Larghi, Alberto; Persiani, Roberto; Larocca, Luigi M

    2015-07-01

    Germline PDGFRA mutations cause multiple heterogeneous gastrointestinal mesenchymal tumors. In its familial form this disease, which was formerly termed intestinal neurofibromatosis/neurofibromatosis 3b (INF/NF3b), has been included among familial gastrointestinal stromal tumors (GISTs) because of its genotype, described when GIST was the only known PDGFRA-mutant gastrointestinal tumor. Shortly afterwards, however, inflammatory fibroid polyps also revealed PDGFRA mutations. Subsequently, gastrointestinal CD34+ 'fibrous tumors' of uncertain classification were described in a germline PDGFRA-mutant context. Our aim was to characterize the syndrome produced by germline PDGFRA mutations and establish diagnostic criteria and management strategies for this hitherto puzzling disease. We studied a kindred displaying multiple gastrointestinal mesenchymal tumors, comparing it with published families/individuals with possible analogous conditions. We identified a novel inherited PDGFRA mutation (P653L), constituting the third reported example of familial PDGFRA mutation. In adult mutants we detected inflammatory fibroid polyps, gastric GISTs and gastrointestinal fibrous tumors of uncertain nosology. We demonstrate that the syndrome formerly defined as INF/NF3b (exemplified by the family reported herein) is simplistically considered a form of familial GIST, because inflammatory fibroid polyps often prevail. Fibrous tumors appear variants of inflammatory fibroid polyps. 'INF/NF3b' and 'familial GIST' are misleading terms which we propose changing to 'PDGFRA-mutant syndrome'. In this condition, unlike KIT-dependent familial GIST syndromes, if present, GISTs are stomach-restricted and diffuse Cajal cell hyperplasia is not observed. This restriction of GISTs to the stomach in PDGFRA-mutant syndrome: (i) focuses oncological concern on gastric masses, as inflammatory fibroid polyps are benign; (ii) supports a selective role of gastric environment for PDGFRA mutations to elicit GISTs

  2. Reduced gravitropic sensitivity in roots of a starch-deficient mutant of Nicotiana sylvestris

    NASA Technical Reports Server (NTRS)

    Kiss, J. Z.; Sack, F. D.

    1989-01-01

    Gravitropism was studied in seedlings of Nicotiana sylvestris Speg. et Comes wild-type (WT) and mutant NS 458 which has a defective plastid phosphoglucomutase (EC 2.7.5.1.). Starch was greatly reduced in NS 458 compared to the WT, but small amounts of starch were detected in rootcap columella cells in NS 458 by light and electron microscopy. The roots of WT are more sensitive to gravity than mutant NS 458 roots since: (1) in mutant roots, curvature was reduced and delayed in the time course of curvature; (2) curvature of mutant roots was 24-56% that of WT roots over the range of induction periods tested; (3) in intermittent-stimulation experiments, curvature of mutant roots was 37% or less than that of WT roots in all treatments tested. The perception time, determined by intermittent-stimulation experiments, was < or = 5 s for WT roots and 30-60 s for mutant roots. The growth rates for WT and NS 458 roots were essentially equal. These results and our previous results with WT and starchless mutant Arabidopsis roots (Kiss et al. 1989, Planta 177, 198-206) support the conclusions that a full complement of starch is necessary for full gravitropic sensitivity and that amyloplasts function in gravity perception. Since a presumed relatively small increase in plastid buoyant mass (N. sylvestris mutant versus Arabidopsis mutant) significantly improves the orientation of the N. sylvestris mutant roots, we suggest that plastids are the likeliest candidates to be triggering gravity perception in roots of both mutants.

  3. Dissecting iron deficiency-induced proton extrusion in Arabidopsis roots.

    PubMed

    Santi, Simonetta; Schmidt, Wolfgang

    2009-01-01

    Here, we have analysed the H(+)-ATPase-mediated extrusion of protons across the plasma membrane (PM) of rhizodermic cells, a process that is inducible by iron (Fe) deficiency and thought to serve in the mobilization of sparingly soluble Fe sources. The induction and function of Fe-responsive PM H(+)-ATPases in Arabidopsis roots was investigated by gene expression analysis and by using mutants defective in the expression or function of one of the isogenes. In addition, the expression of the most responsive isogenes was investigated in natural Arabidopsis accessions that have been selected for their in vivo proton extrusion activity. Our data suggest that the rhizosphere acidification in response to Fe deficiency is chiefly mediated by AHA2, while AHA1 functions as a housekeeping isoform. The aha7 knock-out mutant plants showed a reduced frequency of root hairs, suggesting an involvement of AHA7 in the differentiation of rhizodermic cells. Acidification capacity varied among Arabidopsis accessions and was associated with a high induction of AHA2 and IRT1, a high relative growth rate and a shoot-root ratio that was unaffected by the external Fe supply. An effective regulation of the Fe-responsive genes and a stable shoot-root ratio may represent important characteristics for the Fe uptake efficiency.

  4. Oviposition by pierid butterflies triggers defense responses in Arabidopsis.

    PubMed

    Little, Dawn; Gouhier-Darimont, Caroline; Bruessow, Friederike; Reymond, Philippe

    2007-02-01

    Insect eggs represent a threat for the plant as hatching larvae rapidly start with their feeding activity. Using a whole-genome microarray, we studied the expression profile of Arabidopsis (Arabidopsis thaliana) leaves after oviposition by two pierid butterflies. For Pieris brassicae, the deposition of egg batches changed the expression of hundreds of genes over a period of 3 d after oviposition. The transcript signature was similar to that observed during a hypersensitive response or in lesion-mimic mutants, including the induction of defense and stress-related genes and the repression of genes involved in growth and photosynthesis. Deposition of single eggs by Pieris rapae caused a similar although much weaker transcriptional response. Analysis of the jasmonic acid and salicylic acid mutants coi1-1 and sid2-1 indicated that the response to egg deposition is mostly independent of these signaling pathways. Histochemical analyses showed that egg deposition is causing a localized cell death, accompanied by the accumulation of callose, and the production of reactive oxygen species. In addition, activation of the pathogenesis-related1::beta-glucuronidase reporter gene correlated precisely with the site of egg deposition and was also triggered by crude egg extract. This study provides molecular evidence for the detection of egg deposition by Arabidopsis plants and suggests that oviposition causes a localized response with strong similarity to a hypersensitive response.

  5. Descendants of primed Arabidopsis plants exhibit resistance to biotic stress.

    PubMed

    Slaughter, Ana; Daniel, Xavier; Flors, Victor; Luna, Estrella; Hohn, Barbara; Mauch-Mani, Brigitte

    2012-02-01

    An attack of plants by pathogens or treatment with certain resistance-inducing compounds can lead to the establishment of a unique primed state of defense. Primed plants show enhanced defense reactions upon further challenge with biotic or abiotic stress. Here, we report that the primed state in Arabidopsis (Arabidopsis thaliana) is still functional in the next generation without additional treatment. We compared the reactions of Arabidopsis plants that had been either primed with β-amino-butyric acid (BABA) or with an avirulent isolate of the bacteria Pseudomonas syringae pv tomato (PstavrRpt2). The descendants of primed plants showed a faster and higher accumulation of transcripts of defense-related genes in the salicylic acid signaling pathway and enhanced disease resistance upon challenge inoculation with a virulent isolate of P. syringae. In addition, the progeny of primed plants was also more resistant against the oomycete pathogen Hyaloperonospora arabidopsidis. When transgenerationally primed plants were subjected to an additional priming treatment, their descendants displayed an even stronger primed phenotype, suggesting that plants can inherit a sensitization for the priming phenomenon. Interestingly, this primed to be primed phenotype was much reduced in the Arabidopsis β-amino-butyric acid priming mutant ibs1 (induced BABA sterility1). Our results demonstrate that the primed state of plants is transferred to their progeny and confers improved protection from pathogen attack as compared to the descendants of unprimed plants.

  6. Glutamate functions in stomatal closure in Arabidopsis and fava bean.

    PubMed

    Yoshida, Riichiro; Mori, Izumi C; Kamizono, Nobuto; Shichiri, Yudai; Shimatani, Tetsuo; Miyata, Fumika; Honda, Kenji; Iwai, Sumio

    2016-01-01

    Guard cells are indispensable for higher plants because they control gas exchange and water balance to maintain photosynthetic activity. The signaling processes that govern their movement are controlled by several factors, such as abscisic acid (ABA), blue light, pathogen-associated molecular patterns (PAMPs), and carbon dioxide. Herein, we demonstrated that the amino acid glutamate (Glu), a well-known mammalian neurotransmitter, functions as a novel signaling molecule in stomatal closure in both Arabidopsis and fava bean (Vicia faba L.). Pharmacological and electrophysiological analyses provided important clues for the participation of Glu-receptors, Ca(2+), and protein phosphorylation during the signaling process. Genetic analyses using Arabidopsis ABA-deficient (aba2-1) and ABA-insensitive (abi1-1 and abi2-1) mutants showed that ABA is not required for Glu signaling. However, loss-of-function of the Arabidopsis gene encoding Slow Anion Channel-Associated 1 (SLAC1) and Calcium-Dependent Protein Kinase 6 (CPK6) impaired the Glu response. Moreover, T-DNA knockout mutations of the Arabidopsis Glu receptor-like gene (GLR), GLR3.5, lost their sensitivity to Glu-dependent stomatal closure. Our results strongly support functional Glu-signaling in stomatal closure and the crucial roles of GLRs in this signaling process.

  7. Conservation of floral homeotic gene function between Arabidopsis and antirrhinum.

    PubMed Central

    Irish, V F; Yamamoto, Y T

    1995-01-01

    Several homeotic genes controlling floral development have been isolated in both Antirrhinum and Arabidopsis. Based on the similarities in sequence and in the phenotypes elicited by mutations in some of these genes, it has been proposed that the regulatory hierarchy controlling floral development is comparable in these two species. We have performed a direct experimental test of this hypothesis by introducing a chimeric Antirrhinum Deficiens (DefA)/Arabidopsis APETALA3 (AP3) gene, under the control of the Arabidopsis AP3 promoter, into Arabidopsis. We demonstrated that this transgene is sufficient to partially complement severe mutations at the AP3 locus. In combination with a weak ap3 mutation, this transgene is capable of completely rescuing the mutant phenotype to a fully functional wild-type flower. These observations indicate that despite differences in DNA sequence and expression, DefA coding sequences can compensate for the loss of AP3 gene function. We discuss the implications of these results for the evolution of homeotic gene function in flowering plants. PMID:7580255

  8. Plant chip for high-throughput phenotyping of Arabidopsis.

    PubMed

    Jiang, Huawei; Xu, Zhen; Aluru, Maneesha R; Dong, Liang

    2014-04-07

    We report on the development of a vertical and transparent microfluidic chip for high-throughput phenotyping of Arabidopsis thaliana plants. Multiple Arabidopsis seeds can be germinated and grown hydroponically over more than two weeks in the chip, thus enabling large-scale and quantitative monitoring of plant phenotypes. The novel vertical arrangement of this microfluidic device not only allows for normal gravitropic growth of the plants but also, more importantly, makes it convenient to continuously monitor phenotypic changes in plants at the whole organismal level, including seed germination and root and shoot growth (hypocotyls, cotyledons, and leaves), as well as at the cellular level. We also developed a hydrodynamic trapping method to automatically place single seeds into seed holding sites of the device and to avoid potential damage to seeds that might occur during manual loading. We demonstrated general utility of this microfluidic device by showing clear visible phenotypes of the immutans mutant of Arabidopsis, and we also showed changes occurring during plant-pathogen interactions at different developmental stages. Arabidopsis plants grown in the device maintained normal morphological and physiological behaviour, and distinct phenotypic variations consistent with a priori data were observed via high-resolution images taken in real time. Moreover, the timeline for different developmental stages for plants grown in this device was highly comparable to growth using a conventional agar plate method. This prototype plant chip technology is expected to lead to the establishment of a powerful experimental and cost-effective framework for high-throughput and precise plant phenotyping.

  9. Multiplex micro-respiratory measurements of Arabidopsis tissues.

    PubMed

    Sew, Yun Shin; Ströher, Elke; Holzmann, Cristián; Huang, Shaobai; Taylor, Nicolas L; Jordana, Xavier; Millar, A Harvey

    2013-11-01

    Researchers often want to study the respiratory properties of individual parts of plants in response to a range of treatments. Arabidopsis is an obvious model for this work; however, because of its size, it represents a challenge for gas exchange measurements of respiration. The combination of micro-respiratory technologies with multiplex assays has the potential to bridge this gap, and make measurements possible in this model plant species. We show the adaptation of the commercial technology used for mammalian cell respiration analysis to study three critical tissues of interest: leaf sections, root tips and seeds. The measurement of respiration in single leaf discs has allowed the age dependence of the respiration rate in Arabidopsis leaves across the rosette to be observed. The oxygen consumption of single root tips from plate-grown seedlings shows the enhanced respiration of root tips and their time-dependent susceptibility to salinity. The monitoring of single Arabidopsis seeds shows the kinetics of respiration over 48 h post-imbibition, and the effect of the phytohormones gibberellic acid (GA3 ) and abscisic acid (ABA) on respiration during seed germination. These studies highlight the potential for multiplexed micro-respiratory assays to study oxygen consumption in Arabidopsis tissues, and open up new possibilities to screen and study mutants and to identify differences in ecotypes or populations of different plant species.

  10. The ethylene response pathway in Arabidopsis

    NASA Technical Reports Server (NTRS)

    Kieber, J. J.; Evans, M. L. (Principal Investigator)

    1997-01-01

    The simple gas ethylene influences a diverse array of plant growth and developmental processes including germination, senescence, cell elongation, and fruit ripening. This review focuses on recent molecular genetic studies, principally in Arabidopsis, in which components of the ethylene response pathway have been identified. The isolation and characterization of two of these genes has revealed that ethylene sensing involves a protein kinase cascade. One of these genes encodes a protein with similarity to the ubiquitous Raf family of Ser/Thr protein kinases. A second gene shows similarity to the prokaryotic two-component histidine kinases and most likely encodes an ethylene receptor. Additional elements involved in ethylene signaling have only been identified genetically. The characterization of these genes and mutants will be discussed.

  11. MTHFD1 controls DNA methylation in Arabidopsis

    PubMed Central

    Groth, Martin; Moissiard, Guillaume; Wirtz, Markus; Wang, Haifeng; Garcia-Salinas, Carolina; Ramos-Parra, Perla A.; Bischof, Sylvain; Feng, Suhua; Cokus, Shawn J.; John, Amala; Smith, Danielle C.; Zhai, Jixian; Hale, Christopher J.; Long, Jeff A.; Hell, Ruediger; Díaz de la Garza, Rocío I.; Jacobsen, Steven E.

    2016-01-01

    DNA methylation is an epigenetic mechanism that has important functions in transcriptional silencing and is associated with repressive histone methylation (H3K9me). To further investigate silencing mechanisms, we screened a mutagenized Arabidopsis thaliana population for expression of SDCpro-GFP, redundantly controlled by DNA methyltransferases DRM2 and CMT3. Here, we identify the hypomorphic mutant mthfd1-1, carrying a mutation (R175Q) in the cytoplasmic bifunctional methylenetetrahydrofolate dehydrogenase/methenyltetrahydrofolate cyclohydrolase (MTHFD1). Decreased levels of oxidized tetrahydrofolates in mthfd1-1 and lethality of loss-of-function demonstrate the essential enzymatic role of MTHFD1 in Arabidopsis. Accumulation of homocysteine and S-adenosylhomocysteine, genome-wide DNA hypomethylation, loss of H3K9me and transposon derepression indicate that S-adenosylmethionine-dependent transmethylation is inhibited in mthfd1-1. Comparative analysis of DNA methylation revealed that the CMT3 and CMT2 pathways involving positive feedback with H3K9me are mostly affected. Our work highlights the sensitivity of epigenetic networks to one-carbon metabolism due to their common S-adenosylmethionine-dependent transmethylation and has implications for human MTHFD1-associated diseases. PMID:27291711

  12. Local Evolution of Seed Flotation in Arabidopsis

    PubMed Central

    Saez-Aguayo, Susana; Rondeau-Mouro, Corinne; Macquet, Audrey; Kronholm, Ilkka; Ralet, Marie-Christine; Berger, Adeline; Sallé, Christine; Poulain, Damien; Granier, Fabienne; Botran, Lucy; Loudet, Olivier; de Meaux, Juliette; Marion-Poll, Annie; North, Helen M.

    2014-01-01

    Arabidopsis seeds rapidly release hydrophilic polysaccharides from the seed coat on imbibition. These form a heavy mucilage layer around the seed that makes it sink in water. Fourteen natural Arabidopsis variants from central Asia and Scandinavia were identified with seeds that have modified mucilage release and float. Four of these have a novel mucilage phenotype with almost none of the released mucilage adhering to the seed and the absence of cellulose microfibrils. Mucilage release was modified in the variants by ten independent causal mutations in four different loci. Seven distinct mutations affected one locus, coding the MUM2 β-D-galactosidase, and represent a striking example of allelic heterogeneity. The modification of mucilage release has thus evolved a number of times independently in two restricted geographical zones. All the natural mutants identified still accumulated mucilage polysaccharides in seed coat epidermal cells. Using nuclear magnetic resonance (NMR) relaxometry their production and retention was shown to reduce water mobility into internal seed tissues during imbibition, which would help to maintain seed buoyancy. Surprisingly, despite released mucilage being an excellent hydrogel it did not increase the rate of water uptake by internal seed tissues and is more likely to play a role in retaining water around the seed. PMID:24625826

  13. Local evolution of seed flotation in Arabidopsis.

    PubMed

    Saez-Aguayo, Susana; Rondeau-Mouro, Corinne; Macquet, Audrey; Kronholm, Ilkka; Ralet, Marie-Christine; Berger, Adeline; Sallé, Christine; Poulain, Damien; Granier, Fabienne; Botran, Lucy; Loudet, Olivier; de Meaux, Juliette; Marion-Poll, Annie; North, Helen M

    2014-03-01

    Arabidopsis seeds rapidly release hydrophilic polysaccharides from the seed coat on imbibition. These form a heavy mucilage layer around the seed that makes it sink in water. Fourteen natural Arabidopsis variants from central Asia and Scandinavia were identified with seeds that have modified mucilage release and float. Four of these have a novel mucilage phenotype with almost none of the released mucilage adhering to the seed and the absence of cellulose microfibrils. Mucilage release was modified in the variants by ten independent causal mutations in four different loci. Seven distinct mutations affected one locus, coding the MUM2 β-D-galactosidase, and represent a striking example of allelic heterogeneity. The modification of mucilage release has thus evolved a number of times independently in two restricted geographical zones. All the natural mutants identified still accumulated mucilage polysaccharides in seed coat epidermal cells. Using nuclear magnetic resonance (NMR) relaxometry their production and retention was shown to reduce water mobility into internal seed tissues during imbibition, which would help to maintain seed buoyancy. Surprisingly, despite released mucilage being an excellent hydrogel it did not increase the rate of water uptake by internal seed tissues and is more likely to play a role in retaining water around the seed.

  14. Sm-Like Protein-Mediated RNA Metabolism Is Required for Heat Stress Tolerance in Arabidopsis

    PubMed Central

    Okamoto, Masanori; Matsui, Akihiro; Tanaka, Maho; Morosawa, Taeko; Ishida, Junko; Iida, Kei; Mochizuki, Yoshiki; Toyoda, Tetsuro; Seki, Motoaki

    2016-01-01

    Sm-like proteins play multiple functions in RNA metabolism, which is essential for biological processes such as stress responses in eukaryotes. The Arabidopsis thaliana sad1 mutant has a mutation of sm-like protein 5 (LSM5) and shows impaired drought and salt stress tolerances. The lsm5/sad1 mutant also showed hypersensitivity to heat stress. GFP-fused LSM5/SAD1 was localized in the nucleus under optimal growth conditions. After heat stress treatment, GFP-fused LSM5/SAD1 fluorescence was also observed as small cytoplasmic dots, in addition to nuclear localization. Whole genome transcriptome analysis revealed that many genes in Arabidopsis were drastically changed in response to heat stress. More heat-responsive genes were highly expressed in lsm5/sad1 mutant at both 2 and 6 h after heat stress treatment. Additionally, intron-retained and capped transcripts accumulated in the lsm5/sad1 mutant after heat stress treatment. In this study, we also identified non-Arabidopsis Genome Initiative transcripts that were expressed from unannotated regions. Most of these transcripts were antisense transcripts, and many capped non-AGI transcripts accumulated in the lsm5/sad1 mutant during heat stress treatment. These results indicated that LSM5/SAD1 functions to degrade aberrant transcripts through appropriate mRNA splicing and decapping, and precise RNA metabolic machinery is required for heat stress tolerance. PMID:27493656

  15. Remobilization of Phytol from Chlorophyll Degradation Is Essential for Tocopherol Synthesis and Growth of Arabidopsis

    PubMed Central

    vom Dorp, Katharina; Hölzl, Georg; Plohmann, Christian; Eisenhut, Marion; Abraham, Marion

    2015-01-01

    Phytol from chlorophyll degradation can be phosphorylated to phytyl-phosphate and phytyl-diphosphate, the substrate for tocopherol (vitamin E) synthesis. A candidate for the phytyl-phosphate kinase from Arabidopsis thaliana (At1g78620) was identified via a phylogeny-based approach. This gene was designated VITAMIN E DEFICIENT6 (VTE6) because the leaves of the Arabidopsis vte6 mutants are tocopherol deficient. The vte6 mutant plants are incapable of photoautotrophic growth. Phytol and phytyl-phosphate accumulate, and the phytyl-diphosphate content is strongly decreased in vte6 leaves. Phytol feeding and enzyme assays with Arabidopsis and recombinant Escherichia coli cells demonstrated that VTE6 has phytyl-P kinase activity. Overexpression of VTE6 resulted in increased phytyl-diphosphate and tocopherol contents in seeds, indicating that VTE6 encodes phytyl-phosphate kinase. The severe growth retardation of vte6 mutants was partially rescued by introducing the phytol kinase mutation vte5. Double mutant plants (vte5 vte6) are tocopherol deficient and contain more chlorophyll, but reduced amounts of phytol and phytyl-phosphate compared with vte6 mutants, suggesting that phytol or phytyl-phosphate are detrimental to plant growth. Therefore, VTE6 represents the missing phytyl-phosphate kinase, linking phytol release from chlorophyll with tocopherol synthesis. Moreover, tocopherol synthesis in leaves depends on phytol derived from chlorophyll, not on de novo synthesis of phytyl-diphosphate from geranylgeranyl-diphosphate. PMID:26452599

  16. Abscisic acid and blue light signaling pathways in chloroplast movements in Arabidopsis mesophyll.

    PubMed

    Eckstein, Aleksandra; Krzeszowiec, Weronika; Banaś, Agnieszka Katarzyna; Janowiak, Franciszek; Gabryś, Halina

    2016-01-01

    Abscisic acid (ABA) and phototropins act antagonistically to control stomatal movements. Here, we investigated the role of ABA in phototropin-directed chloroplast movements in mesophyll cells of Arabidopsis thaliana. We analyzed the expression of phototropins at mRNA and protein level under the influence of ABA. PHOT1 mRNA level was decreased by ABA in the dark while it was insensitive to ABA in light. PHOT2 mRNA level was independent of the hormone treatment. The levels of phototropin proteins were down-regulated by ABA, both in darkness and light. No impact of exogenous ABA on amplitudes and kinetics of chloroplast movements was detected. Chloroplast responses in wild type Arabidopsis and three mutants, abi4, abi2 (abscisic acid insensitive4, 2) and aba1 (abscisic acid1), were measured to account for endogenous ABA signaling. The chloroplast responses were slightly reduced in abi2 and aba1 mutants in strong light. To further investigate the effect, abi2 and aba1 mutants were supplemented with exogenous ABA. In the aba1 mutant, the reaction was rescued but in abi2 it was unaffected. Our results show that ABA is not directly involved in phototropin-controlled chloroplast responses in mature leaves of Arabidopsis. However, the disturbance of ABA biosynthesis and signaling in mutants affects some elements of the chloroplast movement mechanism. In line with its role as a stress hormone, ABA appears to enhance plant sensitivity to light and promote the chloroplast avoidance response.

  17. Mechanisms that control knox gene expression in the Arabidopsis shoot.

    PubMed

    Ori, N; Eshed, Y; Chuck, G; Bowman, J L; Hake, S

    2000-12-01

    Knotted1-like homeobox (knox) genes are expressed in specific patterns within shoot meristems and play an important role in meristem maintenance. Misexpression of the knox genes, KNAT1 or KNAT2, in Arabidopsis produces a variety of phenotypes, including lobed leaves and ectopic stipules and meristems in the sinus, the region between lobes. We sought to determine the mechanisms that control knox gene expression in the shoot by examining recessive mutants that share phenotypic characteristics with 35S::KNAT1 plants. Double mutants of serrate (se) with either asymmetric1 (as1) or asymmetric2 (as2) showed lobed leaves, ectopic stipules in the sinuses and defects in the timely elongation of sepals, petals and stamens, similar to 35S::KNAT1 plants. Ectopic stipules and in rare cases, ectopic meristems, were detected in the sinuses on plants that were mutant for pickle and either as1 or as2. KNAT1 and KNAT2 were misexpressed in the leaves and flowers of single as1 and as2 mutants and in the sinuses of leaves of the different double mutants, but not in se or pickle single mutants. These results suggest that AS1 and AS2 promote leaf differentiation through repression of knox expression in leaves, and that SE and PKL globally restrict the competence to respond to genes that promote morphogenesis.

  18. Auxin-Mediated Ribosomal Biogenesis Regulates Vacuolar Trafficking in Arabidopsis[W

    PubMed Central

    Rosado, Abel; Sohn, Eun Ju; Drakakaki, Georgia; Pan, Songqin; Swidergal, Alexandra; Xiong, Yuqing; Kang, Byung-Ho; Bressan, Ray A.; Raikhel, Natasha V.

    2010-01-01

    In plants, the mechanisms that regulate the transit of vacuolar soluble proteins containing C-terminal and N-terminal vacuolar sorting determinants (VSDs) to the vacuole are largely unknown. In a screen for Arabidopsis thaliana mutants affected in the trafficking of C-terminal VSD containing proteins, we isolated the ribosomal biogenesis mutant rpl4a characterized by its partial secretion of vacuolar targeted proteins and a plethora of developmental phenotypes derived from its aberrant auxin responses. In this study, we show that ribosomal biogenesis can be directly regulated by auxins and that the exogenous application of auxins to wild-type plants results in vacuolar trafficking defects similar to those observed in rpl4a mutants. We propose that the influence of auxin on ribosomal biogenesis acts as a regulatory mechanism for auxin-mediated developmental processes, and we demonstrate the involvement of this regulatory mechanism in the sorting of vacuolar targeted proteins in Arabidopsis. PMID:20061553

  19. A Cold-Inducible DEAD-Box RNA Helicase from Arabidopsis thaliana Regulates Plant Growth and Development under Low Temperature

    PubMed Central

    Liu, Yuelin; Tabata, Daisuke; Imai, Ryozo

    2016-01-01

    DEAD-box RNA helicases comprise a large family and are involved in a range of RNA processing events. Here, we identified one of the Arabidopsis thaliana DEAD-box RNA helicases, AtRH7, as an interactor of Arabidopsis COLD SHOCK DOMAIN PROTEIN 3 (AtCSP3), which is an RNA chaperone involved in cold adaptation. Promoter:GUS transgenic plants revealed that AtRH7 is expressed ubiquitously and that its levels of the expression are higher in rapidly growing tissues. Knockout mutant lines displayed several morphological alterations such as disturbed vein pattern, pointed first true leaves, and short roots, which resemble ribosome-related mutants of Arabidopsis. In addition, aberrant floral development was also observed in rh7 mutants. When the mutants were germinated at low temperature (12°C), both radicle and first leaf emergence were severely delayed; after exposure of seedlings to a long period of cold, the mutants developed aberrant, fewer, and smaller leaves. RNA blots and circular RT-PCR revealed that 35S and 18S rRNA precursors accumulated to higher levels in the mutants than in WT under both normal and cold conditions, suggesting the mutants are partially impaired in pre-rRNA processing. Taken together, the results suggest that AtRH7 affects rRNA biogenesis and plays an important role in plant growth under cold. PMID:27116354

  20. A Cold-Inducible DEAD-Box RNA Helicase from Arabidopsis thaliana Regulates Plant Growth and Development under Low Temperature.

    PubMed

    Liu, Yuelin; Tabata, Daisuke; Imai, Ryozo

    2016-01-01

    DEAD-box RNA helicases comprise a large family and are involved in a range of RNA processing events. Here, we identified one of the Arabidopsis thaliana DEAD-box RNA helicases, AtRH7, as an interactor of Arabidopsis COLD SHOCK DOMAIN PROTEIN 3 (AtCSP3), which is an RNA chaperone involved in cold adaptation. Promoter:GUS transgenic plants revealed that AtRH7 is expressed ubiquitously and that its levels of the expression are higher in rapidly growing tissues. Knockout mutant lines displayed several morphological alterations such as disturbed vein pattern, pointed first true leaves, and short roots, which resemble ribosome-related mutants of Arabidopsis. In addition, aberrant floral development was also observed in rh7 mutants. When the mutants were germinated at low temperature (12°C), both radicle and first leaf emergence were severely delayed; after exposure of seedlings to a long period of cold, the mutants developed aberrant, fewer, and smaller leaves. RNA blots and circular RT-PCR revealed that 35S and 18S rRNA precursors accumulated to higher levels in the mutants than in WT under both normal and cold conditions, suggesting the mutants are partially impaired in pre-rRNA processing. Taken together, the results suggest that AtRH7 affects rRNA biogenesis and plays an important role in plant growth under cold.

  1. Mutant fatty acid desaturase

    DOEpatents

    Shanklin, John; Cahoon, Edgar B.

    2004-02-03

    The present invention relates to a method for producing mutants of a fatty acid desaturase having a substantially increased activity towards fatty acid substrates with chains containing fewer than 18 carbons relative to an unmutagenized precursor desaturase having an 18 carbon atom chain length substrate specificity. The method involves inducing one or more mutations in the nucleic acid sequence encoding the precursor desaturase, transforming the mutated sequence into an unsaturated fatty acid auxotroph cell such as MH13 E. coli, culturing the cells in the absence of supplemental unsaturated fatty acids, thereby selecting for recipient cells which have received and which express a mutant fatty acid desaturase with an elevated specificity for fatty acid substrates having chain lengths of less than 18 carbon atoms. A variety of mutants having 16 or fewer carbon atom chain length substrate specificities are produced by this method. Mutant desaturases produced by this method can be introduced via expression vectors into prokaryotic and eukaryotic cells and can also be used in the production of transgenic plants which may be used to produce specific fatty acid products.

  2. The mitochondrial monothiol glutaredoxin S15 is essential for iron-sulfur protein maturation in Arabidopsis thaliana

    PubMed Central

    Moseler, Anna; Aller, Isabel; Wagner, Stephan; Nietzel, Thomas; Przybyla-Toscano, Jonathan; Mühlenhoff, Ulrich; Lill, Roland; Berndt, Carsten; Rouhier, Nicolas; Schwarzländer, Markus; Meyer, Andreas J.

    2015-01-01

    The iron-sulfur cluster (ISC) is an ancient and essential cofactor of many proteins involved in electron transfer and metabolic reactions. In Arabidopsis, three pathways exist for the maturation of iron-sulfur proteins in the cytosol, plastids, and mitochondria. We functionally characterized the role of mitochondrial glutaredoxin S15 (GRXS15) in biogenesis of ISC containing aconitase through a combination of genetic, physiological, and biochemical approaches. Two Arabidopsis T-DNA insertion mutants were identified as null mutants with early embryonic lethal phenotypes that could be rescued by GRXS15. Furthermore, we showed that recombinant GRXS15 is able to coordinate and transfer an ISC and that this coordination depends on reduced glutathione (GSH). We found the Arabidopsis GRXS15 able to complement growth defects based on disturbed ISC protein assembly of a yeast Δgrx5 mutant. Modeling of GRXS15 onto the crystal structures of related nonplant proteins highlighted amino acid residues that after mutation diminished GSH and subsequently ISC coordination, as well as the ability to rescue the yeast mutant. When used for plant complementation, one of these mutant variants, GRXS15K83/A, led to severe developmental delay and a pronounced decrease in aconitase activity by approximately 65%. These results indicate that mitochondrial GRXS15 is an essential protein in Arabidopsis, required for full activity of iron-sulfur proteins. PMID:26483494

  3. Arabidopsis NATA1 Acetylates Putrescine and Decreases Defense-Related Hydrogen Peroxide Accumulation1[OPEN

    PubMed Central

    Preuss, Aileen S.

    2016-01-01

    Biosynthesis of the polyamines putrescine, spermidine, and spermine is induced in response to pathogen infection of plants. Putrescine, which is produced from Arg, serves as a metabolic precursor for longer polyamines, including spermidine and spermine. Polyamine acetylation, which has important regulatory functions in mammalian cells, has been observed in several plant species. Here we show that Arabidopsis (Arabidopsis thaliana) N-ACETYLTRANSFERASE ACTIVITY1 (NATA1) catalyzes acetylation of putrescine to N-acetylputrescine and thereby competes with spermidine synthase for a common substrate. NATA1 expression is strongly induced by the plant defense signaling molecule jasmonic acid and coronatine, an effector molecule produced by DC3000, a Pseudomonas syringae strain that initiates a virulent infection in Arabidopsis ecotype Columbia-0. DC3000 growth is reduced in nata1 mutant Arabidopsis, suggesting a role for NATA1-mediated putrescine acetylation in suppressing antimicrobial defenses. During infection by P. syringae and other plant pathogens, polyamine oxidases use spermidine and spermine as substrates for the production of defense-related H2O2. Compared to wild-type Columbia-0 Arabidopsis, the response of nata1mutants to P. syringae infection includes reduced accumulation of acetylputrescine, greater abundance of nonacetylated polyamines, elevated H2O2 production by polyamine oxidases, and higher expression of genes related to pathogen defense. Together, these results are consistent with a model whereby P. syringae growth is improved in a targeted manner through coronatine-induced putrescine acetylation by NATA1. PMID:27208290

  4. Cinnamate-4-hydroxylase expression in Arabidopsis. Regulation in response to development and the environment.

    PubMed Central

    Bell-Lelong, D A; Cusumano, J C; Meyer, K; Chapple, C

    1997-01-01

    Cinnamate-4-hydroxylase (C4H) is the first Cyt P450-dependent monooxygenase of the phenylpropanoid pathway. To study the expression of this gene in Arabidopsis thaliana, a C4H cDNA clone from the Arabidopsis expressed sequence tag database was identified and used to isolate its corresponding genomic clone. The entire C4H coding sequence plus 2.9 kb of its promoter were isolated on a 5.4-kb HindIII fragment of this cosmid. Inspection of the promoter sequence revealed the presence of a number of putative regulatory motifs previously identified in the promoters of other phenylpropanoid pathway genes. The expression of C4H was analyzed by RNA blot hybridization analysis and in transgenic Arabidopsis carrying a C4H-beta-glucuronidase transcriptional fusion. C4H message accumulation was light-dependent, but was detectable even in dark-grown seedlings. Consistent with these data, C4H mRNA was accumulated to light-grown levels in etiolated det1-1 mutant seedlings. C4H is widely expressed in various Arabidopsis tissues, particularly in roots and cells undergoing lignification. The C4H-driven beta-glucuronidase expression accurately reflected the tissue-specificity and wound-inducibility of the C4H promoter indicated by RNA blot hybridization analysis. A modest increase in C4H expression was observed in the tt8 mutant of Arabidopsis. PMID:9085570

  5. Cell wall glucomannan in Arabidopsis is synthesised by CSLA glycosyltransferases, and influences the progression of embryogenesis.

    PubMed

    Goubet, Florence; Barton, Christopher J; Mortimer, Jennifer C; Yu, Xiaolan; Zhang, Zhinong; Miles, Godfrey P; Richens, Jenny; Liepman, Aaron H; Seffen, Keith; Dupree, Paul

    2009-11-01

    Mannans are hemicellulosic polysaccharides that have previously been implicated as structural constituents of cell walls and as storage reserves but which may serve other functions during plant growth and development. Several members of the Arabidopsis cellulose synthase-like A (CSLA) family have previously been shown to synthesise mannan polysaccharides in vitro when heterologously expressed. It has also been found that CSLA7 is essential for embryogenesis, suggesting a role for the CSLA7 product in development. To determine whether the CSLA proteins are responsible for glucomannan synthesis in vivo, we characterised insertion mutants in each of the nine Arabidopsis CSLA genes and several double and triple mutant combinations. csla9 mutants showed substantially reduced glucomannan, and triple csla2csla3csla9 mutants lacked detectable glucomannan in stems. Nevertheless, these mutants showed no alteration in stem development or strength. Overexpression of CSLA2, CSLA7 and CSLA9 increased the glucomannan content in stems. Increased glucomannan synthesis also caused defective embryogenesis, leading to delayed development and occasional embryo death. The embryo lethality of csla7 was complemented by overexpression of CSLA9, suggesting that the glucomannan products are similar. We conclude that CSLA2, CSLA3 and CSLA9 are responsible for the synthesis of all detectable glucomannan in Arabidopsis stems, and that CSLA7 synthesises glucomannan in embryos. These results are inconsistent with a substantial role for glucomannan in wall strength in Arabidopsis stems, but indicate that glucomannan levels affect embryogenesis. Together with earlier heterologous expression studies, the glucomannan deficiency observed in csla mutant plants demonstrates that the CSLA family encodes glucomannan synthases.

  6. A high throughput genetic screen identifies new early meiotic recombination functions in Arabidopsis thaliana.

    PubMed

    De Muyt, Arnaud; Pereira, Lucie; Vezon, Daniel; Chelysheva, Liudmila; Gendrot, Ghislaine; Chambon, Aurélie; Lainé-Choinard, Sandrine; Pelletier, Georges; Mercier, Raphaël; Nogué, Fabien; Grelon, Mathilde

    2009-09-01

    Meiotic recombination is initiated by the formation of numerous DNA double-strand breaks (DSBs) catalysed by the widely conserved Spo11 protein. In Saccharomyces cerevisiae, Spo11 requires nine other proteins for meiotic DSB formation; however, unlike Spo11, few of these are conserved across kingdoms. In order to investigate this recombination step in higher eukaryotes, we took advantage of a high-throughput meiotic mutant screen carried out in the model plant Arabidopsis thaliana. A collection of 55,000 mutant lines was screened, and spo11-like mutations, characterised by a drastic decrease in chiasma formation at metaphase I associated with an absence of synapsis at prophase, were selected. This screen led to the identification of two populations of mutants classified according to their recombination defects: mutants that repair meiotic DSBs using the sister chromatid such as Atdmc1 or mutants that are unable to make DSBs like Atspo11-1. We found that in Arabidopsis thaliana at least four proteins are necessary for driving meiotic DSB repair via the homologous chromosomes. These include the previously characterised DMC1 and the Hop1-related ASY1 proteins, but also the meiotic specific cyclin SDS as well as the Hop2 Arabidopsis homologue AHP2. Analysing the mutants defective in DSB formation, we identified the previously characterised AtSPO11-1, AtSPO11-2, and AtPRD1 as well as two new genes, AtPRD2 and AtPRD3. Our data thus increase the number of proteins necessary for DSB formation in Arabidopsis thaliana to five. Unlike SPO11 and (to a minor extent) PRD1, these two new proteins are poorly conserved among species, suggesting that the DSB formation mechanism, but not its regulation, is conserved among eukaryotes.

  7. ARABIDOPSIS THALIANA HOMEOBOX25 Uncovers a Role for Gibberellins in Seed Longevity1[C][W

    PubMed Central

    Bueso, Eduardo; Muñoz-Bertomeu, Jesús; Campos, Francisco; Brunaud, Veronique; Martínez, Liliam; Sayas, Enric; Ballester, Patricia; Yenush, Lynne; Serrano, Ramón

    2014-01-01

    Seed longevity is crucial for agriculture and plant genetic diversity, but it is limited by cellular damage during storage. Seeds are protected against aging by cellular defenses and by structures such as the seed coat. We have screened an activation-tagging mutant collection of Arabidopsis (Arabidopsis thaliana) and selected four dominant mutants with improved seed longevity (isl1-1D to isl4-1D) under both natural and accelerated aging conditions. In the isl1-1D mutant, characterized in this work, overexpression of the transcription factor ARABIDOPSIS THALIANA HOMEOBOX25 (ATHB25; At5g65410) increases the expression of GIBBERELLIC ACID3-OXIDASE2, encoding a gibberellin (GA) biosynthetic enzyme, and the levels of GA1 and GA4 are higher (3.2- and 1.4-fold, respectively) in the mutant than in the wild type. The morphological and seed longevity phenotypes of the athb25-1D mutant were recapitulated in transgenic plants with moderate (4- to 6-fold) overexpression of ATHB25. Simultaneous knockdown of ATHB25, ATHB22, and ATHB31 expression decreases seed longevity, as does loss of ATHB25 and ATHB22 function in a double mutant line. Seeds from wild-type plants treated with GA and from a quintuple DELLA mutant (with constitutive GA signaling) are more tolerant to aging, providing additional evidence for a role of GA in seed longevity. A correlation was observed in several genotypes between seed longevity and mucilage formation at the seed surface, suggesting that GA may act by reinforcing the seed coat. This mechanism was supported by the observation of a maternal effect in reciprocal crosses between the wild type and the athb25-1D mutant. PMID:24335333

  8. ARABIDOPSIS THALIANA HOMEOBOX25 uncovers a role for Gibberellins in seed longevity.

    PubMed

    Bueso, Eduardo; Muñoz-Bertomeu, Jesús; Campos, Francisco; Brunaud, Veronique; Martínez, Liliam; Sayas, Enric; Ballester, Patricia; Yenush, Lynne; Serrano, Ramón

    2014-02-01

    Seed longevity is crucial for agriculture and plant genetic diversity, but it is limited by cellular damage during storage. Seeds are protected against aging by cellular defenses and by structures such as the seed coat. We have screened an activation-tagging mutant collection of Arabidopsis (Arabidopsis thaliana) and selected four dominant mutants with improved seed longevity (isl1-1D to isl4-1D) under both natural and accelerated aging conditions. In the isl1-1D mutant, characterized in this work, overexpression of the transcription factor ARABIDOPSIS THALIANA HOMEOBOX25 (ATHB25; At5g65410) increases the expression of GIBBERELLIC ACID3-OXIDASE2, encoding a gibberellin (GA) biosynthetic enzyme, and the levels of GA1 and GA4 are higher (3.2- and 1.4-fold, respectively) in the mutant than in the wild type. The morphological and seed longevity phenotypes of the athb25-1D mutant were recapitulated in transgenic plants with moderate (4- to 6-fold) overexpression of ATHB25. Simultaneous knockdown of ATHB25, ATHB22, and ATHB31 expression decreases seed longevity, as does loss of ATHB25 and ATHB22 function in a double mutant line. Seeds from wild-type plants treated with GA and from a quintuple DELLA mutant (with constitutive GA signaling) are more tolerant to aging, providing additional evidence for a role of GA in seed longevity. A correlation was observed in several genotypes between seed longevity and mucilage formation at the seed surface, suggesting that GA may act by reinforcing the seed coat. This mechanism was supported by the observation of a maternal effect in reciprocal crosses between the wild type and the athb25-1D mutant.

  9. Gravity-regulated gene expression in Arabidopsis thaliana

    NASA Astrophysics Data System (ADS)

    Sederoff, Heike; Brown, Christopher S.; Heber, Steffen; Kajla, Jyoti D.; Kumar, Sandeep; Lomax, Terri L.; Wheeler, Benjamin; Yalamanchili, Roopa

    Plant growth and development is regulated by changes in environmental signals. Plants sense environmental changes and respond to them by modifying gene expression programs to ad-just cell growth, differentiation, and metabolism. Functional expression of genes comprises many different processes including transcription, translation, post-transcriptional and post-translational modifications, as well as the degradation of RNA and proteins. Recently, it was discovered that small RNAs (sRNA, 18-24 nucleotides long), which are heritable and systemic, are key elements in regulating gene expression in response to biotic and abiotic changes. Sev-eral different classes of sRNAs have been identified that are part of a non-cell autonomous and phloem-mobile network of regulators affecting transcript stability, translational kinetics, and DNA methylation patterns responsible for heritable transcriptional silencing (epigenetics). Our research has focused on gene expression changes in response to gravistimulation of Arabidopsis roots. Using high-throughput technologies including microarrays and 454 sequencing, we iden-tified rapid changes in transcript abundance of genes as well as differential expression of small RNA in Arabidopsis root apices after minutes of reorientation. Some of the differentially regu-lated transcripts are encoded by genes that are important for the bending response. Functional mutants of those genes respond faster to reorientation than the respective wild type plants, indicating that these proteins are repressors of differential cell elongation. We compared the gravity responsive sRNAs to the changes in transcript abundances of their putative targets and identified several potential miRNA: target pairs. Currently, we are using mutant and transgenic Arabidopsis plants to characterize the function of those miRNAs and their putative targets in gravitropic and phototropic responses in Arabidopsis.

  10. SHUGOSHINs and PATRONUS protect meiotic centromere cohesion in Arabidopsis thaliana.

    PubMed

    Zamariola, Linda; De Storme, Nico; Vannerum, Katrijn; Vandepoele, Klaas; Armstrong, Susan J; Franklin, F Christopher H; Geelen, Danny

    2014-03-01

    In meiosis, chromosome cohesion is maintained by the cohesin complex, which is released in a two-step manner. At meiosis I, the meiosis-specific cohesin subunit Rec8 is cleaved by the protease Separase along chromosome arms, allowing homologous chromosome segregation. Next, in meiosis II, cleavage of the remaining centromere cohesin results in separation of the sister chromatids. In eukaryotes, protection of centromeric cohesion in meiosis I is mediated by SHUGOSHINs (SGOs). The Arabidopsis genome contains two SGO homologs. Here we demonstrate that Atsgo1 mutants show a premature loss of cohesion of sister chromatid centromeres at anaphase I and that AtSGO2 partially rescues this loss of cohesion. In addition to SGOs, we characterize PATRONUS which is specifically required for the maintenance of cohesion of sister chromatid centromeres in meiosis II. In contrast to the Atsgo1 Atsgo2 double mutant, patronus T-DNA insertion mutants only display loss of sister chromatid cohesion after meiosis I, and additionally show disorganized spindles, resulting in defects in chromosome segregation in meiosis. This leads to reduced fertility and aneuploid offspring. Furthermore, we detect aneuploidy in sporophytic tissue, indicating a role for PATRONUS in chromosome segregation in somatic cells. Thus, ploidy stability is preserved in Arabidopsis by PATRONUS during both meiosis and mitosis.

  11. Gravity perception and gravitropic response of inflorescence stems in Arabidopsis thaliana

    NASA Astrophysics Data System (ADS)

    Fukaki, H.; Tasaka, M.

    1999-01-01

    Shoots of higher plants exhibit negative gravitropism. However, little is known about the site of gravity perception in shoots and the molecular mechanisms of shoot gravitropic responses. Our recent analysis using shoot gravitropism1(sgr1)/scarecrow(scr) and sgr7/short-root (shr) mutants in Arabidopsis thaliana indicated that the endodermis is essential for shoot gravitropism and strongly suggested that the endodermis functions as the gravity-sensing cell layer in dicotyledonous plant shoots. In this paper, we present our recent analysis and model of gravity perception and gravitropic response of inflorescence stems in Arabidopsis thaliana.

  12. Identification and disruption of an Arabidopsis zinc finger gene controlling seed germination

    PubMed Central

    Papi, Maura; Sabatini, Sabrina; Bouchez, David; Camilleri, Christine; Costantino, Paolo; Vittorioso, Paola

    2000-01-01

    We describe here the Arabidopsis gene DAG1, encoding a zinc finger transcription factor of the Dof family, and show that it is involved in the control of seed germination. By a reverse genetics approach, we isolated an Arabidopsis mutant line with one T-DNA insertion in DAG1. Seeds from homozygous knockout dag1-1 plants do not develop dormancy and germinate also in the absence of light. Segregation analysis indicates that the effect of the mutation is maternal. Accordingly, in situ mRNA hybridizations reveal expression of DAG1 in the vascular tissue of the flower and maturing fruit but not in the seed. PMID:10640273

  13. Auxin polar transport in arabidopsis under simulated microgravity conditions - relevance to growth and development

    NASA Astrophysics Data System (ADS)

    Miyamoto, K.; Oka, M.; Yamamoto, R.; Masuda, Y.; Hoson, T.; Kamisaka, S.; Ueda, J.

    1999-01-01

    Activity of auxin polar transport in inflorescence axes of Arabidopsis thaliana grown under simulated microgravity conditions was studied in relation to the growth and development. Seeds were germinated and allowed to grow on an agar medium in test tubes on a horizontal clinostat. Horizontal clinostat rotation substantially reduced the growth of inflorescence axes and the productivity of seeds of Arabidopsis thaliana (ecotypes Landsberg erecta and Columbia), although it little affected seed germination, development of rosette leaves and flowering. The activity of auxin polar transport in inflorescence axes decreased when Arabidopsis plants were grown on a horizontal clinostat from germination stage, being ca. 60% of 1 g control. On the other hand, the auxin polar transport in inflorescence axes of Arabidopsis grown in 1 g conditions was not affected when the segments were exposed to various gravistimuli, including 3-dimensional clinorotation, during transport experiments. Pin-formed mutant of Arabidopsis, having a unique structure of the inflorescence axis with no flower and extremely low levels of the activity of auxin polar transport in inflorescence axes and endogenous auxin, did not continue its vegetative growth under clinostat rotation. These facts suggest that the development of the system of auxin polar transport in Arabidopsis is affected by microgravity, resulting in the inhibition of growth and development, especially during reproductive growth.

  14. Molecular cloning of cryptochrome 1 from apple and its functional characterization in Arabidopsis.

    PubMed

    Li, Yuan-Yuan; Mao, Ke; Zhao, Cheng; Zhang, Rui-Fen; Zhao, Xian-Yan; Zhang, Hua-Lei; Shu, Huai-Rui; Zhao, Yu-Jin

    2013-06-01

    Cryptochromes are blue-light photoreceptors involved in regulating many aspects of plant growth and development. Investigations of cryptochromes in plants have largely focused on Arabidopsis (Arabidopsis thaliana), tomato (Solanum lycopersicum), rice (Oryza sativa) and pea (Pisum sativum). Here, we isolated the cryptochrome 1 gene from apple (Malus domestica) (MdCRY1) and analyzed its function in transgenic Arabidopsis. The predicted MdCRY1 protein was most closely homologous to strawberry CRY1. In terms of transcript levels, MdCRY1 expression was up-regulated by light. The function of MdCRY1 was analyzed through heterologous expression in Arabidopsis. Overexpression of MdCRY1 in Arabidopsis is able to rescue the cry1 mutant phenotype, inhibit hypocotyl elongation, promote root growth, and enhance anthocyanin accumulation in wild-type seedlings under blue light. These data provide functional evidence for a role of MdCRY1 in controlling photomorphogenesis under blue light and indicate that CRY1 function is conserved between Arabidopsis and apple. Furthermore, we found that MdCRY1 interacts with AtCOP1 in both yeast and onion cells. This interaction may represent an important regulatory mechanism in blue-light signaling pathway in apple.

  15. Using Arabidopsis to Study Shoot Branching in Biomass Willow1[C][W][OA

    PubMed Central

    Ward, Sally P.; Salmon, Jemma; Hanley, Steven J.; Karp, Angela; Leyser, Ottoline

    2013-01-01

    The success of the short-rotation coppice system in biomass willow (Salix spp.) relies on the activity of the shoot-producing meristems found on the coppice stool. However, the regulation of the activity of these meristems is poorly understood. In contrast, our knowledge of the mechanisms behind axillary meristem regulation in Arabidopsis (Arabidopsis thaliana) has grown rapidly in the past few years through the exploitation of integrated physiological, genetic, and molecular assays. Here, we demonstrate that these assays can be directly transferred to study the control of bud activation in biomass willow and to assess similarities with the known hormone regulatory system in Arabidopsis. Bud hormone response was found to be qualitatively remarkably similar in Salix spp. and Arabidopsis. These similarities led us to test whether Arabidopsis hormone mutants could be used to assess allelic variation in the cognate Salix spp. hormone genes. Allelic differences in Salix spp. strigolactone genes were observed using this approach. These results demonstrate that both knowledge and assays from Arabidopsis axillary meristem biology can be successfully applied to Salix spp. and can increase our understanding of a fundamental aspect of short-rotation coppice biomass production, allowing more targeted breeding. PMID:23610219

  16. Inducible Expression of Arabidopsis Response Regulator 22 (ARR22), a Type-C ARR, in Transgenic Arabidopsis Enhances Drought and Freezing Tolerance

    PubMed Central

    Kim, Jungmook

    2013-01-01

    The Arabidopsis two-component signaling system, which is comprised of sensor histidine kinases, histidine phosphotransfer proteins, and response regulators, mediates cytokinin response as well as various other plant responses including abiotic stress responses. Arabidopsis response regulators (ARRs) are classified into type-A, -B, and -C. Although the roles of type-A and -B ARRs are well established in Arabidopsis plant signaling, roles of type-C ARRs, ARR22 and ARR24, remain elusive. ARR22, a preferentially cytosolic protein, interacts with certain Arabidopsis histidine phosphotransfer proteins (AHPs) and displays phosphatase activity on AHP5. ARR22 is induced by cold and dehydration. Here, we show that inducible overexpression of ARR22 in Arabidopsis enhanced dehydration, drought, and cold tolerance in a dexamethasone-dependent manner, whereas mutation of the putative phospho-accepting Asp to Asn in ARR22 (ARR22D74N) abolished these tolerance phenotypes. Overexpression of ARR22 decreased electrolyte leakage in dehydration-, drought-, or cold-stressed transgenic Arabidopsis plants compared with that of ARR22D74N or compared with wild-type plants. Transpiration rates and stomatal apertures were not affected by ARR22 overexpression. No significant difference in both dehydration and freezing tolerance was observed between wild-type and arr22 mutants with or without cytokinin preincubation, consistent with the lack of phenotypes of arr22 mutants in their vegetative development. Meta-profile analyses of the microarray data on ARR22-responsive genes indicate that ARR22 modulates expression of a variety of abiotic stress-responsive genes, which might contribute to increasing drought and freezing tolerance. Taken together, these results suggest that ARR22 plays a positive role in the stress tolerance response in part via enhancing cell membrane integrity and that phospho-histidine phosphatase activity of ARR22 may be required for this function. PMID:24244460

  17. Aspartate oxidase plays an important role in Arabidopsis stomatal immunity.

    PubMed

    Macho, Alberto P; Boutrot, Freddy; Rathjen, John P; Zipfel, Cyril

    2012-08-01

    Perception of pathogen-associated molecular patterns (PAMPs), such as bacterial flagellin (or the peptide flg22), by surface-localized receptors activates defense responses and subsequent immunity. In a previous forward-genetic screen aimed at the identification of Arabidopsis (Arabidopsis thaliana) flagellin-insensitive (fin) mutants, we isolated fin4, which is severely affected in flg22-triggered reactive oxygen species (ROS) bursts. Here, we report that FIN4 encodes the chloroplastic enzyme ASPARTATE OXIDASE (AO), which catalyzes the first irreversible step in the de novo biosynthesis of NAD. Genetic studies on the role of NAD have been hindered so far by the lethality of null mutants in NAD biosynthetic enzymes. Using newly identified knockdown fin alleles, we found that AO is required for the ROS burst mediated by the NADPH oxidase RBOHD triggered by the perception of several unrelated PAMPs. AO is also required for RBOHD-dependent stomatal closure. However, full AO activity is not required for flg22-induced responses that are RBOHD independent. Interestingly, although the fin4 mutation dramatically affects RBOHD function, it does not affect functions carried out by other members of the RBOH family, such as RBOHC and RBOHF. Finally, we determined that AO is required for stomatal immunity against the bacterium Pseudomonas syringae. Altogether, our work reveals a novel specific requirement for AO activity in PAMP-triggered RBOHD-dependent ROS burst and stomatal immunity. In addition, the availability of viable mutants for the chloroplastic enzyme AO will enable future detailed studies on the role of NAD metabolism in different cellular processes, including immunity, in Arabidopsis.

  18. Starch synthesis in Arabidopsis. Granule synthesis, composition, and structure.

    PubMed

    Zeeman, Samuel C; Tiessen, Axel; Pilling, Emma; Kato, K Lisa; Donald, Athene M; Smith, Alison M

    2002-06-01

    The aim of this work was to characterize starch synthesis, composition, and granule structure in Arabidopsis leaves. First, the potential role of starch-degrading enzymes during starch accumulation was investigated. To discover whether simultaneous synthesis and degradation of starch occurred during net accumulation, starch was labeled by supplying (14)CO(2) to intact, photosynthesizing plants. Release of this label from starch was monitored during a chase period in air, using different light intensities to vary the net rate of starch synthesis. No release of label was detected unless there was net degradation of starch during the chase. Similar experiments were performed on a mutant line (dbe1) that accumulates the soluble polysaccharide, phytoglycogen. Label was not released from phytoglycogen during the chase indicating that, even when in a soluble form, glucan is not appreciably degraded during accumulation. Second, the effect on starch composition of growth conditions and mutations causing starch accumulation was studied. An increase in starch content correlated with an increased amylose content of the starch and with an increase in the ratio of granule-bound starch synthase to soluble starch synthase activity. Third, the structural organization and morphology of Arabidopsis starch granules was studied. The starch granules were birefringent, indicating a radial organization of the polymers, and x-ray scatter analyses revealed that granules contained alternating crystalline and amorphous lamellae with a periodicity of 9 nm. Granules from the wild type and the high-starch mutant sex1 were flattened and discoid, whereas those of the high-starch mutant sex4 were larger and more rounded. These larger granules contained "growth rings" with a periodicity of 200 to 300 nm. We conclude that leaf starch is synthesized without appreciable turnover and comprises similar polymers and contains similar levels of molecular organization to storage starches, making Arabidopsis

  19. Arabidopsis GLASSY HAIR genes promote trichome papillae development

    PubMed Central

    Kirik, Viktor

    2013-01-01

    Specialized plant cells form cell walls with distinct composition and properties pertinent to their function. Leaf trichomes in Arabidopsis form thick cell walls that support the upright growth of these large cells and, curiously, have strong light-reflective properties. To understand the process of trichome cell-wall maturation and the molecular origins of this optical property, mutants affected in trichome light reflection were isolated and characterized. It was found that GLASSY HAIR (GLH) genes are required for the formation of surface papillae structures at late stages of trichome development. Trichomes in these mutants appeared transparent due to unobstructed light transmission. Genetic analysis of the isolated mutants revealed seven different gene loci. Two—TRICHOME BIREFRINGENCE (TBR) and NOK (Noeck)—have been reported previously to have the glassy trichome mutant phenotype. The other five glh mutants were analysed for cell-wall-related phenotypes. A significant reduction was found in cellulose content in glh2 and glh4 mutant trichomes. In addition to the glassy trichome phenotype, the glh6 mutants showed defects in leaf cuticular wax, and glh6 was found to represent a new allele of the eceriferum 10 (cer10) mutation. Trichomes of the glh1 and glh3 mutants did not show any other phenotypes beside reduced papillae formation. These data suggest that the GLH1 and GLH3 genes may have specific functions in trichome papillae formation, whereas GLH2, GLH4, and GLH6 genes are also involved in deposition of other cell-wall components. PMID:24014871

  20. The Receptor Kinase IMPAIRED OOMYCETE SUSCEPTIBILITY1 Attenuates Abscisic Acid Responses in Arabidopsis1[C][W

    PubMed Central

    Hok, Sophie; Allasia, Valérie; Andrio, Emilie; Naessens, Elodie; Ribes, Elsa; Panabières, Franck; Attard, Agnès; Ris, Nicolas; Clément, Mathilde; Barlet, Xavier; Marco, Yves; Grill, Erwin; Eichmann, Ruth; Weis, Corina; Hückelhoven, Ralph; Ammon, Alexandra; Ludwig-Müller, Jutta; Voll, Lars M.; Keller, Harald

    2014-01-01

    In plants, membrane-bound receptor kinases are essential for developmental processes, immune responses to pathogens and the establishment of symbiosis. We previously identified the Arabidopsis (Arabidopsis thaliana) receptor kinase IMPAIRED OOMYCETE SUSCEPTIBILITY1 (IOS1) as required for successful infection with the downy mildew pathogen Hyaloperonospora arabidopsidis. We report here that IOS1 is also required for full susceptibility of Arabidopsis to unrelated (hemi)biotrophic filamentous oomycete and fungal pathogens. Impaired susceptibility in the absence of IOS1 appeared to be independent of plant defense mechanism. Instead, we found that ios1-1 plants were hypersensitive to the plant hormone abscisic acid (ABA), displaying enhanced ABA-mediated inhibition of seed germination, root elongation, and stomatal opening. These findings suggest that IOS1 negatively regulates ABA signaling in Arabidopsis. The expression of ABA-sensitive COLD REGULATED and RESISTANCE TO DESICCATION genes was diminished in Arabidopsis during infection. This effect on ABA signaling was alleviated in the ios1-1 mutant background. Accordingly, ABA-insensitive and ABA-hypersensitive mutants were more susceptible and resistant to oomycete infection, respectively, showing that the intensity of ABA signaling affects the outcome of downy mildew disease. Taken together, our findings suggest that filamentous (hemi)biotrophs attenuate ABA signaling in Arabidopsis during the infection process and that IOS1 participates in this pathogen-mediated reprogramming of the host. PMID:25274985

  1. A Chemical Genetic Screening Procedure for Arabidopsis thaliana Seedlings

    PubMed Central

    Bjornson, Marta; Song, Xingshun; Dandekar, Abhaya; Franz, Annaliese; Drakakaki, Georgia; Dehesh, Katayoon

    2016-01-01

    Unbiased screening approaches are powerful tools enabling identification of novel players in biological processes. Chemical genetic screening refers to the technique of using a reporter response, such as expression of luciferase driven by a promoter of interest, to discover small molecules that affect a given process when applied to plants. These chemicals then act as tools for identification of regulatory components that could not otherwise be detected by forward genetic screens due to gene family redundancy or mutant lethality. This protocol describes a chemical genetic screen using Arabidopsis thaliana seedlings, which has led to recognition of novel players in the plant general stress response. PMID:27446980

  2. Proteomics and transcriptomics analyses of Arabidopsis floral buds uncover important functions of ARABIDOPSIS SKP1-LIKE1

    DOE PAGES

    Lu, Dihong; Ni, Weimin; Stanley, Bruce A.; ...

    2016-03-03

    The ARABIDOPSIS SKP1-LIKE1 (ASK1) protein functions as a subunit of SKP1-CUL1-F-box (SCF) E3 ubiquitin ligases. Previous genetic studies showed that ASK1 plays important roles in Arabidopsis flower development and male meiosis. However, the molecular impact of ASK1-containing SCF E3 ubiquitin ligases (ASK1-E3s) on the floral proteome and transcriptome is unknown. Here we identified proteins that are potentially regulated by ASK1-E3s by comparing floral bud proteomes of wild-type and the ask1 mutant plants. More than 200 proteins were detected in the ask1 mutant but not in wild-type and >300 were detected at higher levels in the ask1 mutant than in wild-type,more » but their RNA levels were not significantly different between wild-type and ask1 floral buds as shown by transcriptomics analysis, suggesting that they are likely regulated at the protein level by ASK1-E3s. Integrated analyses of floral proteomics and transcriptomics of ask1 and wild-type uncovered several potential aspects of ASK1-E3 functions, including regulation of transcription regulators, kinases, peptidases, and ribosomal proteins, with implications on possible mechanisms of ASK1-E3 functions in floral development. In conclusion, our results suggested that ASK1-E3s play important roles in Arabidopsis protein degradation during flower development. This study opens up new possibilities for further functional studies of these candidate E3 substrates.« less

  3. Disruption of the Arabidopsis Defense Regulator Genes SAG101, EDS1, and PAD4 Confers Enhanced Freezing Tolerance

    PubMed Central

    Chen, Qin-Fang; Xu, Le; Tan, Wei-Juan; Chen, Liang; Qi, Hua; Xie, Li-Juan; Chen, Mo-Xian; Liu, Bin-Yi; Yu, Lu-Jun; Yao, Nan; Zhang, Jian-Hua; Shu, Wensheng; Xiao, Shi

    2017-01-01

    In Arabidopsis, three lipase-like regulators, SAG101, EDS1, and PAD4, act downstream of resistance protein-associated defense signaling. Although the roles of SAG101, EDS1, and PAD4 in biotic stress have been extensively studied, little is known about their functions in plant responses to abiotic stresses. Here, we show that SAG101, EDS1, and PAD4 are involved in the regulation of freezing tolerance in Arabidopsis. With or without cold acclimation, the sag101, eds1, and pad4 single mutants, as well as their double mutants exhibited similarly enhanced tolerance to freezing temperatures. Upon cold exposure, the sag101, eds1, and pad4 mutants showed increased transcript levels of C-REPEAT/DRE BINDING FACTORs and their regulons, compared with wild type. Moreover, freezing-induced cell death and accumulation of hydrogen peroxide were ameliorated in sag101, eds1, and pad4 mutants. The sag101, eds1, and pad4 mutants had much lower salicylic acid (SA) and diacylglycerol (DAG) contents than wild type and exogenous application of SA and DAG compromised the freezing tolerance of the mutants. Furthermore, SA suppressed the cold-induced expression of DGATs and DGKs in wild-type leaves. These findings indicate that SAG101, EDS1, and PAD4 are involved in freezing response in Arabidopsis, at least in part, by modulating the homeostasis of SA and DAG. PMID:26149542

  4. Jasmonate is essential for insect defense in Arabidopsis.

    PubMed

    McConn, M; Creelman, R A; Bell, E; Mullet, J E; Browse, J

    1997-05-13

    The signaling pathways that allow plants to mount defenses against chewing insects are known to be complex. To investigate the role of jasmonate in wound signaling in Arabidopsis and to test whether parallel or redundant pathways exist for insect defense, we have studied a mutant (fad3-2 fad7-2 fad8) that is deficient in the jasmonate precursor linolenic acid. Mutant plants contained negligible levels of jasmonate and showed extremely high mortality ( approximately 80%) from attack by larvae of a common saprophagous fungal gnat, Bradysia impatiens (Diptera: Sciaridae), even though neighboring wild-type plants were largely unaffected. Application of exogenous methyl jasmonate substantially protected the mutant plants and reduced mortality to approximately 12%. These experiments precisely define the role of jasmonate as being essential for the induction of biologically effective defense in this plant-insect interaction. The transcripts of three wound-responsive genes were shown not to be induced by wounding of mutant plants but the same transcripts could be induced by application of methyl jasmonate. By contrast, measurements of transcript levels for a gene encoding glutathione S-transferase demonstrated that wound induction of this gene is independent of jasmonate synthesis. These results indicate that the mutant will be a good genetic model for testing the practical effectiveness of candidate defense genes.

  5. The ASK1 gene regulates development and interacts with the UFO gene to control floral organ identity in Arabidopsis.

    PubMed

    Zhao, D; Yang, M; Solava, J; Ma, H

    1999-09-01

    Normal flower development likely requires both specific and general regulators. We have isolated an Arabidopsis mutant ask1-1 (for -Arabidopsis skp1-like1-1), which exhibits defects in both vegetative and reproductive development. In the ask1-1mutant, rosette leaf growth is reduced, resulting in smaller than normal rosette leaves, and internodes in the floral stem are shorter than normal. Examination of cell sizes in these organs indicates that cell expansion is normal in the mutant, but cell number is reduced. In the mutant, the numbers of petals and stamens are reduced, and many flowers have one or more petals with a reduced size. In addition, all mutant flowers have short stamen filaments. Furthermore, petal/stamen chimeric organs are found in many flowers. These results indicate that the ASK1 gene affects the size of vegetative and floral organs. The ask1 floral phenotype resembles somewhat that of the Arabidopsis ufo mutants in that both genes affect whorls 2 and 3. We therefore tested for possible interactions between ASK1 and UFO by analyzing the phenotypes of ufo-2 ask1-1 double mutant plants. In these plants, vegetative development is similar to that of the ask1-1 single mutant, whereas the floral defects are more severe than those in either single mutant. Interior to the first whorl, the double mutant flowers have more sepals or sepal-like organs than are found in ufo-2, and less petals than ask1-1. Our results suggest that ASK1 interacts with UFO to control floral organ identity in whorls 2 and 3. This is very intriguing because ASK1 is very similar in sequence to the yeast SKP1 protein and UFO contains an F-box, a motif known to interact with SKP1 in yeast. Although the precise mechanism of ASK1 and UFO action is unknown, our results support the hypothesis that these two proteins physically interact in vivo.

  6. Modification of AtGRDP1 gene expression affects silique and seed development in Arabidopsis thaliana.

    PubMed

    Rodríguez-Hernández, Aída Araceli; Muro-Medina, Carlos Vladimir; Ramírez-Alonso, Jocelin Itzel; Jiménez-Bremont, Juan Francisco

    2017-03-08

    Glycine Rich Proteins (GRPs) are induced at different developmental stages and in specific plant tissues. Recently, we described a novel Arabidopsis gene encoding a short glycine-rich domain protein (AtGRDP1). This gene is involved in abiotic stress responsiveness; the Atgrdp1-null mutant seeds were more sensitive to stress, while the opposite phenotype was achieved by AtGRDP1 overexpression. In this study, we analyzed the phenotype of the fruits produced by Arabidopsis Atgrdp1 mutants and 35S::AtGRDP1 overexpression lines. Our analyses revealed important changes in silique length, seed number, seed weight and morphology in the analyzed lines. In particular, Atgrdp1 mutant lines exhibited several defects including short siliques, a diminished number of seeds per silique, and reduction in seed size and weight as compared to Col-0. The overexpression of the AtGRDP1 gene also generated phenotypes with alterations in size of silique, number of seeds per silique, and size and weight of seed. In addition, the expression analysis of AtGRDP1 gene showed that it was expressed in floral and fruit organs, with the highest expression level in mature siliques. The alterations in the siliques and seeds traits in the Atgrdp1 mutant line, as well as the phenotypes observed in AtGRDP1 overexpression lines, suggest a role of the AtGRDP1 gene in the Arabidopsis fruit development.

  7. Conserved Arabidopsis ECHIDNA protein mediates trans-Golgi-network trafficking and cell elongation.

    PubMed

    Gendre, Delphine; Oh, Jaesung; Boutté, Yohann; Best, Jacob G; Samuels, Lacey; Nilsson, Robert; Uemura, Tomohiro; Marchant, Alan; Bennett, Malcolm J; Grebe, Markus; Bhalerao, Rishikesh P

    2011-05-10

    Multiple steps of plant growth and development rely on rapid cell elongation during which secretory and endocytic trafficking via the trans-Golgi network (TGN) plays a central role. Here, we identify the ECHIDNA (ECH) protein from Arabidopsis thaliana as a TGN-localized component crucial for TGN function. ECH partially complements loss of budding yeast TVP23 function and a Populus ECH complements the Arabidopsis ech mutant, suggesting functional conservation of the genes. Compared with wild-type, the Arabidopsis ech mutant exhibits severely perturbed cell elongation as well as defects in TGN structure and function, manifested by the reduced association between Golgi bodies and TGN as well as mislocalization of several TGN-localized proteins including vacuolar H(+)-ATPase subunit a1 (VHA-a1). Strikingly, ech is defective in secretory trafficking, whereas endocytosis appears unaffected in the mutant. Some aspects of the ech mutant phenotype can be phenocopied by treatment with a specific inhibitor of vacuolar H(+)-ATPases, concanamycin A, indicating that mislocalization of VHA-a1 may account for part of the defects in ech. Hence, ECH is an evolutionarily conserved component of the TGN with a central role in TGN structure and function.

  8. Identification of An Arsenic Tolerant Double Mutant With a Thiol-Mediated Component And Increased Arsenic Tolerance in PhyA Mutants

    SciTech Connect

    Sung, D.Y.; Lee, D.; Harris, H.; Raab, A.; Feldmann, J.; Meharg, A.; Kumabe, B.; Komives, E.A.; Schroeder, J.I.; /SLAC, SSRL /Sydney U. /Aberdeen U. /UC, San Diego

    2007-04-06

    A genetic screen was performed to isolate mutants showing increased arsenic tolerance using an Arabidopsis thaliana population of activation tagged lines. The most arsenic-resistant mutant shows increased arsenate and arsenite tolerance. Genetic analyses of the mutant indicate that the mutant contains two loci that contribute to arsenic tolerance, designated ars4 and ars5. The ars4ars5 double mutant contains a single T-DNA insertion, ars4, which co-segregates with arsenic tolerance and is inserted in the Phytochrome A (PHYA) gene, strongly reducing the expression of PHYA. When grown under far-red light conditions ars4ars5 shows the same elongated hypocotyl phenotype as the previously described strong phyA-211 allele. Three independent phyA alleles, ars4, phyA-211 and a new T-DNA insertion allele (phyA-t) show increased tolerance to arsenate, although to a lesser degree than the ars4ars5 double mutant. Analyses of the ars5 single mutant show that ars5 exhibits stronger arsenic tolerance than ars4, and that ars5 is not linked to ars4. Arsenic tolerance assays with phyB-9 and phot1/phot2 mutants show that these photoreceptor mutants do not exhibit phyA-like arsenic tolerance. Fluorescence HPLC analyses show that elevated levels of phytochelatins were not detected in ars4, ars5 or ars4ars5, however increases in the thiols cysteine, gamma-glutamylcysteine and glutathione were observed. Compared with wild type, the total thiol levels in ars4, ars5 and ars4ars5 mutants were increased up to 80% with combined buthionine sulfoximine and arsenic treatments, suggesting the enhancement of mechanisms that mediate thiol synthesis in the mutants. The presented findings show that PHYA negatively regulates a pathway conferring arsenic tolerance, and that an enhanced thiol synthesis mechanism contributes to the arsenic tolerance of ars4ars5.

  9. Data for a comparative proteomic analysis of chloroplast biogenesis (clb) mutants.

    PubMed

    de Luna-Valdez, L A; Martínez-Batallar, A G; Hernández-Ortiz, M; Encarnación-Guevara, S; Ramos-Vega, M; López-Bucio, J S; León, P; Guevara-García, A A

    2014-12-01

    This data article contains data related to the research article titled Proteomic analysis of chloroplast biogenesis (clb) mutants uncovers novel proteins potentially involved in the development of Arabidopsis thaliana chloroplasts (de Luna-Valdez et al., 2014) [1]. This research article describes the 2-D PAGE-based proteomic analysis of wild-type and four mutant lines (cla1-1, clb2, clb5 and clb19) affected in the development of Arabidopsis thaliana chloroplasts. The report concludes with the discovery of three proteins potentially involved in chloroplast biogenesis. The information presented here represent the tables and figures that detail the processing of the raw data obtained from the image analysis of the 2-D PAGE gels.

  10. Data for a comparative proteomic analysis of chloroplast biogenesis (clb) mutants

    PubMed Central

    de Luna-Valdez, L.A.; Martínez-Batallar, A.G.; Hernández-Ortiz, M.; Encarnación-Guevara, S.; Ramos-Vega, M.; López-Bucio, J.S.; León, P.; Guevara-García, A.A.

    2014-01-01

    This data article contains data related to the research article titled Proteomic analysis of chloroplast biogenesis (clb) mutants uncovers novel proteins potentially involved in the development ofArabidopsis thalianachloroplasts (de Luna-Valdez et al., 2014) [1]. This research article describes the 2-D PAGE-based proteomic analysis of wild-type and four mutant lines (cla1-1, clb2, clb5 and clb19) affected in the development of Arabidopsis thaliana chloroplasts. The report concludes with the discovery of three proteins potentially involved in chloroplast biogenesis. The information presented here represent the tables and figures that detail the processing of the raw data obtained from the image analysis of the 2-D PAGE gels. PMID:26217679

  11. Each of the chloroplast potassium efflux antiporters affects photosynthesis and growth of fully developed Arabidopsis rosettes under short-day photoperiod.

    PubMed

    Dana, Somnath; Herdean, Andrei; Lundin, Björn; Spetea, Cornelia

    2016-12-01

    In Arabidopsis thaliana, the chloroplast harbors three potassium efflux antiporters (KEAs), namely KEA1 and KEA2 in the inner envelope and KEA3 in the thylakoid membrane. They may play redundant physiological roles as in our previous analyses of young developing Arabidopsis rosettes under long-day photoperiod (16 h light per day), chloroplast kea single mutants resembled the wild-type plants, whereas kea1kea2 and kea1kea2kea3 mutants were impaired in chloroplast development and photosynthesis resulting in stunted growth. Here, we aimed to study whether chloroplast KEAs play redundant roles in chloroplast function of older Arabidopsis plants with fully developed rosettes grown under short-day photoperiod (8 h light per day). Under these conditions, we found defects in photosynthesis and growth in the chloroplast kea single mutants, and most dramatic defects in the kea1kea2 double mutant. The mechanism behind these defects in the single mutants involves reduction in the electron transport rate (kea1 and kea3), and stomata conductance (kea1, kea2 and kea3), which in turn affect CO2 fixation rates. The kea1kea2 mutant, in addition to these alterations, displayed reduced levels of photosynthetic machinery. Taken together, our data suggest that, in addition to the previously reported roles in chloroplast development in young rosettes, each chloroplast KEA affects photosynthesis and growth of Arabidopsis fully developed rosettes.

  12. Redox regulation of ascorbate and glutathione by a chloroplastic dehydroascorbate reductase is required for high-light stress tolerance in Arabidopsis.

    PubMed

    Noshi, Masahiro; Hatanaka, Risa; Tanabe, Noriaki; Terai, Yusuke; Maruta, Takanori; Shigeoka, Shigeru

    2016-05-01

    Chloroplasts are a significant site for reactive oxygen species production under illumination and, thus, possess a well-organized antioxidant system involving ascorbate. Ascorbate recycling occurs in different manners in this system, including a dehydroascorbate reductase (DHAR) reaction. We herein investigated the physiological significance of DHAR3 in photo-oxidative stress tolerance in Arabidopsis. GFP-fused DHAR3 protein was targeted to chloroplasts in Arabidopsis leaves. A DHAR3 knockout mutant exhibited sensitivity to high light (HL). Under HL, the ascorbate redox states were similar in mutant and wild-type plants, while total ascorbate content was significantly lower in the mutant, suggesting that DHAR3 contributes, at least to some extent, to ascorbate recycling. Activation of monodehydroascorbate reductase occurred in dhar3 mutant, which might compensate for the lack of DHAR3. Interestingly, glutathione oxidation was consistently inhibited in dhar3 mutant. These findings indicate that DHAR3 regulates both ascorbate and glutathione redox states to acclimate to HL.

  13. Amyloplast movement and gravityperception in Arabidopsis endoderm

    NASA Astrophysics Data System (ADS)

    Tasaka, M.; Saito, T.; Morita, M. T.

    Gravitropism of higher plant is a growth response regulating the orientation of organs elongation, which includes four sequential steps, the perception of gravistimulus, transduction of the physical stimulus to chemical signal, transmission of the signal, and differential cell elongation depending on the signal. To elucidate the molecular mechanism of these steps, we have isolated a number of Arabidopsis mutants with abnormal shoot gravitropic response. zig (zigzag)/sgr4(shoot gravitropism 4) shows little gravitropism in their shoots. Besides, their inflorescence stems elongate in a zigzag-fashion to bend at each node. ZIG encodes a SNARE, AtVTI11. sgr3 with reduced gravitropic response in inflorescence stems had a missense mutation in other SNARE, AtVAM3. These two SNAREs make a complex in the shoot endoderm cells that are gravity-sensing cells, suggesting that the vesicle transport from trans-Golgi network (TGN) to prevacuolar compartment (PVC) and/or vacuole is involved in gravitropism. Abnormal vesicular/vacuolar structures were observed in several tissues of both mutants. Moreover, SGR2 encodes phospholipase A1-like protein that resides in the vacuolar membrane. Endodermis-specific expression of these genes could complement gravitropism in each mutant. In addition, amyloplasts thought to be statoliths localized abnormally in their endoderm cells. These results strongly suggest that formation and function of vacuole in the endoderm cells are important for amyloplasts sedimentation, which is involved in the early process of shoot gravitropism. To reveal this, we constructed vertical stage microscope system to visualize the behavior of amyloplasts and vacuolar membrane in living endodermal cells. We hope to discuss the mechanism of gravity perception after showing their movements.

  14. Arabidopsis STERILE APETALA, a multifunctional gene regulating inflorescence, flower, and ovule development

    PubMed Central

    Byzova, Marina V.; Franken, John; Aarts, Mark G.M.; de Almeida-Engler, Janice; Engler, Gilbert; Mariani, Celestina; Van Lookeren Campagne, Michiel M.; Angenent, Gerco C.

    1999-01-01

    A recessive mutation in the Arabidopsis STERILE APETALA (SAP) causes severe aberrations in inflorescence and flower and ovule development. In sap flowers, sepals are carpelloid, petals are short and narrow or absent, and anthers are degenerated. Megasporogenesis, the process of meiotic divisions preceding the female gametophyte formation, is arrested in sap ovules during or just after the first meiotic division. More severe aberrations were observed in double mutants between sap and mutant alleles of the floral homeotic gene APETALA2 (AP2) suggesting that both genes are involved in the initiation of female gametophyte development. Together with the organ identity gene AGAMOUS (AG) SAP is required for the maintenance of floral identity acting in a manner similar to APETALA1. In contrast to the outer two floral organs in sap mutant flowers, normal sepals and petals develop in ag/sap double mutants, indicating that SAP negatively regulates AG expression in the perianth whorls. This supposed cadastral function of SAP is supported by in situ hybridization experiments showing ectopic expression of AG in the sap mutant. We have cloned the SAP gene by transposon tagging and revealed that it encodes a novel protein with sequence motifs, that are also present in plant and animal transcription regulators. Consistent with the mutant phenotype, SAP is expressed in inflorescence and floral meristems, floral organ primordia, and ovules. Taken together, we propose that SAP belongs to a new class of transcription regulators essential for a number of processes in Arabidopsis flower development. PMID:10215627

  15. Suppression of Dwarf and irregular xylem Phenotypes Generates Low-Acetylated Biomass Lines in Arabidopsis.

    PubMed

    Bensussan, Matthieu; Lefebvre, Valérie; Ducamp, Aloïse; Trouverie, Jacques; Gineau, Emilie; Fortabat, Marie-Noëlle; Guillebaux, Alexia; Baldy, Aurélie; Naquin, Delphine; Herbette, Stéphane; Lapierre, Catherine; Mouille, Gregory; Horlow, Christine; Durand-Tardif, Mylène

    2015-06-01

    eskimo1-5 (esk1-5) is a dwarf Arabidopsis (Arabidopsis thaliana) mutant that has a constitutive drought syndrome and collapsed xylem vessels, along with low acetylation levels in xylan and mannan. ESK1 has xylan O-acetyltransferase activity in vitro. We used a suppressor strategy on esk1-5 to screen for variants with wild-type growth and low acetylation levels, a favorable combination for ethanol production. We found a recessive mutation in the KAKTUS (KAK) gene that suppressed dwarfism and the collapsed xylem character, the cause of decreased hydraulic conductivity in the esk1-5 mutant. Backcrosses between esk1-5 and two independent knockout kak mutants confirmed suppression of the esk1-5 effect. kak single mutants showed larger stem diameters than the wild type. The KAK promoter fused with a reporter gene showed activity in the vascular cambium, phloem, and primary xylem in the stem and hypocotyl. However, suppression of the collapsed xylem phenotype in esk1 kak double mutants was not associated with the recovery of cell wall O-acetylation or any major cell wall modifications. Therefore, our results indicate that, in addition to its described activity as a repressor of endoreduplication, KAK may play a role in vascular development. Furthermore, orthologous esk1 kak double mutants may hold promise for ethanol production in crop plants.

  16. Defense against Sclerotinia sclerotiorum in Arabidopsis is dependent on jasmonic acid, salicylic acid, and ethylene signaling.

    PubMed

    Guo, Xiaomei; Stotz, Henrik U

    2007-11-01

    Genotypic differences in susceptibility of Arabidopsis thaliana to Sclerotinia sclerotiorum have not been reported due to the extreme susceptibility of this cruciferous plant. To overcome this limitation, we have established inoculation conditions that enable evaluation of differences in susceptibility to S. sclerotiorum among Arabidopsis mutants and ecotypes. Two coil mutant alleles conferred hypersusceptibility to S. sclerotiorum. The plant defensin gene PDF1.2 was no longer induced after challenging the coi1-2 mutant with S. sclerotiorum. Hypersusceptibility of the coi1-2 mutant to S. sclerotiorum was not correlated with oxalate sensitivity. The mutants npr1 and ein2 were also hypersusceptible to S. sclerotiorum. Induction of PDF1.2 and the pathogenesis-related gene PR1 was reduced in ein2 and npr1 mutants, respectively. Actigard, a commercial formulation of the systemic acquired resistance inducer benzothiadiazole, reduced susceptibility to S. sclerotiorum. Based on histochemical analysis of oxalate-deficient and wild-type strains of S. sclerotiorum, oxalate caused a decrease in hydrogen peroxide production but no detectable changes in plant superoxide production or gene expression.

  17. Light-Stimulated Cotyledon Expansion in Arabidopsis Seedlings (The Role of Phytochrome B).

    PubMed Central

    Neff, M. M.; Van Volkenburgh, E.

    1994-01-01

    Leaf and cotyledon expansion in dicotyledonous plants is a light-dependent developmental process. The unique role of phytochrome B has been tested by investigating expansion of cotyledons in wild-type and phytochrome-deficient mutants of Arabidopsis thaliana (L.) Heynh. A relatively rapid method for measuring cotyledon area was developed to quantify growth in large populations (average n [greater than or equal to] 100) of wild-type or mutant seedlings under different light and chemical treatments. Three-day-old wild-type (La-er) Arabidopsis seedlings, grown in saturating, low-fluence red light (2-4 [mu]mol m-2 s-1), showed a >250% increase in cotyledon area after 48 h of bright-red light when compared with the phytochrome mutants hy1, hy2, and hy3. An increase in epidermal cell area was observed in wild-type cotyledons but not in hy3, indicating that light-stimulated growth is due in part to cell expansion. The mutant phenotype was rescued by feeding the chromophore precursor biliverdin to the chromophore biosynthesis mutants hy1 and hy6. This treatment did not rescue the hy3 mutant. Since the hy3 lesion is specific to phytochrome B, we conclude that this pigment is involved in the enhancement of cotyledon cell expansion in bright-red light. PMID:12232145

  18. 'Gr