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Sample records for arabidopsis cyp51a2 mutant

  1. Arabidopsis mutants impaired in cosuppression.

    PubMed Central

    Elmayan, T; Balzergue, S; Béon, F; Bourdon, V; Daubremet, J; Guénet, Y; Mourrain, P; Palauqui, J C; Vernhettes, S; Vialle, T; Wostrikoff, K; Vaucheret, H

    1998-01-01

    Post-transcriptional gene silencing (cosuppression) results in the degradation of RNA after transcription. A transgenic Arabidopsis line showing post-transcriptional silencing of a 35S-uidA transgene and uidA-specific methylation was mutagenized using ethyl methanesulfonate. Six independent plants were isolated in which uidA mRNA accumulation and beta-glucuronidase activity were increased up to 3500-fold, whereas the transcription rate of the 35S-uidA transgene was increased only up to threefold. These plants each carried a recessive monogenic mutation that is responsible for the release of silencing. These mutations defined two genetic loci, called sgs1 and sgs2 (for suppressor of gene silencing). Transgene methylation was distinctly modified in sgs1 and sgs2 mutants. However, methylation of centromeric repeats was not affected, indicating that sgs mutants differ from ddm (for decrease in DNA methylation) and som (for somniferous) mutants. Indeed, unlike ddm and som mutations, sgs mutations were not able to release transcriptional silencing of a 35S-hpt transgene. Conversely, both sgs1 and sgs2 mutations were able to release cosuppression of host Nia genes and 35S-Nia2 transgenes. These results therefore indicate that sgs mutations act in trans to impede specifically transgene-induced post-transcriptional gene silencing. PMID:9761800

  2. Arabidopsis mutants with a reduced seed dormancy.

    PubMed Central

    Léon-Kloosterziel, K M; van de Bunt, G A; Zeevaart, J A; Koornneef, M

    1996-01-01

    The development of seed dormancy is an aspect of seed maturation, the last stage of seed development. To isolate mutants of Arabidopsis thaliana that are affected in this process, we selected directly for the absence of dormancy among freshly harvested M2 seeds. The screen yielded two mutants exhibiting a reduced dormancy, rdo1 and rdo2, that are specifically affected in dormancy determined by the embryo. The rdo1 and rdo2 mutants show normal levels of abscisic acid and the same sensitivity to abscisic acid, ethylene, auxin, and cytokinin as the wild type. The rdo2 mutant but not the rdo1 mutant has a reduced sensitivity to the gibberellin biosynthesis inhibitor tetcyclacis. Double-mutant analysis suggested that the RDO1 and RDO2 genes are involved in separate pathways leading to the development of dormancy. We assume that the RDO2 gene controls a step in the induction of dormancy that is most likely induced by abscisic acid and is expressed as an increase of the gibberellin requirement for germination. PMID:8587986

  3. 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.

  4. 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 ...

  5. 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

  6. An Arabidopsis mutant defective in the general phenylpropanoid pathway.

    PubMed Central

    Chapple, C C; Vogt, T; Ellis, B E; Somerville, C R

    1992-01-01

    Mutants of Arabidopsis deficient in a major leaf phenylpropanoid ester, 2-O-sinapoyl-L-malate, were identified by thin-layer chromatographic screening of methanolic leaf extracts from several thousand mutagenized plants. Mutations at a locus designated SIN1 also eliminate accumulation of the sinapic acid esters characteristic of seed tissues. Because of increased transparency to UV light, the sin1 mutants exhibit a characteristic red fluorescence under UV light, whereas wild-type plants have a blue-green appearance due to the fluorescence of sinapoyl malate in the upper epidermis. As determined by in vivo radiotracer feeding experiments, precursor supplementation studies, and enzymatic assays, the defect in the sin1 mutants appears to block the conversion of ferulate to 5-hydroxyferulate in the general phenylpropanoid pathway. As a result, the lignin of the mutant lacks the sinapic acid-derived components typical of wild-type lignin. PMID:1477555

  7. 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.

  8. An Arabidopsis mutant with enhanced resistance to powdery mildew.

    PubMed Central

    Frye, C A; Innes, R W

    1998-01-01

    We have identified an Arabidopsis mutant that displays enhanced disease resistance to the fungus Erysiphe cichoracearum, causal agent of powdery mildew. The edr1 mutant does not constitutively express the pathogenesis-related genes PR-1, BGL2, or PR-5 and thus differs from previously described disease-resistant mutants of Arabidopsis. E. cichoracearum conidia (asexual spores) germinated normally and formed extensive hyphae on edr1 plants, indicating that the initial stages of infection were not inhibited. Production of conidiophores on edr1 plants, however, was <16% of that observed on wild-type Arabidopsis. Reduction in sporulation correlated with a more rapid induction of defense responses. Autofluorescent compounds and callose accumulated in edr1 leaves 3 days after inoculation with E. cichoracearum, and dead mesophyll cells accumulated in edr1 leaves starting 5 days after inoculation. Macroscopic patches of dead cells appeared 6 days after inoculation. This resistance phenotype is similar to that conferred by "late-acting" powdery mildew resistance genes of wheat and barley. The edr1 mutation is recessive and maps to chromosome 1 between molecular markers ATEAT1 and NCC1. We speculate that the edr1 mutation derepresses multiple defense responses, making them more easily induced by virulent pathogens. PMID:9634583

  9. Characterization of a new Arabidopsis mutant exhibiting enhanced disease resistance.

    PubMed

    Silva, H; Yoshioka, K; Dooner, H K; Klessig, D F

    1999-12-01

    In many plant-pathogen interactions, resistance is associated with the synthesis and accumulation of salicylic acid (SA) and pathogenesis-related (PR) proteins. At least two general classes of mutants with altered resistance to pathogen attack have been identified in Arabidopsis. One class exhibits increased susceptibility to pathogen infection; the other class exhibits enhanced resistance to pathogens. In an attempt to identify mutations in resistance-associated loci, we screened a population of T-DNA tagged Arabidopsis thaliana ecotype Wassilewskija (Ws) for mutants showing constitutive expression of the PR-1 gene (cep). A mutant was isolated and shown to constitutively express PR-1, PR-2, and PR-5 genes. This constitutive phenotype segregated as a single recessive trait in the Ws genetic background. The mutant also had elevated levels of SA, which are responsible for the cep phenotype. The cep mutant spontaneously formed hypersensitive response (HR)-like lesions on the leaves and cotyledons and also exhibited enhanced resistance to virulent bacterial and fungal pathogens. Genetic analyses of segregating progeny from outcrosses to other ecotypes unexpectedly revealed that alterations in more than one gene condition the constitutive expression of PR genes in the original mutant. One of the mutations, designated cpr20, maps to the lower arm of chromosome 4 and is required for the cep phenotype. Another mutation, which has been termed cpr21, maps to chromosome 1 and is often, but not always, associated with this phenotype. The recessive nature of the cep trait suggests that the CPR20 and CPR21 proteins may act as negative regulators in the disease resistance signal transduction pathway. PMID:10624014

  10. Arabidopsis thaliana nucleosidase mutants provide new insights into nucleoside degradation

    PubMed Central

    Riegler, Heike; Geserick, Claudia; Zrenner, Rita

    2011-01-01

    A central step in nucleoside and nucleobase salvage pathways is the hydrolysis of nucleosides to their respective nucleobases. In plants this is solely accomplished by nucleosidases (EC 3.2.2.x). To elucidate the importance of nucleosidases for nucleoside degradation, general metabolism, and plant growth, thorough phenotypic and biochemical analyses were performed using Arabidopsis thaliana T-DNA insertion mutants lacking expression of the previously identified genes annotated as uridine ribohydrolases (URH1 and URH2). Comprehensive functional analyses of single and double mutants demonstrated that both isoforms are unimportant for seedling establishment and plant growth, while one participates in uridine degradation. Rather unexpectedly, nucleoside and nucleotide profiling and nucleosidase activity screening of soluble crude extracts revealed a deficiency of xanthosine and inosine hydrolysis in the single mutants, with substantial accumulation of xanthosine in one of them. Mixing of the two mutant extracts, and by in vitro activity reconstitution using a mixture of recombinant URH1 and URH2 proteins, both restored activity, thus providing biochemical evidence that at least these two isoforms are needed for inosine and xanthosine hydrolysis. This mutant study demonstrates the utility of in vivo systems for the examination of metabolic activities, with the discovery of the new substrate xanthosine and elucidation of a mechanism for expanding the nucleosidase substrate spectrum. PMID:21599668

  11. 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

  12. Isolation of Arabidopsis mutants with altered seed fatty acid composition

    SciTech Connect

    Lemieux, B.; Browse, J.; Somerville, C. Washington State Univ., Pullman )

    1989-04-01

    By direct screening of Arabidopsis seed fatty acid methyl esters, we have isolated mutants which are deficient in the elongation of 18:1 to 20:1 and the desaturation of 18:2 to 18:3. Both the elongation and the desaturation mutants, designated MB14 and BL1 respectively, have only 10% of the wild-type levels of 20:1 and 18:3 in their seeds. The intermediate levels of 20:1 and 18:3 in F1 seeds of crosses to the wild type indicate that the level of enzyme is regulating the amount of 20:1 and 18:3 in seeds. Consistent with this observation, the mutations were found to segregate 1:2:1 in F2 seeds. We have found that the 18:2 desaturase mutation is clearly expressed in root phosphatidylcholine.

  13. 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.

  14. Flavonoid Accumulation Patterns of Transparent Testa Mutants of Arabidopsis1

    PubMed Central

    Peer, Wendy Ann; Brown, Dana E.; Tague, Brian W.; Muday, Gloria K.; Taiz, Lincoln; Murphy, Angus 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. PMID:11402185

  15. Genetic analysis of salt-tolerant mutants in Arabidopsis thaliana.

    PubMed Central

    Quesada, V; Ponce, M R; Micol, J L

    2000-01-01

    Stress caused by the increased salinity of irrigated fields impairs plant growth and is one of the major constraints that limits crop productivity in many important agricultural areas. As a contribution to solving such agronomic problems, we have carried out a large-scale screening for Arabidopsis thaliana mutants induced on different genetic backgrounds by EMS treatment, fast neutron bombardment, or T-DNA insertions. From the 675,500 seeds we screened, 17 mutant lines were isolated, all but one of which yielded 25-70% germination levels on 250 mm NaCl medium, a condition in which their ancestor ecotypes are unable to germinate. Monogenic recessive inheritance of NaCl-tolerant germination was displayed with incomplete penetrance by all the selected mutants, which fell into five complementation groups. These were named SALOBRENO (SAN) and mapped relative to polymorphic microsatellites, the map positions of three of them suggesting that they are novel genes. Strains carrying mutations in the SAN1-SAN4 genes display similar responses to both ionic effects and osmotic pressure, their germination being NaCl and mannitol tolerant but KCl and Na(2)SO(4) sensitive. In addition, NaCl-, KCl-, and mannitol-tolerant as well as abscisic-acid-insensitive germination was displayed by sañ5, whose genetic and molecular characterization indicates that it carries an extremely hypomorphic or null allele of the ABI4 gene, its deduced protein product lacking the APETALA2 DNA binding domain. PMID:10629000

  16. Arabidopsis AMY1 expressions and early flowering mutant phenotype.

    PubMed

    Jie, Wang; Dashi, Yu; XinHong, Guo; Xuanming, Liu

    2009-02-28

    The homozygous T-DNA mutant of the AMY1 gene in Arabidopsis was identified and importantly, shown to cause an early flowering phenotype. We found that the disruption of AMY1 enhanced expression of CO and FT. The expression analyses of genes related to starch metabolism revealed that expression of the AGPase small subunit APS1 in the wild type was higher than in the amy1 mutant. However, there were no significant differences in expression levels of the AGPase large subunit genes ApL1, AMY2, or AMY3 between wild type and the amy1 mutant. Expression profiling showed that AMY1 was highly expressed in leaves, stems, and flowers, and expressed less in leafstalks and roots. Furthermore, the level of AMY1 mRNA was highly elevated with age and in senescing leaves. RT-PCR analyses showed that the expression of AMY1 was induced by heat shock, GA, and ABA, while salt stress had no apparent effect on its expression. PMID:19250611

  17. 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

  18. Arabidopsis Flavonoid Mutants Are Hypersensitive to UV-B Irradiation.

    PubMed Central

    Li, J; Ou-Lee, TM; Raba, R; Amundson, RG; Last, RL

    1993-01-01

    Increases in the terrestrial levels of ultraviolet-B (UV-B) radiation (280 to 320 nm) due to diminished stratospheric ozone have prompted an investigation of the protective mechanisms that contribute to UV-B tolerance in plants. In response to UV-B stress, flowering plants produce a variety of UV-absorptive secondary products derived from phenylalanine. Arabidopsis mutants with defects in the synthesis of these compounds were tested for UV-B sensitivity. The transparent testa-4 (tt4) mutant, which has reduced flavonoids and normal levels of sinapate esters, is more sensitive to UV-B than the wild type when grown under high UV-B irradiance. The tt5 and tt6 mutants, which have reduced levels of UV-absorptive leaf flavonoids and the monocyclic sinapic acid ester phenolic compounds, are highly sensitive to the damaging effects of UV-B radiation. These results demonstrate that both flavonoids and other phenolic compounds play important roles in vivo in plant UV-B protection. PMID:12271060

  19. Molecular analysis of ethylene-insensitive mutants in arabidopsis

    SciTech Connect

    Meyerowitz, E.

    1991-01-01

    The subject of this study is the biochemical basis of ethylene reception. The Arabidopsis thaliana ETR gene codes for the ethylene receptor or is involved in transduction of the ethylene-generated signal. We have cloned an etr mutation which results in a decrease in the ethylene response of the plant, with a decrease in ethylene binding of about five-fold. Two genes have been found in the cloned region which confer resistance. By sequence analysis, the first protein contains three distinct regions: a transmembrane region, a serine/threonine protein kinase region, and a control region similar to the RAS-binding region of yeast adenylate cyclase. The second protein contains a zinc-finger; since sequence of the first protein shows no mutant-dependent changes, and transition metals have been implicated in ethylene binding, this protein could be the ETR gene product. However, no mutant dependent differences have been found in this protein, either. The mutation could be upstream of the coding region of either gene and involve regulatory elements, so we are continuing to sequence. (MHB)

  20. Leaf Epicuticular Waxes of the Eceriferum Mutants in Arabidopsis.

    PubMed Central

    Jenks, M. A.; Tuttle, H. A.; Eigenbrode, S. D.; Feldmann, K. A.

    1995-01-01

    Wild-type Arabidopsis leaf epicuticular wax (EW) occurs as a smooth layer over the epidermal surface, whereas stem EW has a crystalline microstructure. Wild-type EW load was more than 10-fold lower on leaves than on stems. Compared with the EW on wild-type stems, EW on wild-type leaves had a much higher proportion of their total EW load in the form of alkanes and 1-alcohols; a large reduction in secondary alcohols, ketones, and esters; and a chain-length distribution for major EW classes that was skewed toward longer lengths. The eceriferum (cer) mutations often differentially affected leaf and stem EW chemical compositions. For example, the cer2 mutant EW phenotype was expressed on the stem but not on the leaf. Compared to wild type, the amount of primary alcohols on cer9 mutants was reduced on leaves but elevated on stems, whereas an opposite differential effect for primary alcohols was observed on cer16 leaves and stems. Putative functions for CER gene products are discussed. The CER4 and CER6 gene products may be involved in fatty aldehyde reduction and C26 fatty acylcoenzyme A elongation, respectively. CER1, CER8, CER9, and CER16 gene products may be involved in EW substrate transfer. The CER3 gene product may be involved in release of fatty acids from elongase complexes. CER2 gene product may have regulatory functions. PMID:12228482

  1. 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.

  2. Characterization of grape Gibberellin Insensitive 1 mutant alleles in transgenic Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We generated a dozen of different mutations in the grape Gibberellin Insensitive or GAI sequence, transformed them into Arabidopsis under the control of 35S, Arabidopsis or grape GAI promoter, and evaluated the impact of these mutant alleles on plant growth and development. These GAI sequence varian...

  3. Metabolomic Characterization of Knockout Mutants in Arabidopsis: Development of a Metabolite Profiling Database for Knockout Mutants in Arabidopsis1[W][OPEN

    PubMed Central

    Fukushima, Atsushi; Kusano, Miyako; Mejia, Ramon Francisco; Iwasa, Mami; Kobayashi, Makoto; Hayashi, Naomi; Watanabe-Takahashi, Akiko; Narisawa, Tomoko; Tohge, Takayuki; Hur, Manhoi; Wurtele, Eve Syrkin; Nikolau, Basil J.; Saito, Kazuki

    2014-01-01

    Despite recent intensive research efforts in functional genomics, the functions of only a limited number of Arabidopsis (Arabidopsis thaliana) genes have been determined experimentally, and improving gene annotation remains a major challenge in plant science. As metabolite profiling can characterize the metabolomic phenotype of a genetic perturbation in the plant metabolism, it provides clues to the function(s) of genes of interest. We chose 50 Arabidopsis mutants, including a set of characterized and uncharacterized mutants, that resemble wild-type plants. We performed metabolite profiling of the plants using gas chromatography-mass spectrometry. To make the data set available as an efficient public functional genomics tool for hypothesis generation, we developed the Metabolite Profiling Database for Knock-Out Mutants in Arabidopsis (MeKO). It allows the evaluation of whether a mutation affects metabolism during normal plant growth and contains images of mutants, data on differences in metabolite accumulation, and interactive analysis tools. Nonprocessed data, including chromatograms, mass spectra, and experimental metadata, follow the guidelines set by the Metabolomics Standards Initiative and are freely downloadable. Proof-of-concept analysis suggests that MeKO is highly useful for the generation of hypotheses for genes of interest and for improving gene annotation. MeKO is publicly available at http://prime.psc.riken.jp/meko/. PMID:24828308

  4. 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.

  5. Arabidopsis genes essential for seedling viability: isolation of insertional mutants and molecular cloning.

    PubMed Central

    Budziszewski, G J; Lewis, S P; Glover, L W; Reineke, J; Jones, G; Ziemnik, L S; Lonowski, J; Nyfeler, B; Aux, G; Zhou, Q; McElver, J; Patton, D A; Martienssen, R; Grossniklaus, U; Ma, H; Law, M; Levin, J Z

    2001-01-01

    We have undertaken a large-scale genetic screen to identify genes with a seedling-lethal mutant phenotype. From screening approximately 38,000 insertional mutant lines, we identified >500 seedling-lethal mutants, completed cosegregation analysis of the insertion and the lethal phenotype for >200 mutants, molecularly characterized 54 mutants, and provided a detailed description for 22 of them. Most of the seedling-lethal mutants seem to affect chloroplast function because they display altered pigmentation and affect genes encoding proteins predicted to have chloroplast localization. Although a high level of functional redundancy in Arabidopsis might be expected because 65% of genes are members of gene families, we found that 41% of the essential genes found in this study are members of Arabidopsis gene families. In addition, we isolated several interesting classes of mutants and genes. We found three mutants in the recently discovered nonmevalonate isoprenoid biosynthetic pathway and mutants disrupting genes similar to Tic40 and tatC, which are likely to be involved in chloroplast protein translocation. Finally, we directly compared T-DNA and Ac/Ds transposon mutagenesis methods in Arabidopsis on a genome scale. In each population, we found only about one-third of the insertion mutations cosegregated with a mutant phenotype. PMID:11779813

  6. 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

  7. Detecting Functional Groups of Arabidopsis Mutants by Metabolic Profiling and Evaluation of Pleiotropic Responses

    PubMed Central

    Hofmann, Jörg; Börnke, Frederik; Schmiedl, Alfred; Kleine, Tatjana; Sonnewald, Uwe

    2011-01-01

    Metabolic profiles and fingerprints of Arabidopsis thaliana plants with various defects in plastidic sugar metabolism or photosynthesis were analyzed to elucidate if the genetic mutations can be traced by comparing their metabolic status. Using a platform of chromatographic and spectrometric tools data from untargeted full MS scans as well as from selected metabolites including major carbohydrates, phosphorylated intermediates, carboxylates, free amino acids, major antioxidants, and plastidic pigments were evaluated. Our key observations are that by multivariate statistical analysis each mutant can be separated by a unique metabolic signature. Closely related mutants come close. Thus metabolic profiles of sugar mutants are different but more similar than those of photosynthesis mutants. All mutants show pleiotropic responses mirrored in their metabolic status. These pleiotropic responses are typical and can be used for separating and grouping of the mutants. Our findings show that metabolite fingerprints can be taken to classify mutants and hence may be used to sort genes into functional groups. PMID:22639613

  8. 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.

  9. 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

  10. Comparative proteomic study of Arabidopsis mutants mpk4 and mpk6.

    PubMed

    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

  11. 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

  12. 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.

  13. 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...

  14. Mutant and overexpression analysis of a C2H2 single zinc finger gene of Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The goal of this work was to characterize an Arabidopsis C2H2 single zinc finger gene, named AtZFP11, that is similar to SUPERMAN (SUP) and RABBIT EARS (RBE). No altered phenotype was observed in mutants analyzed that were derived through TILLING, nor a T-DNA insertion into the exon of AtZFP11. Pl...

  15. Azetidine-2-carboxylic acid resistant mutants of Arabidopsis thaliana with increased salt tolerance

    SciTech Connect

    Lehle, F.R.; Murphy, M.A.; Khan, R.A. )

    1989-04-01

    Nineteen mutant Arabidopsis families resistant to the proline analog azetidine-2-carboxylic acid (ACA) were characterized in terms of NaCl tolerance and proline content. Mutants were selected from about 64,000 progeny of about 16,000 self-pollinated Columbia parents which had been mutated with ethyl methane sulfonate during seed imbibition. Selections were performed during seed germination on aseptic agar medium containing 0.2 to 0.25 mM ACA. Nineteen mutant families, 12 clearly independent, retained resistance to ACA in the M{sub 4} generation. Based on germination on 150 mM NaCl, 13 of the mutant families were more tolerant than the wild type. Two mutants of intermediate resistance to ACA were markedly more salt tolerant than the others. Four mutant families appeared to overproduce proline. Of these, only 3 showed slight increases in salt tolerance.

  16. 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.

  17. The Arabidopsis elch mutant reveals functions of an ESCRT component in cytokinesis.

    PubMed

    Spitzer, Christoph; Schellmann, Swen; Sabovljevic, Aneta; Shahriari, Mojgan; Keshavaiah, Channa; Bechtold, Nicole; Herzog, Michel; Müller, Stefan; Hanisch, Franz-Georg; Hülskamp, Martin

    2006-12-01

    Recently, an alternative route to the proteasomal protein-degradation pathway was discovered that specifically targets transmembrane proteins marked with a single ubiquitin to the endosomal multivesicular body (MVB) and, subsequently, to the vacuole (yeast) or lysosome (animals), where they are degraded by proteases. Vps23p/TSG101 is a key component of the ESCRT I-III machinery in yeast and animals that recognizes mono-ubiquitylated proteins and sorts them into the MVB. Here, we report that the Arabidopsis ELCH (ELC) gene encodes a Vps23p/TSG101 homolog, and that homologs of all known ESCRT I-III components are present in the Arabidopsis genome. As with its animal and yeast counterparts, ELC binds ubiquitin and localizes to endosomes. Gel-filtration experiments indicate that ELC is a component of a high-molecular-weight complex. Yeast two-hybrid and immunoprecipitation assays showed that ELC interacts with Arabidopsis homologs of the ESCRT I complex. The elc mutant shows multiple nuclei in various cell types, indicating a role in cytokinesis. Double-mutant analysis with kaktus shows that increased ploidy levels do not influence the cytokinesis effect of elc mutants, suggesting that ELC is only important during the first endoreduplication cycle. Double mutants with tubulin folding cofactor a mutants show a synergistic phenotype, suggesting that ELC regulates cytokinesis through the microtubule cytoskeleton. PMID:17090720

  18. RARGE II: An Integrated Phenotype Database of Arabidopsis Mutant Traits Using a Controlled Vocabulary

    PubMed Central

    Akiyama, Kenji; Kurotani, Atsushi; Iida, Kei; Kuromori, Takashi; Shinozaki, Kazuo; Sakurai, Tetsuya

    2014-01-01

    Arabidopsis thaliana is one of the most popular experimental plants. However, only 40% of its genes have at least one experimental Gene Ontology (GO) annotation assigned. Systematic observation of mutant phenotypes is an important technique for elucidating gene functions. Indeed, several large-scale phenotypic analyses have been performed and have generated phenotypic data sets from many Arabidopsis mutant lines and overexpressing lines, which are freely available online. Since each Arabidopsis mutant line database uses individual phenotype expression, the differences in the structured term sets used by each database make it difficult to compare data sets and make it impossible to search across databases. Therefore, we obtained publicly available information for a total of 66,209 Arabidopsis mutant lines, including loss-of-function (RATM and TARAPPER) and gain-of-function (AtFOX and OsFOX) lines, and integrated the phenotype data by mapping the descriptions onto Plant Ontology (PO) and Phenotypic Quality Ontology (PATO) terms. This approach made it possible to manage the four different phenotype databases as one large data set. Here, we report a publicly accessible web-based database, the RIKEN Arabidopsis Genome Encyclopedia II (RARGE II; http://rarge-v2.psc.riken.jp/), in which all of the data described in this study are included. Using the database, we demonstrated consistency (in terms of protein function) with a previous study and identified the presumed function of an unknown gene. We provide examples of AT1G21600, which is a subunit in the plastid-encoded RNA polymerase complex, and AT5G56980, which is related to the jasmonic acid signaling pathway. PMID:24272250

  19. An Arabidopsis neutral ceramidase mutant ncer1 accumulates hydroxyceramides and is sensitive to oxidative stress

    PubMed Central

    Li, Jian; Bi, Fang-Cheng; Yin, Jian; Wu, Jian-Xin; Rong, Chan; Wu, Jia-Li; Yao, Nan

    2015-01-01

    Ceramidases hydrolyze ceramide into sphingosine and fatty acids and, although ceramidases function as key regulators of sphingolipid homeostasis in mammals, their roles in plants remain largely unknown. Here, we characterized the Arabidopsis thaliana ceramidase AtNCER1, a homolog of human neutral ceramidase. AtNCER1 localizes predominantly on the endoplasmic reticulum. The ncer1 T-DNA insertion mutants had no visible phenotype, but accumulated hydroxyceramides, and showed increased sensitivity to oxidative stress induced by methyl viologen. Plants over-expressing AtNCER1 showed increased tolerance to oxidative stress. These data indicate that the Arabidopsis neutral ceramidase affects sphingolipid homeostasis and oxidative stress responses. PMID:26150824

  20. 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.

  1. 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

  2. A mutant of Arabidopsis thaliana (L.) Heynh. with modified control of aspartate kinase by threonine.

    PubMed

    Heremans, B; Jacobs, M

    1997-04-01

    Mutagenesis and subsequent selection of Arabidopsis thaliana plantlets on a growth inhibitory concentration of lysine has led to the isolation of lysine-resistant mutants. The ability to grown on 2 mM lysine has been used to isolate mutants that may contain an aspartate kinase with altered regulatory-feedback properties. One of these mutants (RL 4) was characterized by a relative enhancement of soluble lysine. The recessive monogenic nuclear transmission of the resistance trait was established. It was associated with an aspartate kinase less sensitive to feedback inhibition by threonine. Two mutants (RLT 40 and RL 4) in Arabidopsis, characterized by an altered regulation of aspartate kinase, were crossed to assess the effects of the simultaneous presence of these different aspartate kinase forms. A double mutant (RLT40 x RL4) was isolated and characterized by two feedback-desensitized isozymes of aspartate kinase to, respectively, lysine and threonine but no threonine and/or lysine overproduction was observed. Genetical analysis of this unique double aspartate kinase mutant indicated that both mutations were located on chromosome 2, but their loci (ak1 and ak2) were found to be unlinked. PMID:9241437

  3. Genetic analysis of incurvata mutants reveals three independent genetic operations at work in Arabidopsis leaf morphogenesis.

    PubMed Central

    Serrano-Cartagena, J; Candela, H; Robles, P; Ponce, M R; Pérez-Pérez, J M; Piqueras, P; Micol, J L

    2000-01-01

    In an attempt to identify genes involved in the control of leaf morphogenesis, we have studied 13 Arabidopsis thaliana mutants with curled, involute leaves, a phenotype herein referred to as Incurvata (Icu), which were isolated by G. Röbbelen and belong to the Arabidopsis Information Service Form Mutants collection. The Icu phenotype was inherited as a single recessive trait in 10 mutants, with semidominance in 2 mutants and with complete dominance in the remaining 1. Complementation analyses indicated that the studied mutations correspond to five genes, representative alleles of which were mapped relative to polymorphic microsatellites. Although most double-mutant combinations displayed additivity of the Icu phenotypes, those of icu1 icu2 and icu3 icu4 double mutants were interpreted as synergistic, which suggests that the five genes studied represent three independent genetic operations that are at work for the leaf to acquire its final form at full expansion. We have shown that icu1 mutations are alleles of the Polycomb group gene CURLY LEAF (CLF) and that the leaf phenotype of the icu2 mutant is suppressed in an agamous background, as is known for clf mutants. In addition, we have tested by means of multiplex RT-PCR the transcription of several floral genes in Icu leaves. Ectopic expression of AGAMOUS and APETALA3 was observed in clf and icu2, but not in icu3, icu4, and icu5 mutants. Taken together, these results suggest that CLF and ICU2 play related roles, the latter being a candidate to belong to the Polycomb group of regulatory genes. We propose that, as flowers evolved, a new major class of genes, including CLF and ICU2, may have been recruited to prevent the expression of floral homeotic genes in the leaves. PMID:11063708

  4. 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 ...

  5. 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.

  6. 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.

  7. An Arabidopsis thaliana copper-sensitive mutant suggests a role of phytosulfokine in ethylene production

    PubMed Central

    Wu, Tao; Kamiya, Takehiro; Yumoto, Hiroko; Sotta, Naoyuki; Katsushi, Yamaguchi; Shigenobu, Shuji; Matsubayashi, Yoshikatsu; Fujiwara, Toru

    2015-01-01

    To increase our understanding of the adaptation for copper (Cu) deficiency, Arabidopsis mutants with apparent alterations under Cu deficiency were identified. In this report, a novel mutant, tpst-2, was found to be more sensitive than wild-type (Col-0) plants to Cu deficiency during root elongation. The positional cloning of tpst-2 revealed that this gene encodes a tyrosylprotein sulfotransferase (TPST). Moreover, the ethylene production of tpst-2 mutant was higher than that of Col-0 under Cu deficiency, and adding the ethylene response inhibitor AgNO3 partially rescued defects in root elongation. Interestingly, peptide hormone phytosulfokine (PSK) treatment also repressed the ethylene production of tpst-2 mutant plants. Our results revealed that TPST suppressed ethylene production through the action of PSK. PMID:25908239

  8. An Arabidopsis thaliana copper-sensitive mutant suggests a role of phytosulfokine in ethylene production.

    PubMed

    Wu, Tao; Kamiya, Takehiro; Yumoto, Hiroko; Sotta, Naoyuki; Katsushi, Yamaguchi; Shigenobu, Shuji; Matsubayashi, Yoshikatsu; Fujiwara, Toru

    2015-07-01

    To increase our understanding of the adaptation for copper (Cu) deficiency, Arabidopsis mutants with apparent alterations under Cu deficiency were identified. In this report, a novel mutant, tpst-2, was found to be more sensitive than wild-type (Col-0) plants to Cu deficiency during root elongation. The positional cloning of tpst-2 revealed that this gene encodes a tyrosylprotein sulfotransferase (TPST). Moreover, the ethylene production of tpst-2 mutant was higher than that of Col-0 under Cu deficiency, and adding the ethylene response inhibitor AgNO3 partially rescued defects in root elongation. Interestingly, peptide hormone phytosulfokine (PSK) treatment also repressed the ethylene production of tpst-2 mutant plants. Our results revealed that TPST suppressed ethylene production through the action of PSK. PMID:25908239

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

    PubMed

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

    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. PMID:11537704

  10. A Direct Screening Procedure for Gravitropism Mutants in Arabidopsis thaliana (L.) Heynh. 1

    PubMed Central

    Bullen, Bertha L.; Best, Thérèse R.; Gregg, Mary M.; Barsel, Sara-Ellen; Poff, Kenneth L.

    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. PMID:11537704

  11. Identification of Mutants of Arabidopsis Defective in Acclimation of Photosynthesis to the Light Environment1

    PubMed Central

    Walters, Robin G.; Shephard, Freya; Rogers, Jennifer J.M.; Rolfe, Stephen A.; Horton, Peter

    2003-01-01

    In common with many other higher plant species, Arabidopsis undergoes photosynthetic acclimation, altering the composition of the photosynthetic apparatus in response to fluctuations in its growth environment. The changes in photosynthetic function that result from acclimation can be detected in a noninvasive manner by monitoring chlorophyll (Chl) fluorescence. This technique has been used to develop a screen that enables the rapid identification of plants defective at ACCLIMATION OF PHOTOSYNTHESIS TO THE ENVIRONMENT (APE) loci. The application of this screen to a population of T-DNA-transformed Arabidopsis has successfully led to the identification of a number of mutant lines with altered Chl fluorescence characteristics. Analysis of photosynthesis and pigment composition in leaves from three such mutants showed that they had altered acclimation responses to the growth light environment, each having a distinct acclimation-defective phenotype, demonstrating that screening for mutants using Chl fluorescence is a viable strategy for the investigation of acclimation. Sequencing of the genomic DNA flanking the T-DNA elements showed that in the ape1 mutant, a gene was disrupted that encodes a protein of unknown function but that appears to be specific to photosynthetic organisms, whereas the ape2 mutant carries an insertion in the region of the TPT gene encoding the chloroplast inner envelope triose phosphate/phosphate translocator. PMID:12586872

  12. 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. PMID:24591708

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

    PubMed Central

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

    2014-01-01

    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. PMID:24591708

  14. 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.

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

    PubMed

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

    2016-07-01

    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

  16. Effect of light quality and vernalization on late-flowering mutants of Arabidopsis thaliana

    SciTech Connect

    Martinez-Zapater, J.M. ); Somerville, C.R. )

    1990-03-01

    We have analyzed the response to vernalization and light quality of six classes of late-flowering mutants (fb, fca, fe, fg, ft, and fy) previously isolated following mutagenesis of the early Landsberg race of Arabidopsis thaliana (L.) Heynh. When grown in continuous fluorescent illumination, four mutants (fca, fe, ft, and fy) and the Landsberg wild type exhibited a reduction in both flowering time and leaf number following 6 weeks of vernalization. A significant decrease in flowering time was also observed for all the mutants and the wild type when constant fluorescent illumination was supplemented with irradiation enriched in the red and far red regions of the spectrum. In the most extreme case, the late-flowering phenotype of the fca mutant was completely suppressed by vernalization, suggesting that this mutation has a direct effect on flowering. The fe and fy mutants also showed a more pronounced response than wild type to both vernalization and incandescent supplementation. The ft mutant showed a similar response to that of the wild type. The fb and fg mutants were substantially less sensitive to these treatments. These results are interpreted in the context of a multifactorial pathway for induction of flowering, in which the various mutations affect different steps of the pathway.

  17. 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.

  18. 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 ...

  19. 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...

  20. An Arabidopsis mutant impaired in coenzyme A biosynthesis is sugar dependent for seedling establishment.

    PubMed

    Rubio, Silvia; Larson, Tony R; Gonzalez-Guzman, Miguel; Alejandro, Santiago; Graham, Ian A; Serrano, Ramón; Rodriguez, Pedro L

    2006-03-01

    Once the plant coenzyme A (CoA) biosynthetic pathway has been elucidated by comparative genomics, it is feasible to analyze the physiological relevance of CoA biosynthesis in plant life. To this end, we have identified and characterized Arabidopsis (Arabidopsis thaliana) T-DNA knockout mutants of two CoA biosynthetic genes, HAL3A and HAL3B. The HAL3A gene encodes a 4'-phosphopantothenoyl-cysteine decarboxilase that generates 4'-phosphopantetheine. A second gene, HAL3B, whose gene product is 86% identical to that of HAL3A, is present in the Arabidopsis genome. HAL3A appears to have a predominant role over HAL3B according to their respective mRNA expression levels. The hal3a-1, hal3a-2, and hal3b mutants were viable and showed a similar growth rate as that in wild-type plants; in contrast, a hal3a-1 hal3b double mutant was embryo lethal. Unexpectedly, seedlings that were null for HAL3A and heterozygous for HAL3B (aaBb genotype) displayed a sucrose (Suc)-dependent phenotype for seedling establishment, which is in common with mutants defective in beta-oxidation. This phenotype was genetically complemented in aaBB siblings of the progeny and chemically complemented by pantethine. In contrast, seedling establishment of Aabb plants was not Suc dependent, proving a predominant role of HAL3A over HAL3B at this stage. Total fatty acid and acyl-CoA measurements of 5-d-old aaBb seedlings in medium lacking Suc revealed stalled storage lipid catabolism and impaired CoA biosynthesis; in particular, acetyl-CoA levels were reduced by approximately 80%. Taken together, these results provide in vivo evidence for the function of HAL3A and HAL3B, and they point out the critical role of CoA biosynthesis during early postgerminative growth. PMID:16415216

  1. 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

    2016-01-01

    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. PMID:27160427

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

    PubMed Central

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

    2016-01-01

    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. MPK6F364L and MPK6F368L 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. PMID:27160427

  3. 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.

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

    PubMed Central

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

    2013-01-01

    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. PMID:23299430

  5. 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. PMID:23299430

  6. [Disturbed thylakoid-formation in chloroplasts of a xantha-mutant of Arabidopsis thaliana (L.) HEYNH].

    PubMed

    Röbbeilen, G

    1966-03-01

    A monohybrid recessive xantha-mutant of Arabidopsis thaliana was grown aseptically on mineral agar +2% sucrose up to maturity. The cotyledons and rosette leaves were non-green during all developmental stages, their yellow colour bleaching with age.Palisade cells of various leaves of mutant plants were investigated light- and electron microscopically. All of the organelles of these cells, except the plastids, had a normal appearance. Special attention was paid to the existence of "dense bodies", and their relationship to sphaerosomes and lysosomes was outlined.In the mutant plastids the typical thylakoid differentiation present in the chloroplasts of the green Arabidopsis plant is entirely lacking. Within the palisade cells the plastids are normal in number and size. Under the light microscope they appear as hyaline vesicles with usually two "primary grana". Their outline is characterized by numerous invaginations and protrusions; however, it is still uncertain to what extent this "amoeboidy" is only an artifact.The specific gene dependent block in the plastid development is apparently effective during the conversion of tubules of the prolamellar body to thylakoids. From the persistent "prolamellar bodies" in the mutant only single abnormal thylakoids originate. These often have a deformed cup- or ventricle-like shape and occasionally are piled up as magnograna. In a characteristic manner tubules and fibrils are closely associated in parallel rows with the outsides of the thylakoids. The fibrils having a mean diameter of 75 Å are connected with the tubules which are derived from the "prolamellar body". From this striking aggregation the possibility of a laminal fusion of tubules as a process of normal thylakoid growth is deduced.By pigment analysis of mutant leaves no chlorophyll but only carotenoids were detected. However, many osmiophilic globules as well as large bodies of unknown composition (proteinaceous?) appear in the plastid matrix. Therefore the question

  7. Comparison of rhizosphere bacterial communities in Arabidopsis thaliana mutants for systemic acquired resistance.

    PubMed

    Hein, John W; Wolfe, Gordon V; Blee, Kristopher A

    2008-02-01

    Systemic acquired resistance (SAR) is an inducible systemic plant defense against a broad spectrum of plant pathogens, with the potential to secrete antimicrobial compounds into the soil. However, its impact on rhizosphere bacteria is not known. In this study, we examined fingerprints of bacterial communities in the rhizosphere of the model plant Arabidopsis thaliana to determine the effect of SAR on bacterial community structure and diversity. We compared Arabidopsis mutants that are constitutive and non-inducible for SAR and verified SAR activation by measuring pathogenesis-related protein activity via a beta-glucoronidase (GUS) reporter construct driven by the beta-1-3 glucanase promoter. We used terminal restriction fragment length polymorphism (T-RFLP) analysis of MspI- and HaeIII-digested 16S rDNA to estimate bacterial rhizosphere community diversity, with Lactobacillus sp. added as internal controls. T-RFLP analysis showed a clear rhizosphere effect on community structure, and diversity analysis of both rhizosphere and bulk soil operational taxonomic units (as defined by terminal restriction fragments) using richness, Shannon-Weiner, and Simpson's diversity indices and evenness confirmed that the presence of Arabidopsis roots significantly altered bacterial communities. This effect of altered soil microbial community structure by plants was also seen upon multivariate cluster analysis of the terminal restriction fragments. We also found visible differences in the rhizosphere community fingerprints of different Arabidopsis SAR mutants; however, there was no clear decrease of rhizosphere diversity because of constitutive SAR expression. Our study suggests that SAR can alter rhizosphere bacterial communities, opening the door to further understanding and application of inducible plant defense as a driving force in structuring soil bacterial assemblages. PMID:17619212

  8. The Arabidopsis her1 mutant implicates GABA in E-2-hexenal responsiveness.

    PubMed

    Mirabella, Rossana; Rauwerda, Han; Struys, Eduard A; Jakobs, Cornelis; Triantaphylidès, Christian; Haring, Michel A; Schuurink, Robert C

    2008-01-01

    When wounded or attacked by herbivores or pathogens, plants produce a blend of six-carbon alcohols, aldehydes and esters, known as C6-volatiles. Undamaged plants, when exposed to C6-volatiles, respond by inducing defense-related genes and secondary metabolites, suggesting that C6-volatiles can act as signaling molecules regulating plant defense responses. However, to date, the molecular mechanisms by which plants perceive and respond to these volatiles are unknown. To elucidate such mechanisms, we decided to isolate Arabidopsis thaliana mutants in which responses to C6-volatiles were altered. We observed that treatment of Arabidopsis seedlings with the C6-volatile E-2-hexenal inhibits root elongation. Among C6-volatiles this response is specific to E-2-hexenal, and is not dependent on ethylene, jasmonic and salicylic acid. Using this bioassay, we isolated 18 E-2-hexenal-response (her) mutants that showed sustained root growth after E-2-hexenal treatment. Here, we focused on the molecular characterization of one of these mutants, her1. Microarray and map-based cloning revealed that her1 encodes a gamma-amino butyric acid transaminase (GABA-TP), an enzyme that degrades GABA. As a consequence of the mutation, her1 plants accumulate high GABA levels in all their organs. Based on the observation that E-2-hexenal treatment induces GABA accumulation, and that high GABA levels confer resistance to E-2-hexenal, we propose a role for GABA in mediating E-2-hexenal responses. PMID:17971036

  9. 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.

  10. Chloroplast 2010: a database for large-scale phenotypic screening of Arabidopsis mutants.

    PubMed

    Lu, Yan; Savage, Linda J; Larson, Matthew D; Wilkerson, Curtis G; Last, Robert L

    2011-04-01

    Large-scale phenotypic screening presents challenges and opportunities not encountered in typical forward or reverse genetics projects. We describe a modular database and laboratory information management system that was implemented in support of the Chloroplast 2010 Project, an Arabidopsis (Arabidopsis thaliana) reverse genetics phenotypic screen of more than 5,000 mutants (http://bioinfo.bch.msu.edu/2010_LIMS; www.plastid.msu.edu). The software and laboratory work environment were designed to minimize operator error and detect systematic process errors. The database uses Ruby on Rails and Flash technologies to present complex quantitative and qualitative data and pedigree information in a flexible user interface. Examples are presented where the database was used to find opportunities for process changes that improved data quality. We also describe the use of the data-analysis tools to discover mutants defective in enzymes of leucine catabolism (heteromeric mitochondrial 3-methylcrotonyl-coenzyme A carboxylase [At1g03090 and At4g34030] and putative hydroxymethylglutaryl-coenzyme A lyase [At2g26800]) based upon a syndrome of pleiotropic seed amino acid phenotypes that resembles previously described isovaleryl coenzyme A dehydrogenase (At3g45300) mutants. In vitro assay results support the computational annotation of At2g26800 as hydroxymethylglutaryl-coenzyme A lyase. PMID:21224340

  11. Arabidopsis mutant of AtABCG26, an ABC transporter gene, is defective in pollen maturation.

    PubMed

    Kuromori, Takashi; Ito, Takuya; Sugimoto, Eriko; Shinozaki, Kazuo

    2011-11-01

    In plants, pollen is the male gametophyte that is generated from microspores, which are haploid cells produced after meiosis of diploid pollen mother cells in floral anthers. In normal maturation, microspores interact with the tapetum, which consists of one layer of metabolically active cells enclosing the locule in anthers. The tapetum plays several important roles in the maturation of microspores. ATP-binding cassette (ABC) transporters are a highly conserved protein super-family that uses the energy released in ATP hydrolysis to transport substrates. The ABC transporter gene family is more diverse in plants than in animals. Previously, we reported that an Arabidopsis half-size type ABC transporter gene, COF1/AtWBC11/AtABCG11, is involved in lipid transport for the construction of cuticle layers and pollen coats in normal organ formation, as compared to CER5/AtWBC12/AtABCG12. However, physiological functions of most other ABCG members are unknown. Here, we identified another family gene, AtABCG26, which is required for pollen development in Arabidopsis. An AtABCG26 mutant developed very few pollen grains, resulting in a male-sterile phenotype. By investigating microspore and pollen development in this mutant, we observed that there was a slight abnormality in tetrad morphology prior to the formation of haploid microspores. At a later stage, we could not detect exine deposition on the microspore surface. During pollen maturation, many grains in the mutant anthers got aborted, and surviving grains were found to be defective in mitosis. Transmission of the mutant allele through male gametophytes appeared to be normal in genetic transmission analysis, supporting the view that the pollen function was disturbed by sporophytic defects in the AtABCG26 mutant. AtABCG26 can be expected to be involved in the transport of substrates such as sporopollenin monomers from tapetum to microspores, which both are plant-specific structures critical to pollen development. PMID

  12. Indole-3-butyric acid synthesis in ecotypes and mutants of Arabidopsis thaliana under different growth conditions.

    PubMed

    Ludwig-Müller, Jutta

    2007-01-01

    Although IBA is a naturally occurring auxin, its role in plant development is still under debate. In this study a set of Arabidopsis mutants was used to analyze the biosynthesis of IBA in vitro. The mutants chosen for this study can be classified as: (1) involvement in auxin metabolism, transport or synthesis (amt1, aux1, ilr1, nit1, rib1, sur1, trp1-100); (2) other hormones possibly involved in the regulation of IBA synthesis (aba1, aba3, eto2, fae1, hls1, jar1); (3) photomorphogenesis (det1, det2, det3); and (4) root architecture (cob1, cob2, scr1). In addition, two transgenic lines overexpressing the IAA glucose synthase (iaglu) gene from maize were analyzed. The ecotypes No-0 and Wassilewskija showed the highest IBA synthetase activity under control conditions, followed by Columbia, Enkheim and Landsberg erecta. In the mutant lines IBA synthetase activity differed in most cases from the wild type, however no particular pattern of up- or down-regulation, which could be correlated to their possible function, was found. For rib1 mutant seedlings it was tested whether reduced IBA synthetase activity correlates with the endogenous IBA levels. Free IBA differed only depending on the culture conditions, but gave no clear correlation with IBA synthetase activity compared to the wild type. Since drought and osmotic stress as well as abscisic acid (ABA) application enhanced IBA synthesis in maize, it was tested whether IBA synthetase from Arabidopsis is also inducible by drought stress conditions. This was confirmed for the two ecotypes Col and Ler which showed different IBA synthetase activity when cultivated with various degrees of drought stress. IBA synthetase was also determined in photomorphogenic mutants under different light regimes. Induction of IBA synthetase in det1 and det3 plants was found under short day plus a red light pulse or in the dark, respectively. The results are discussed with respect to the functions of the mutated genes. PMID:16325963

  13. Mutants of Arabidopsis lacking a chloroplastic isoamylase accumulate phytoglycogen and an abnormal form of amylopectin.

    PubMed

    Wattebled, Fabrice; Dong, Ying; Dumez, Sylvain; Delvallé, David; Planchot, Véronique; Berbezy, Pierre; Vyas, Darshna; Colonna, Paul; Chatterjee, Manash; Ball, Steven; D'Hulst, Christophe

    2005-05-01

    Mutant lines defective for each of the four starch debranching enzyme (DBE) genes (AtISA1, AtISA2, AtISA3, and AtPU1) detected in the nuclear genome of Arabidopsis (Arabidopsis thaliana) were produced and analyzed. Our results indicate that both AtISA1 and AtISA2 are required for the production of a functional isoamylase-type of DBE named Iso1, the major isoamylase activity found in leaves. The absence of Iso1 leads to an 80% decrease in the starch content in both lines and to the accumulation of water-soluble polysaccharides whose structure is similar to glycogen. In addition, the residual amylopectin structure in the corresponding mutant lines displays a strong modification when compared to the wild type, suggesting a direct, rather than an indirect, function of Iso1 during the synthesis of amylopectin. Mutant lines carrying a defect in AtISA3 display a strong starch-excess phenotype at the end of both the light and the dark phases accompanied by a small modification of the amylopectin structure. This result suggests that this isoamylase-type of DBE plays a major role during starch mobilization. The analysis of the Atpu1 single-mutant lines did not lead to a distinctive phenotype. However, Atisa2/Atpu1 double-mutant lines display a 92% decrease in starch content. This suggests that the function of pullulanase partly overlaps that of Iso1, although its implication remains negligible when Iso1 is present within the cell. PMID:15849301

  14. Identification and Biochemical Characterization of Mutants in the Proanthocyanidin Pathway in Arabidopsis1

    PubMed Central

    Abrahams, Sharon; Tanner, Gregory J.; Larkin, Philip J.; Ashton, Anthony R.

    2002-01-01

    Proanthocyanidin (PA), or condensed tannin, is a polymeric flavanol that accumulates in a number of tissues in a wide variety of plants. In Arabidopsis, we found that PA precursors (detected histochemically using OsO4) accumulate in the endothelial cell layer of the seed coat from the two-terminal cell stage of embryo development onwards. To understand how PA is made, we screened mature seed pools of T-DNA-tagged Arabidopsis lines to identify mutants defective in the synthesis of PA and found six tds (tannin-deficient seed) complementation groups defective in PA synthesis. Mutations in these loci disrupt the amount (tds1, tds2, tds3, tds5, and tds6) or location and amount of PA (tds4) in the endothelial cell layer. The PA intermediate epicatechin has been identified in wild type and mutants tds1, tds2, tds3, and tds5 (which do not produce PA) and tds6 (6% of wild-type PA), whereas tds4 (2% of wild-type PA) produces an unidentified dimethylaminocinnamaldehyde-reacting compound, indicating that the mutations may be acting on genes beyond leucoanthocyanidin reductase, the first enzymatic reduction step dedicated to PA synthesis. Two other mutants were identified, an allele of tt7, which has a spotted pattern of PA deposition and produces only 8% of the wild-type level of type PA as propelargonidin, and an allele of tt8 producing no PA. Spotted patterns of PA deposition observed in seed of mutants tds4 and tt7-3 result from altered PA composition and distribution in the cell. Our mutant screen, which was not exhaustive, suggests that the cooperation of many genes is required for successful PA accumulation. PMID:12376625

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

    PubMed Central

    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

  16. 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

  17. Embryo and endosperm development is disrupted in the female gametophytic capulet mutants of Arabidopsis.

    PubMed Central

    Grini, Paul E; Jürgens, Gerd; Hülskamp, Martin

    2002-01-01

    The female gametophyte of higher plants gives rise, by double fertilization, to the diploid embryo and triploid endosperm, which develop in concert to produce the mature seed. What roles gametophytic maternal factors play in this process is not clear. The female-gametophytic effects on embryo and endosperm development in the Arabidopsis mea, fis, and fie mutants appear to be due to gametic imprinting that can be suppressed by METHYL TRANSFERASE1 antisense (MET1 a/s) transgene expression or by mutation of the DECREASE IN DNA METHYLATION1 (DDM1) gene. Here we describe two novel gametophytic maternal-effect mutants, capulet1 (cap1) and capulet2 (cap2). In the cap1 mutant, both embryo and endosperm development are arrested at early stages. In the cap2 mutant, endosperm development is blocked at very early stages, whereas embryos can develop to the early heart stage. The cap mutant phenotypes were not rescued by wild-type pollen nor by pollen from tetraploid plants. Furthermore, removal of silencing barriers from the paternal genome by MET1 a/s transgene expression or by the ddm1 mutation also failed to restore seed development in the cap mutants. Neither cap1 nor cap2 displayed autonomous seed development, in contrast to mea, fis, and fie mutants. In addition, cap2 was epistatic to fis1 in both autonomous endosperm and sexual development. Finally, both cap1 and cap2 mutant endosperms, like wild-type endosperms, expressed the paternally inactive endosperm-specific FIS2 promoter GUS fusion transgene only when the transgene was introduced via the embryo sac, indicating that imprinting was not affected. Our results suggest that the CAP genes represent novel maternal functions supplied by the female gametophyte that are required for embryo and endosperm development. PMID:12524359

  18. 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.

  19. 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.

  20. 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

  1. 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.

  2. Light-stimulated cotyledon expansion in the blu3 and hy4 mutants of Arabidopsis thaliana.

    PubMed

    Blum, D E; Neff, M M; Van Volkenburgh, E

    1994-08-01

    Cotyledon expansion in response to blue light was compared for wild-type Arabidopsis thaliana (L.) Heynh. and the mutants blu3 and hy4, which show reduced inhibition of hypocotyl growth in blue light. White, blue, and red light stimulated cotyledon expansion in both intact and excised cotyledons of wild-type seedlings (ecotypes No-0, WS, Co-0, La-er). Cotyledons on intact blu3 and hy4 seedlings did not grow as well as those on the wild type in response to blue light, but pretreatment of blu3 seedlings with low fluence rates of red light increased their responsiveness to blue light. Excision of cotyledons alleviated the mutant phenotype so that both mutant and wild-type cotyledons grew equally well in blue light. The loss of the mutant cotyledon phenotype upon excision indicates that the blu3 and hy4 lesions affect cotyledon expansion indirectly via a whole-plant response to light. Furthermore, the ability of excised, mutant cotyledons to grow normally in blue light shows that this growth response to blue light is mediated by a photosystem other than the ones impaired by the blu3 and hy4 lesions. PMID:7972499

  3. The Critical Requirement for Linolenic Acid Is Pollen Development, Not Photosynthesis, in an Arabidopsis Mutant.

    PubMed Central

    McConn, M; Browse, J

    1996-01-01

    The very high proportions of trienoic fatty acids found in chloroplast membranes of all higher plants suggest that these lipid structures might be essential for photosynthesis. We report here on the production of Arabidopsis triple mutants that contain negligible levels of trienoic fatty acids. Photosynthesis at 22[deg]C was barely affected, and vegetative growth of the mutants was identical with that of the wild type, demonstrating that any requirement for trienoic acyl groups in membrane structure and function is relatively subtle. Although vegetative growth and development were unaffected, the triple mutants are male sterlle and produce no seed under normal conditions. Comparisons of pollen development in wild-type and triple mutant flowers established that pollen grains in the mutant developed to the tricellular stage. Exogenous applications of [alpha]-llnolenate or jasmonate restored fertility. Taken together, the results demonstrate that the critical role of trienoic acids in the life cycle of plants is as the precursor of oxylipin, a signaling compound that regulates final maturation processes and the release of pollen. PMID:12239389

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

    PubMed Central

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

    2010-01-01

    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. PMID:20547563

  5. Large genetic screens for gynogenesis and androgenesis haploid inducers in Arabidopsis thaliana failed to identify mutants

    PubMed Central

    Portemer, Virginie; Renne, Charlotte; Guillebaux, Alexia; Mercier, Raphael

    2015-01-01

    Gynogenesis is a process in which the embryo genome originates exclusively from female origin, following embryogenesis stimulation by a male gamete. In contrast, androgenesis is the development of embryos that contain only the male nuclear genetic background. Both phenomena are of great interest in plant breeding as haploidization is an efficient tool to reduce the length of breeding schemes to create varieties. Although few inducer lines have been described, the genetic control of these phenomena is poorly understood. We developed genetic screens to identify mutations that would induce gynogenesis or androgenesis in Arabidopsis thaliana. The ability of mutant pollen to induce either gynogenesis or androgenesis was tested by crossing mutagenized plants as males. Seedlings from these crosses were screened with recessive phenotypic markers, one genetically controlled by the female genome and another by the male genome. Positive and negative controls confirmed the unambiguous detection of both gynogenesis and androgenesis events. This strategy was applied to 1,666 EMS-mutagenised lines and 47 distant Arabidopsis strains. While an internal control suggested that the mutagenesis reached saturation, no gynogenesis or androgenesis inducer was found. However, spontaneous gynogenesis was observed at a frequency of 1/10,800. Altogether, these results suggest that no simple EMS-induced mutation in the male genome is able to induce gynogenesis or androgenesis in Arabidopsis. PMID:25814999

  6. 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.

  7. Isolation and Characterization of Mutants Defective in Seed Coat Mucilage Secretory Cell Development in Arabidopsis1

    PubMed Central

    Western, Tamara L.; Burn, Joanne; Tan, Wei Ling; Skinner, Debra J.; Martin-McCaffrey, Luke; Moffatt, Barbara A.; Haughn, George W.

    2001-01-01

    In Arabidopsis, fertilization induces the epidermal cells of the outer ovule integument to differentiate into a specialized seed coat cell type producing extracellular pectinaceous mucilage and a volcano-shaped secondary cell wall. Differentiation involves a regulated series of cytological events including growth, cytoplasmic rearrangement, mucilage synthesis, and secondary cell wall production. We have tested the potential of Arabidopsis seed coat epidermal cells as a model system for the genetic analysis of these processes. A screen for mutants defective in seed mucilage identified five novel genes (MUCILAGE-MODIFIED [MUM]1–5). The seed coat development of these mutants, and that of three previously identified ones (TRANSPARENT TESTA GLABRA1, GLABRA2, and APETALA2) were characterized. Our results show that the genes identified define several events in seed coat differentiation. Although APETALA2 is needed for differentiation of both outer layers of the seed coat, TRANSPARENT TESTA GLABRA1, GLABRA2, and MUM4 are required for complete mucilage synthesis and cytoplasmic rearrangement. MUM3 and MUM5 may be involved in the regulation of mucilage composition, whereas MUM1 and MUM2 appear to play novel roles in post-synthesis cell wall modifications necessary for mucilage extrusion. PMID:11706181

  8. 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

  9. 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.

  10. 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. PMID:25740148

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

    PubMed

    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

  12. Differential sensitivity of Arabidopsis siRNA biogenesis mutants to genotoxic stress.

    PubMed

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

    2010-12-01

    Plant response to stress has been linked to different RNA-silencing processes and epigenetic mechanisms. Our recent results showed that Arabidopsis thaliana Dicer-like (DCL) mutants were impaired in transgenerational changes, recombination frequency and stress tolerance. We also found that transgenerational changes were dependent on changes in DNA methylation. Here, we hypothesized that plants deficient in the production of small RNAs would show an impaired abiotic stress response. To test this, we exposed A. thaliana dcl2, dcl3, dcl4, dcl2 dcl3 (d2d3), dcl2 dcl4 (d2d4), dcl2 dcl3 dcl4 (d2d3d4), nrpd1a, rdr2 and rdr6 mutants to methyl methane sulfonate (MMS). We found dcl4 and rdr6 to be more sensitive and dcl2, dcl3, d2d3 and rdr2 plants more resistant to MMS, as shown by fresh weight, root length and survival rate. The in vitro repair assay showed the lower ability of dcl2 and dcl3 to repair UV-damaged DNA. To summarize, we found that whereas mutants impaired in transactivating siRNA biogenesis were more sensitive to MMS, mutants impaired in natural antisense siRNA and heterochromatic siRNA biogeneses were more tolerant. Our data suggest that plant response to MMS is in part regulated through biogenesis of various siRNAs. PMID:20953786

  13. 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

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

    PubMed Central

    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. PMID:25610446

  15. 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.

  16. 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

  17. 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'. PMID:27122571

  18. 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

  19. Properties of proteins and the glassy matrix in maturation-defective mutant seeds of Arabidopsis thaliana.

    PubMed

    Wolkers, W F; Alberda, M; Koornneef, M; Léon-Kloosterziel, K M; Hoekstra, F A

    1998-10-01

    In situ Fourier transform infrared microspectroscopy was used to study the heat stability of proteins and hydrogen bonding interactions in dry maturation-defective mutant seeds of Arabidopsis thaliana. alpha-Helical, turn and beta-sheet conformations were the major protein secondary structures in all of these seeds. On heating, intermolecular extended beta-sheet structures, typical of protein denaturation, were formed in abscisic acid-insensitive (abi3) and leafy cotyledon (lec) mutant seeds. Proteins in dry wild-type seeds did not denature up to 150 degrees C, but those in dry desiccation-sensitive, lec1-1, lec1-3 and abi3-5 seeds did at 68, 89 and 87 degrees C, respectively. In the desiccation-tolerant abi3-7 and abi3-1 seeds, denaturation commenced above 120 and 135 degrees C, respectively. Seeds of the aba1-1 abi3-1 double mutant showed signs of denaturation already upon drying. The molecular packing in the seeds was studied by observing the shift in the position of the OH-stretching vibration band with temperature. The maximal rate of change of this band with temperature was much higher in the desiccation-sensitive abi3-5, aba1-1 abi3-1, lec1-1, and lec1-3 mutant seeds than in the desiccation-tolerant wild-type, abi3-1, abi3-7, and lec2-1 seeds. We interpret this to mean that the molecular packing density is higher in dry desiccation-tolerant than in dry desiccation-sensitive seeds, which is associated with a higher or lower protein denaturation temperature, respectively. The results are discussed in relation to the physiological and biochemical characteristics of these mutant seeds. PMID:9839460

  20. 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. PMID:26520913

  1. Biochemical characterization of the aba2 and aba3 mutants in Arabidopsis thaliana.

    PubMed Central

    Schwartz, S H; Léon-Kloosterziel, K M; Koornneef, M; Zeevaart, J A

    1997-01-01

    Abscisic acid (ABA)-deficient mutants in a variety of species have been identified by screening for precocious germination and a wilty phenotype. Mutants at two new loci, aba2 and aba3, have recently been isolated in Arabidopsis thaliana (L.) Hynh. (K.M. Léon-Kloosterziel, M. Alvarez-Gil, G.J. Ruijs, S.E. Jacobsen, N.E. Olszewski, S.H. Schwartz, J.A.D. Zeevaart, M. Koornneef [1996] Plant J 10: 655-661), and the biochemical characterization of these mutants is presented here. Protein extracts from aba2 and aba3 plants displayed a greatly reduced ability to convert xanthoxin to ABA relative to the wild type. The next putative intermediate in ABA synthesis, ABA-aldehyde, was efficiently converted to ABA by extracts from aba2 but not by extracts from aba3 plants. This indicates that the aba2 mutant is blocked in the conversion of xanthoxin to ABA-aldehyde and that aba3 is impaired in the conversion of ABA-aldehyde to ABA. Extracts from the aba3 mutant also lacked additional activities that require a molybdenum cofactor (Moco). Nitrate reductase utilizes a Moco but its activity was unaffected in extracts from aba3 plants. Moco hydroxylases in animals require a desulfo moiety of the cofactor. A sulfido ligand can be added to the Moco by treatment with Na2S and dithionite. Treatment of aba3 extracts with Na2S restored ABA-aldehyde oxidase activity. Therefore, the genetic lesion in aba3 appears to be in the introduction of S into the Moco. PMID:9159947

  2. Biochemical characterization of the aba2 and aba3 mutants in Arabidopsis thaliana.

    PubMed

    Schwartz, S H; Léon-Kloosterziel, K M; Koornneef, M; Zeevaart, J A

    1997-05-01

    Abscisic acid (ABA)-deficient mutants in a variety of species have been identified by screening for precocious germination and a wilty phenotype. Mutants at two new loci, aba2 and aba3, have recently been isolated in Arabidopsis thaliana (L.) Hynh. (K.M. Léon-Kloosterziel, M. Alvarez-Gil, G.J. Ruijs, S.E. Jacobsen, N.E. Olszewski, S.H. Schwartz, J.A.D. Zeevaart, M. Koornneef [1996] Plant J 10: 655-661), and the biochemical characterization of these mutants is presented here. Protein extracts from aba2 and aba3 plants displayed a greatly reduced ability to convert xanthoxin to ABA relative to the wild type. The next putative intermediate in ABA synthesis, ABA-aldehyde, was efficiently converted to ABA by extracts from aba2 but not by extracts from aba3 plants. This indicates that the aba2 mutant is blocked in the conversion of xanthoxin to ABA-aldehyde and that aba3 is impaired in the conversion of ABA-aldehyde to ABA. Extracts from the aba3 mutant also lacked additional activities that require a molybdenum cofactor (Moco). Nitrate reductase utilizes a Moco but its activity was unaffected in extracts from aba3 plants. Moco hydroxylases in animals require a desulfo moiety of the cofactor. A sulfido ligand can be added to the Moco by treatment with Na2S and dithionite. Treatment of aba3 extracts with Na2S restored ABA-aldehyde oxidase activity. Therefore, the genetic lesion in aba3 appears to be in the introduction of S into the Moco. PMID:9159947

  3. 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

  4. A NEW ARABIDOPSIS THALIANA MUTANT DEFICIENT IN THE EXPRESSION OF OMETHYLTRANSFERASE 1: IMPACT ON LIGNINS AND ON SINAPOYL ESTERI

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A promoter-trap screen allowed us to identify an Arabidopsis line expressing GUS in the root vascular tissues. T-DNA border sequencing showed that the line was mutated in the COMT 1 gene (AtOMT1) and therefore deficient in OMT1 activity. In this knockout mutant and relative to the wild type, lignins...

  5. ahg12 is a dominant proteasome mutant that affects multiple regulatory systems for germination of Arabidopsis.

    PubMed

    Hayashi, Shimpei; Hirayama, Takashi

    2016-01-01

    The ubiquitin-proteasome system is fundamentally involved in myriad biological phenomena of eukaryotes. In plants, this regulated protein degradation system has a pivotal role in the cellular response mechanisms for both internal and external stimuli, such as plant hormones and environmental stresses. Information about substrate selection by the ubiquitination machinery has accumulated, but there is very little information about selectivity for substrates at the proteasome. Here, we report characterization of a novel abscisic acid (ABA)-hypersensitive mutant named ABA hypersensitive germination12 (ahg12) in Arabidopsis. The ahg12 mutant showed a unique pleiotropic phenotype, including hypersensitivity to ABA and ethylene, and hyposensitivity to light. Map-based cloning identified the ahg12 mutation to cause an amino acid conversion in the L23 loop of RPT5a, which is predicted to form the pore structure of the 19S RP complex of the proteasome. Transient expression assays demonstrated that some plant-specific signaling components accumulated at higher levels in the ahg12 mutant. These results suggest that the ahg12 mutation led to changes in the substrate preference of the 26S proteasome. The discovery of the ahg12 mutation thus will contribute to elucidate the characteristics of the regulated protein degradation system. PMID:27139926

  6. ahg12 is a dominant proteasome mutant that affects multiple regulatory systems for germination of Arabidopsis

    PubMed Central

    Hayashi, Shimpei; Hirayama, Takashi

    2016-01-01

    The ubiquitin-proteasome system is fundamentally involved in myriad biological phenomena of eukaryotes. In plants, this regulated protein degradation system has a pivotal role in the cellular response mechanisms for both internal and external stimuli, such as plant hormones and environmental stresses. Information about substrate selection by the ubiquitination machinery has accumulated, but there is very little information about selectivity for substrates at the proteasome. Here, we report characterization of a novel abscisic acid (ABA)-hypersensitive mutant named ABA hypersensitive germination12 (ahg12) in Arabidopsis. The ahg12 mutant showed a unique pleiotropic phenotype, including hypersensitivity to ABA and ethylene, and hyposensitivity to light. Map-based cloning identified the ahg12 mutation to cause an amino acid conversion in the L23 loop of RPT5a, which is predicted to form the pore structure of the 19S RP complex of the proteasome. Transient expression assays demonstrated that some plant-specific signaling components accumulated at higher levels in the ahg12 mutant. These results suggest that the ahg12 mutation led to changes in the substrate preference of the 26S proteasome. The discovery of the ahg12 mutation thus will contribute to elucidate the characteristics of the regulated protein degradation system. PMID:27139926

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

    PubMed Central

    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. PMID:27468291

  8. 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

  9. A mutant of Arabidopsis deficient in desaturation of palmitic acid in leaf lipids

    SciTech Connect

    Kunst, L.; Somerville, C. ); Browse, J. )

    1989-07-01

    The overall fatty acid composition of leaf lipids in a mutant of Arabidopsis thaliana was characterized by elevated amounts of palmitic acid and a decreased amount of unsaturated 16-carbon fatty acids as a consequence of a single nuclear mutation. Quantitative analysis of the fatty acid composition of individual lipids suggested that the mutant is deficient in the activity of a chloroplast {omega}9 fatty acid desaturase which normally introduces a double bond in 16-carbon acyl chains esterified to monogalactosyldiacylglycerol (MGD). The mutant exhibited an increased ratio of 18- to 16-carbon fatty acids in MGD due to a change in the relative contribution of the prokaryotic and eukaryotic pathways of lipid biosynthesis. This appears to be a regulated response to the loss of chloroplast {omega}9 desaturase and presumably reflects a requirement for polyunsaturated fatty acids for the normal assembly of chloroplast membranes. The reduction in mass of prokaryotic MGD species involved both a reduction in synthesis of MGD by the prokaryotic pathway and increased turnover of MGD molecular species which contain 16:0.

  10. Biochemical characterization of Arabidopsis wild-type and mutant phytochrome B holoproteins.

    PubMed Central

    Elich, T D; Chory, J

    1997-01-01

    Although phytochrome B (phyB) plays a particularly important role throughout the life cycle of a plant, it has not been studied in detail at the molecular level due to its low abundance. Here, we report on the expression, assembly with chromophore, and purification of epitope-tagged Arabidopsis phyB. In addition, we have reconstructed two missense mutations, phyB-4 and phyB-101, isolated in long hypocotyl screens. We show that mutant proteins phyB-4 and phyB-101 exhibit altered spectrophotometric and biochemical properties relative to the wild-type protein. In particular, we demonstrate that phyB-101 Pfr exhibits rapid nonphotochemical (dark) reversion to Pr that results in a lower photoequilibrium level of the active Pfr form. We conclude that this occurs in vivo as well because phyB-101 mutants are shown to lack an end-of-day-far-red hypocotyl elongation response that requires a stable Pfr species. We propose that this Pfr instability may be the primary molecular mechanism underlying the phyB-101 mutant phenotype. PMID:9437866

  11. 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

  12. 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.

  13. Mutants of Arabidopsis thaliana with Decreased Amplitude in Their Phototropic Response 1

    PubMed Central

    Khurana, Jitendra P.; Ren, Zhangling; Steinitz, Benjamin; Parks, Brian; Best, Thérèse R.; Poff, Kenneth 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. PMID:11537461

  14. An Arabidopsis syntaxin homologue isolated by functional complementation of a yeast pep12 mutant

    SciTech Connect

    Bassham, D.C.; Conceicao, A.S. Da; Gal, S.

    1995-08-01

    The syntaxin family of integral membrane proteins are thought to function as receptors for transport vesicles, with different isoforms of this family localized to various membranes throughout the cell. The yeast Pep12 protein is a syntaxin homologue which may function in the trafficking of vesicles from the trans-Golgi network to the vacuole. We have isolated an Arabidopsis thaliana cDNA by functional complementation of a yeast pep12 mutant. The Arabidopsis cDNA (aPEP12) potentially encodes a 31-kDa protein which is homologous to yeast Pep12 and to other members of the syntaxin family, indicating that this protein may function in the docking or fusion of transport vesicles with the vacuolar membrane in plant cells. Northern blot analysis indicates that the mRNA is expressed in all tissues examined, although at a very low level in leaves. The mRNA is found in all cell types in roots and leaves, as shown by in situ hybridization experiments. The existence of plant homologues of proteins of the syntaxin family indicates that the basic vesicle docking and fusion machinery may be conserved in plants as it is in yeast and mammals. 41 refs., 6 figs.

  15. Reduced Na+ uptake in the NaCl-hypersensitive sos1 mutant of Arabidopsis thaliana.

    PubMed Central

    Ding, L; Zhu, J K

    1997-01-01

    Sos1 is an Arabidopsis thaliana mutant with > 20 times higher sensitivity toward Na+ inhibition due to a defective high-affinity potassium-uptake system. We report here that sos1 accumulates less Na+ than the wild type in response to NaCl stress. The Na+ contents in sos1 seedlings exposed to 25 mM NaCl for 2 or more d are about 43% lower than those in the wild type. When assayed at 20 mM external NaCl, sos1 seedlings pretreated with low potassium have 32% lower Na+ uptake than the wild type. However, little difference in Na+ uptake could be measured when the seedlings were not pretreated with low potassium. Low-potassium treatment was shown to induce high-affinity potassium-uptake activity in Arabidopsis seedlings. No substantial difference in Na+ efflux between sos1 and the wild type was detected. The results show that the reduced Na+ accumulation in sos1 is due to a lower Na+ influx rate. Therefore, the sos1 mutation appears to disrupt low-affinity Na+ uptake in addition to its impairment of high-affinity K+ uptake. PMID:9085573

  16. 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

  17. Cortical microtubule patterning in roots of Arabidopsis thaliana primary cell wall mutants reveals the bidirectional interplay with cell expansion.

    PubMed

    Panteris, Emmanuel; Adamakis, Ioannis-Dimosthenis S; Daras, Gerasimos; Rigas, Stamatis

    2014-04-01

    Cell elongation requires directional deposition of cellulose microfibrils regulated by transverse cortical microtubules. Microtubules respond differentially to suppression of cell elongation along the developmental zones of Arabidopsis thaliana root apex. Cortical microtubule orientation is particularly affected in the fast elongation zone but not in the meristematic or transition zones of thanatos and pom2-4 cellulose-deficient mutants of Arabidopsis thaliana. Here, we report that a uniform phenotype is established among the primary cell wall mutants, as cortical microtubules of root epidermal cells of rsw1 and prc1 mutants exhibit the same pattern described in thanatos and pom2-4. Whether cortical microtubules assume transverse orientation or not is determined by the demand for cellulose synthesis, according to each root zone's expansion rate. It is suggested that cessation of cell expansion may provide a biophysical signal resulting in microtubule reorientation. PMID:24717634

  18. Cortical microtubule patterning in roots of Arabidopsis thaliana primary cell wall mutants reveals the bidirectional interplay with cell expansion

    PubMed Central

    Panteris, Emmanuel; Adamakis, Ioannis-Dimosthenis S; Daras, Gerasimos; Rigas, Stamatis

    2014-01-01

    Cell elongation requires directional deposition of cellulose microfibrils regulated by transverse cortical microtubules. Microtubules respond differentially to suppression of cell elongation along the developmental zones of Arabidopsis thaliana root apex. Cortical microtubule orientation is particularly affected in the fast elongation zone but not in the meristematic or transition zones of thanatos and pom2–4 cellulose-deficient mutants of Arabidopsis thaliana. Here, we report that a uniform phenotype is established among the primary cell wall mutants, as cortical microtubules of root epidermal cells of rsw1 and prc1 mutants exhibit the same pattern described in thanatos and pom2–4. Whether cortical microtubules assume transverse orientation or not is determined by the demand for cellulose synthesis, according to each root zone’s expansion rate. It is suggested that cessation of cell expansion may provide a biophysical signal resulting in microtubule reorientation. PMID:24717634

  19. Cortical microtubule patterning in roots of Arabidopsis thaliana primary cell wall mutants reveals the bidirectional interplay with cell expansion

    PubMed Central

    Panteris, Emmanuel; Adamakis, Ioannis-Dimosthenis S; Daras, Gerasimos; Rigas, Stamatis

    2015-01-01

    Cell elongation requires directional deposition of cellulose microfibrils regulated by transverse cortical microtubules. Microtubules respond differentially to suppression of cell elongation along the developmental zones of Arabidopsis thaliana root apex. Cortical microtubule orientation is particularly affected in the fast elongation zone but not in the meristematic or transition zones of thanatos and pom2–4 cellulose-deficient mutants of Arabidopsis thaliana. Here, we report that a uniform phenotype is established among the primary cell wall mutants, as cortical microtubules of root epidermal cells of rsw1 and prc1 mutants exhibit the same pattern described in thanatos and pom2–4. Whether cortical microtubules assume transverse orientation or not is determined by the demand for cellulose synthesis, according to each root zone's expansion rate. It is suggested that cessation of cell expansion may provide a biophysical signal resulting in microtubule reorientation. PMID:26042727

  20. Cortical microtubule patterning in roots of Arabidopsis thaliana primary cell wall mutants reveals the bidirectional interplay with cell expansion.

    PubMed

    Panteris, Emmanuel; Adamakis, Ioannis-Dimosthenis S; Daras, Gerasimos; Rigas, Stamatis

    2015-01-01

    Cell elongation requires directional deposition of cellulose microfibrils regulated by transverse cortical microtubules. Microtubules respond differentially to suppression of cell elongation along the developmental zones of Arabidopsis thaliana root apex. Cortical microtubule orientation is particularly affected in the fast elongation zone but not in the meristematic or transition zones of thanatos and pom2-4 cellulose-deficient mutants of Arabidopsis thaliana. Here, we report that a uniform phenotype is established among the primary cell wall mutants, as cortical microtubules of root epidermal cells of rsw1 and prc1 mutants exhibit the same pattern described in thanatos and pom2-4. Whether cortical microtubules assume transverse orientation or not is determined by the demand for cellulose synthesis, according to each root zone's expansion rate. It is suggested that cessation of cell expansion may provide a biophysical signal resulting in microtubule reorientation. PMID:26042727

  1. Regulated ethylene insensitivity through the inducible expression of the Arabidopsis etr1-1 mutant ethylene receptor in tomato.

    PubMed

    Gallie, Daniel R

    2010-04-01

    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

  2. Transformation of shoots into roots in Arabidopsis embryos mutant at the TOPLESS locus.

    PubMed

    Long, Jeff A; Woody, Scott; Poethig, Scott; Meyerowitz, Elliot M; Barton, M Kathryn

    2002-06-01

    We describe a novel phenotype in Arabidopsis embryos homozygous for the temperature-sensitive topless-1 mutation. This mutation causes the transformation of the shoot pole into a root. Developing topless embryos fail to express markers for the shoot apical meristem (SHOOT MERISTEMLESS and UNUSUAL FLORAL ORGANS) and the hypocotyl (KNAT1). By contrast, the pattern of expression of root markers is either duplicated (LENNY, J1092) or expanded (SCARECROW). Shifts of developing topless embryos between permissive and restrictive temperatures show that apical fates (cotyledons plus shoot apical meristem) can be transformed to basal fates (root) as late as transition stage. As the apical pole of transition stage embryos shows both morphological and molecular characteristics of shoot development, this demonstrates that the topless 1 mutation is capable of causing structures specified as shoot to be respecified as root. Finally, our experiments fail to show a clear link between auxin signal transduction and topless-1 mutant activity: the development of the apical root in topless mutant individuals is not dependent on the activity of the predicted auxin response factor MONOPTEROS nor is the expression of DR5, a proposed 'auxin maximum reporter', expanded in the apical domain of topless embryos. PMID:12050130

  3. Overproduction of stomatal lineage cells in Arabidopsis mutants defective in active DNA demethylation

    PubMed Central

    Yamamuro, Chizuko; Miki, Daisuke; Zheng, Zhimin; Ma, Jun; Wang, Jing; Yang, Zhenbiao; Dong, Juan; Zhu, Jian-Kang

    2014-01-01

    DNA methylation is a reversible epigenetic mark regulating genome stability and function in many eukaryotes. In Arabidopsis, active DNA demethylation depends on the function of the ROS1 subfamily of genes that encode 5-methylcytosine DNA glycosylases/lyases. ROS1-mediated DNA demethylation plays a critical role in the regulation of transgenes, transposable elements and some endogenous genes, but there have been no reports of clear developmental phenotypes in ros1 mutant plants. Here we report that, in the ros1 mutant, the promoter region of the peptide ligand gene EPF2 is hypermethylated, which greatly reduces EPF2 expression and thereby leads to a phenotype of overproduction of stomatal lineage cells. EPF2 gene expression in ros1 is restored and the defective epidermal cell patterning is suppressed by mutations in genes in the RNA-directed DNA methylation pathway. Our results show that active DNA demethylation combats the activity of RNA-directed DNA methylation to influence the initiation of stomatal lineage cells. PMID:24898766

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

    PubMed

    Pacurar, Daniel Ioan; Pacurar, Monica Lacramioara; Bussell, John Desmond; Schwambach, Joseli; Pop, Tiberia Ioana; Kowalczyk, Mariusz; Gutierrez, Laurent; Cavel, Emilie; Chaabouni, Salma; Ljung, Karin; Fett-Neto, Arthur Germano; Pamfil, Doru; Bellini, Catherine

    2014-04-01

    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

  5. The arc mutants of Arabidopsis with fewer large chloroplasts have a lower mesophyll conductance.

    PubMed

    Weise, Sean E; Carr, David J; Bourke, Ashley M; Hanson, David T; Swarthout, Debbie; Sharkey, Thomas D

    2015-04-01

    Photosynthetic cells of most land plant lineages have numerous small chloroplasts even though most algae, and even the early diverging land plant group the hornworts, tend to have one or a few large chloroplasts. One constraint that small chloroplasts could improve is the resistance to CO2 diffusion from the atmosphere to the chloroplast stroma. We examined the mesophyll conductance (inverse of the diffusion resistance) of mutant Arabidopsis thaliana plants with one or only a few large chloroplasts per cell. The accumulation and replication of chloroplasts (arc) mutants of A. thaliana were studied by model fitting to gas exchange data and (13)CO2 discrimination during carbon fixation. The two methods generally agreed, but the value of the CO2 compensation point of Rubisco (Γ *) used in the model had a large impact on the estimated photosynthetic parameters, including mesophyll conductance. We found that having only a few large chloroplasts per cell resulted in a 25-50 % reduction in the mesophyll conductance at ambient CO2. PMID:25733184

  6. Protection of Chloroplast Membranes by VIPP1 Rescues Aberrant Seedling Development in Arabidopsis nyc1 Mutant

    PubMed Central

    Zhang, Lingang; Kusaba, Makoto; Tanaka, Ayumi; Sakamoto, Wataru

    2016-01-01

    Chlorophylls (Chl) in photosynthetic apparatuses, along with other macromolecules in chloroplasts, are known to undergo degradation during leaf senescence. Several enzymes involved in Chl degradation, by which detoxification of Chl is safely implemented, have been identified. Chl degradation also occurs during embryogenesis and seedling development. Some genes encoding Chl degradation enzymes such as Chl b reductase (CBR) function during these developmental stages. Arabidopsis mutants lacking CBR (NYC1 and NOL) have been reported to exhibit reduced seed storability, compromised germination, and cotyledon development. In this study, we examined aberrant cotyledon development and found that NYC1 is solely responsible for this phenotype. We inferred that oxidative damage of chloroplast membranes caused the aberrant cotyledon. To test the inference, we attempted to trans-complement nyc1 mutant with overexpressing VIPP1 protein that is unrelated to Chl degradation but which supports chloroplast membrane integrity. VIPP1 expression actually complemented the aberrant cotyledon of nyc1, whereas stay-green phenotype during leaf senescence remained. The swollen chloroplasts observed in unfixed cotyledons of nyc1, which are characteristics of chloroplasts receiving envelope membrane damage, were recovered by overexpressing VIPP1. These results suggest that chloroplast membranes are a target for oxidative damage caused by the impairment in Chl degradation. Trans-complementation of nyc1 with VIPP1 also suggests that VIPP1 is useful for protecting chloroplasts against oxidative stress. PMID:27200011

  7. 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.

  8. Hormonal Studies of fass, an Arabidopsis Mutant That Is Altered in Organ Elongation.

    PubMed Central

    Fisher, R. H.; Barton, M. K.; Cohen, J. D.; Cooke, T. J.

    1996-01-01

    We have isolated an allele of fass, an Arabidopsis thaliana mutation that separates plant development and organ differentiation from plant elongation, and studied its hormonal regulation. Micro-surgically isolated fass roots elongate 2.5 times as much as the roots on intact mutant plants. Wild-type heart embryos, when cultured with a strong auxin, naphthaleneacetic acid, phenocopy fass embryos. fass seedlings contain variable levels of free auxin, which average 2.5 times higher than wild-type seedling levels, and fass seedlings evolve 3 times as much ethylene as wild-type seedlings on a per-plant basis over a 24-h period. The length-to-width ratios of fass seedlings can be changed by several compounds that affect their endogenous ethylene levels, but fass is epistatic to etr1, an ethylene-insensitive mutant. fass's high levels of free auxin may be inducing its high levels of ethylene, which may, in turn, result in the fass phenotype. We postulate that FASS may be acting as a negative regulator to maintain wild-type auxin levels and that the mutation may be in an auxin-conjugating enzyme. PMID:12226245

  9. Hyperspectral imaging techniques for rapid identification of Arabidopsis mutants with altered leaf pigment status.

    PubMed

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

    2012-06-01

    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

  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. MicroRNAs and other small RNAs enriched in the Arabidopsis RNA-dependent RNA polymerase-2 mutant.

    PubMed

    Lu, Cheng; Kulkarni, Karthik; Souret, Frédéric F; MuthuValliappan, Ramesh; Tej, Shivakundan Singh; Poethig, R Scott; Henderson, Ian R; Jacobsen, Steven E; Wang, Wenzhong; Green, Pamela J; Meyers, Blake C

    2006-10-01

    The Arabidopsis genome contains a highly complex and abundant population of small RNAs, and many of the endogenous siRNAs are dependent on RNA-Dependent RNA Polymerase 2 (RDR2) for their biogenesis. By analyzing an rdr2 loss-of-function mutant using two different parallel sequencing technologies, MPSS and 454, we characterized the complement of miRNAs expressed in Arabidopsis inflorescence to considerable depth. Nearly all known miRNAs were enriched in this mutant and we identified 13 new miRNAs, all of which were relatively low abundance and constitute new families. Trans-acting siRNAs (ta-siRNAs) were even more highly enriched. Computational and gel blot analyses suggested that the minimal number of miRNAs in Arabidopsis is approximately 155. The size profile of small RNAs in rdr2 reflected enrichment of 21-nt miRNAs and other classes of siRNAs like ta-siRNAs, and a significant reduction in 24-nt heterochromatic siRNAs. Other classes of small RNAs were found to be RDR2-independent, particularly those derived from long inverted repeats and a subset of tandem repeats. The small RNA populations in other Arabidopsis small RNA biogenesis mutants were also examined; a dcl2/3/4 triple mutant showed a similar pattern to rdr2, whereas dcl1-7 and rdr6 showed reductions in miRNAs and ta-siRNAs consistent with their activities in the biogenesis of these types of small RNAs. Deep sequencing of mutants provides a genetic approach for the dissection and characterization of diverse small RNA populations and the identification of low abundance miRNAs. PMID:16954541

  12. 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. PMID:21388819

  13. Ectopic expression of a Catalpa bungei (Bignoniaceae) PISTILLATA homologue rescues the petal and stamen identities in Arabidopsis pi-1 mutant.

    PubMed

    Jing, Danlong; Xia, Yan; Chen, Faju; Wang, Zhi; Zhang, Shougong; Wang, Junhui

    2015-02-01

    PISTILLATA (PI) plays crucial roles in Arabidopsis flower development by specifying petal and stamen identities. To investigate the molecular mechanisms underlying organ development of woody angiosperm in Catalpa, we isolated and identified a PI homologue, referred to as CabuPI (C. bungei PISTILLATA), from two genetically cognate C. bungei (Bignoniaceae) bearing single and double flowers. Sequence and phylogenetic analyses revealed that the gene is closest related to the eudicot PI homologues. Moreover, a highly conserved PI-motif is found in the C-terminal regions of CabuPI. Semi-quantitative and quantitative real time PCR analyses showed that the expression of CabuPI was restricted to petals and stamens. However, CabuPI expression in the petals and stamens persisted throughout all floral development stages, but the expression levels were different. In 35S::CabuPI transgenic homozygous pi-1 mutant Arabidopsis, the second and the third whorl floral organs produced normal petals and a different number of stamens, respectively. Furthermore, ectopic expression of the CabuPI in transgenic wild-type or heterozygote pi-1 mutant Arabidopsis caused the first whorl sepal partially converted into a petal-like structure. These results clearly reveal the functional conservation of PI homologues between C. bungei and Arabidopsis. PMID:25575990

  14. The Arabidopsis ABHD11 Mutant Accumulates Polar Lipids in Leaves as a Consequence of Absent Acylhydrolase Activity1[OPEN

    PubMed Central

    Vijayakumar, Anitha; Vijayaraj, Panneerselvam; Vijayakumar, Arun Kumar; Rajasekharan, Ram

    2016-01-01

    Alpha/beta hydrolase domain (ABHD)-containing proteins are structurally related with diverse catalytic activities. In various species, some ABHD proteins have been characterized and shown to play roles in lipid homeostasis. However, little is known about ABHD proteins in plants. Here, we characterized AT4G10030 (AtABHD11), an Arabidopsis (Arabidopsis thaliana) homolog of a human ABHD11 gene. In silico analyses of AtABHD11 revealed homology with other plant species with a conserved GXSXG lipid motif. Interestingly, Arabidopsis abhd11 mutant plants exhibited an enhanced growth rate compared with wild-type plants. Quantitative analyses of the total lipids showed that the mutant abhd11 has a high amount of phospholipid and galactolipid in Arabidopsis leaves. The overexpression of AtABHD11 in Escherichia coli led to a reduction in phospholipid levels. The bacterially expressed recombinant AtABHD11 hydrolyzed lyso(phospho)lipid and monoacylglycerol. Furthermore, using whole-genome microarray and real-time PCR analyses of abhd11 and wild-type plants, we noted the up-regulation of MGD1, -2, and -3 and DGD1. Together, these findings suggested that AtABHD11 is a lyso(phospho)lipase. The disruption of AtABHD11 caused the accumulation of the polar lipids in leaves, which in turn promoted a higher growth rate compared with wild-type plants. PMID:26589672

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

    PubMed

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

    2015-07-01

    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. Taken together, these results indicate that GL2 negatively regulates anthocyanin biosynthesis in Arabidopsis by directly repressing the expression of some MBW component genes. PMID:26017690

  16. Cold acclimation of the Arabidopsis dgd1 mutant results in recovery from photosystem I-limited photosynthesis.

    PubMed

    Hendrickson, Luke; Vlcková, Alexandra; Selstam, Eva; Huner, Norman; Oquist, Gunnar; Hurry, Vaughan

    2006-09-01

    We compared the thylakoid membrane composition and photosynthetic properties of non- and cold-acclimated leaves from the dgd1 mutant (lacking >90% of digalactosyl-diacylglycerol; DGDG) and wild type (WT) Arabidopsis thaliana. In contrast to warm grown plants, cold-acclimated dgd1 leaves recovered pigment-protein pools and photosynthetic function equivalent to WT. Surprisingly, this recovery was not correlated with an increase in DGDG. When returned to warm temperatures the severe dgd1 mutant phenotype reappeared. We conclude that the relative recovery of photosynthetic activity at 5 degrees C resulted from a temperature/lipid interaction enabling the stable assembly of PSI complexes in the thylakoid. PMID:16930596

  17. 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

  18. Classification and identification of Arabidopsis cell wall mutants using Fourier-Transform InfraRed (FT-IR) microspectroscopy.

    PubMed

    Mouille, Grégory; Robin, Stéphane; Lecomte, Mannaïg; Pagant, Silvère; Höfte, Herman

    2003-08-01

    We have developed a novel procedure for the rapid classification and identification of Arabidopsis mutants with altered cell wall architecture based on Fourier-Transform Infrared (FT-IR) microspectroscopy. FT-IR transmission spectra were sampled from native 4-day-old dark-grown hypocotyls of 46 mutants and the wild type treated with various drugs. The Mahalanobis distance between mutants, calculated from the spectral information after compression with the Discriminant Variables Selection procedure, was used for alpha hierarchical cluster analysis. Despite the completely unsupervised nature of the classification procedure, we show that all mutants with cellulose defects appeared in the same cluster. In addition, mutant alleles of similar strength for several unrelated loci were also clustered, which demonstrates the sensitivity of the method to detect a wide array of cell wall defects. Comparing the cellulose-deficient cluster with the cluster that contained wild-type controls led to the identification of wave numbers that were diagnostic for altered cellulose content in the context of an intact cell wall. The results show that FT-IR spectra can be used to identify different classes of mutants and to characterize cell wall changes at a microscopic level in unknown mutants. This procedure significantly accelerates the identification and classification of cell wall mutants, which makes cell wall polysaccharides more accessible to functional genomics approaches. PMID:12887590

  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. 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.

  1. 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.

  2. Structure of cellulose-deficient secondary cell walls from the irx3 mutant of Arabidopsis thaliana.

    PubMed

    Ha, Marie-Ann; MacKinnon, Iain M; Sturcová, Adriana; Apperley, David C; McCann, Maureen C; Turner, Simon R; Jarvis, Michael C

    2002-09-01

    In the Arabidopsis mutant irx3, truncation of the AtCesA7 gene encoding a xylem-specific cellulose synthase results in reduced cellulose synthesis in the affected xylem cells and collapse of mature xylem vessels. Here we describe spectroscopic experiments to determine whether any cellulose, normal or abnormal, remained in the walls of these cells and whether there were consequent effects on other cell-wall polysaccharides. Xylem cell walls from irx3 and its wild-type were prepared by anatomically specific isolation and were examined by solid-state NMR spectroscopy and FTIR microscopy. The affected cell walls of irx3 contained low levels of crystalline cellulose, probably associated with primary cell walls. There was no evidence that crystalline cellulose was replaced by less ordered glucans. From the molecular mobility of xylans and lignin it was deduced that these non-cellulosic polymers were cross-linked together in both irx3 and the wild-type. The disorder previously observed in the spatial pattern of non-cellulosic polymer deposition in the secondary walls of irx3 xylem could not be explained by any alteration in the structure or cross-linking of these polymers and may be attributed directly to the absence of cellulose microfibrils which, in the wild-type, scaffold the organisation of the other polymers into a coherent secondary cell wall. PMID:12165296

  3. 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.

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

    PubMed

    Rutherford, R; Masson, P H

    1996-08-01

    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. PMID:8756492

  5. Disturbed excitation energy transfer in Arabidopsis thaliana mutants lacking minor antenna complexes of photosystem II.

    PubMed

    Dall'Osto, Luca; Ünlü, Caner; Cazzaniga, Stefano; van Amerongen, Herbert

    2014-12-01

    Minor light-harvesting complexes (Lhcs) CP24, CP26 and CP29 occupy a position in photosystem II (PSII c' plants between the major light-harvesting complexes LHCII and the PSII core subunits. Lack of minor Lhcs in vivo causes impairment of PSII organization, and negatively affects electron transport rates anc photoprotection capacity. Here we used picosecond-fluorescence spectroscopy to study excitation-energy transfer (EET) in thylakoid membranes isolated from Arabidopsis thaliana wild-type plants and knockout lines depleted of either two (koCP26/24 and koCP29/24) or all minor Lhcs (NoM). In the absence of all minor Lhcs. the functional connection ofLHCII to the PSII cores appears to be seriously impaired whereas the "disconnected" LHCII is substantially quenched. For both double knock-out mutants, excitation trapping in PSII is faster than in NoM thylakoids but slower than in WT thylakoids. In NoM thylakoids, the loss of all minor Lhcs is accompanied by an over-accumulation ofLHCII, suggesting a compensating response to the reduced trapping efficiency in limiting light, which leads to a photosynthetic phenotype resembling that of low-light-acclimated plants. Finally. fluorescence kinetics and biochemical results show that the missing minor complexes are not replaced by other Lhcs, implying that they are unique among the antenna subunits and crucial for the functioning and macroorganization of PSII. PMID:25291424

  6. L-ascorbic acid metabolism in the ascorbate-deficient arabidopsis mutant vtc1.

    PubMed Central

    Conklin, P L; Pallanca, J E; Last, R L; Smirnoff, N

    1997-01-01

    The biosynthesis of L-ascorbic acid (vitamin C) is not well understood in plants. The ozone-sensitive Arabidopsis thaliana mutant vitamin c-1 (vtc1; formerly known as soz1) is deficient in ascorbic acid, accumulating approximately 30% of wild-type levels. This deficiency could result from elevated catabolism or decreased biosynthesis. No differences that could account for the deficiency were found in the activities of enzymes that catalyze the oxidation or reduction of ascorbic acid. The absolute rate of ascorbic acid turnover is actually less in vtc1 than in wild type; however, the turnover rate relative to the pool of ascorbic acid is not significantly different. The results from [U-14C]Glc labeling experiments suggest that the deficiency is the result of a biosynthetic defect: less L-[14C]ascorbic acid as a percentage of total soluble 14C accumulates in vtc1 than in wild type. The feeding of two putative biosynthetic intermediates, D-glucosone and L-sorbosone, had no positive effect on ascorbic acid levels in either genotype. The vtc1 defect does not appear to be the result of a deficiency in L-galactono-1,4-lactone dehydrogenase, an enzyme able to convert L-galactono-1,4-lactone to ascorbic acid. PMID:9390448

  7. 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. PMID:26965486

  8. 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.

  9. Utilization of mutants to analyze the interaction between Arabidopsis thaliana and its naturally root-associated Pseudomonas.

    PubMed

    Persello-Cartieaux, F; David, P; Sarrobert, C; Thibaud, M C; Achouak, W; Robaglia, C; Nussaume, L

    2001-01-01

    A model system based on the Arabidopsis thaliana (L.) Heynh. Ws ecotype and its naturally colonizing Pseudomonas thivervalensis rhizobacteria was defined. Pseudomonas strains colonizing A. thaliana were found to modify the root architecture either in vivo or in vitro. A gnotobiotic system using bacteria labelled with green fluorescent protein revealed that P. thivervalensis exhibited a colonization profile similar to that of other rhizobacterial species. Mutants of A. thaliana affected in root hair development and possible hormone perception were used to analyze the plant genetic determinants of bacterial colonization. A screen for mutants insensitive to P. thivervalensis colonization yielded two mutants found to be auxin resistant. This further supports a proposed role for bacterial auxin in inducing morphological modifications of roots. This work paves the way for studying the interaction between plants and non-pathogenic rhizobacteria in a gnotobiotic system, derived from a natural association, where interactions between both partners can be genetically dissected. PMID:11216839

  10. The Arabidopsis cax1 Mutant Exhibits Impaired Ion Homeostasis, Development, and Hormonal Responses and Reveals Interplay among Vacuolar Transporters

    PubMed Central

    Cheng, Ning-Hui; Pittman, Jon K.; Barkla, Bronwyn J.; Shigaki, Toshiro; Hirschi, Kendal D.

    2003-01-01

    The Arabidopsis Ca2+/H+ transporter CAX1 (Cation Exchanger1) may be an important regulator of intracellular Ca2+ levels. Here, we describe the preliminary localization of CAX1 to the tonoplast and the molecular and biochemical characterization of cax1 mutants. We show that these mutants exhibit a 50% reduction in tonoplast Ca2+/H+ antiport activity, a 40% reduction in tonoplast V-type H+-translocating ATPase activity, a 36% increase in tonoplast Ca2+-ATPase activity, and increased expression of the putative vacuolar Ca2+/H+ antiporters CAX3 and CAX4. Enhanced growth was displayed by the cax1 lines under Mn2+ and Mg2+ stress conditions. The mutants exhibited altered plant development, perturbed hormone sensitivities, and altered expression of an auxin-regulated promoter-reporter gene fusion. We propose that CAX1 regulates myriad plant processes and discuss the observed phenotypes with regard to the compensatory alterations in other transporters. PMID:12566577

  11. 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

  12. AUXIN RESPONSE FACTOR7 Restores the Expression of Auxin-Responsive Genes in Mutant Arabidopsis Leaf Mesophyll ProtoplastsW⃞

    PubMed Central

    Wang, Shucai; Tiwari, Shiv B.; Hagen, Gretchen; Guilfoyle, Tom J.

    2005-01-01

    AUXIN RESPONSE FACTOR7 (ARF7) is one of five ARF transcriptional activators in Arabidopsis thaliana that is proposed to regulate auxin-responsive expression of genes containing TGTCTC auxin response elements in their promoters. An Arabidopsis mutant (nonphototropic hypocotyl4-1 [nph4-1]) that is a null for ARF7 showed strongly reduced expression of integrated auxin-responsive reporter genes and natural genes that were monitored in Arabidopsis leaf mesophyll protoplasts. Expression of the reporter and natural genes was restored in an auxin-dependent manner when protoplasts were transfected with a 35S:ARF7 effector gene, encoding a full-length ARF7 protein. Transfection of effector genes encoding other ARF activators restored auxin-responsive gene expression to varying degrees, but less than that observed with the ARF7 effector gene. Arabidopsis lines that were null for ARF6, ARF8, or ARF19 were not defective in expression of the reporter and natural auxin response genes assayed in mesophyll protoplasts, suggesting that ARF7 plays a major role in regulating expression of a subset of auxin response genes in leaf mesophyll cells. Auxin-responsive gene expression was induced in wild-type protoplasts and restored in nph4-1 protoplasts only with auxin and not with other hormones, including brassinolide. In the presence of auxin, however, brassinolide modestly enhanced auxin-responsive gene expression. PMID:15923351

  13. 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.

  14. Altered chloroplast structure and function in a mutant of Arabidopsis deficient in plastid glycerol-3-phosphate acyltransferase activity

    SciTech Connect

    Kunst, L.; Somerville, C. ); Browse, J. )

    1989-07-01

    Mutants of Arabidopsis thaliana deficient in plastid glycerol-3-phosphate acyltransferase activity have altered chloroplast membrane lipid composition. This caused an increase in the number of regions of appressed membrane per chloroplast and a decrease in the average number of thylakoid membranes in the appressed regions. The net effect was a significant decrease in the ratio of appressed to nonappressed membranes. A comparison of 77 K fluorescence emission spectra of thylakoid membranes from the mutant and wild type indicated that the ultrastructural changes were associated with an altered distribution of excitation energy transfer from antenna chlorophyll to photosystem II and photosystem I in the mutant. The changes in leaf lipid composition did not significantly affect growth or development of the mutant under standard conditions. However, at temperatures above 28{degree}C the mutant grew slightly more rapidly than the wild type, and measurements of temperature-induced fluorescence yield enhancement suggested an increased thermal stability of the photosynthetic apparatus of the mutant. These effects are consistent with other evidence suggesting that membrane lipid composition is an important determinant of chloroplast structure but has relatively minor direct effects on the function of the membrane proteins associated with photosynthetic electron transport.

  15. Cell-wall structure and anisotropy in procuste, a cellulose synthase mutant of Arabidopsis thaliana.

    PubMed

    MacKinnon, Iain M; Sturcová, Adriana; Sugimoto-Shirasu, Keiko; His, Isabelle; McCann, Maureen C; Jarvis, Michael C

    2006-07-01

    In dark-grown hypocotyls of the Arabidopsis procuste mutant, a mutation in the CesA6 gene encoding a cellulose synthase reduces cellulose synthesis and severely inhibits elongation growth. Previous studies had left it uncertain why growth was inhibited, because cellulose synthesis was affected before, not during, the main phase of elongation. We characterised the quantity, structure and orientation of the cellulose remaining in the walls of affected cells. Solid-state NMR spectroscopy and infrared microscopy showed that the residual cellulose did not differ in structure from that of the wild type, but the cellulose content of the prc-1 cell walls was reduced by 28%. The total mass of cell-wall polymers per hypocotyl was reduced in prc-1 by about 20%. Therefore, the fourfold inhibition of elongation growth in prc-1 does not result from aberrant cellulose structure, nor from uniform reduction in the dimensions of the cell-wall network due to reduced cellulose or cell-wall mass. Cellulose orientation was quantified by two quantitative methods. First, the orientation of newly synthesised microfibrils was measured in field-emission scanning electron micrographs of the cytoplasmic face of the inner epidermal cell wall. The ordered transverse orientation of microfibrils at the inner face of the cell wall was severely disrupted in prc-1 hypocotyls, particularly in the early growth phase. Second, cellulose orientation distributions across the whole cell-wall thickness, measured by polarised infrared microscopy, were much broader. Analysis of the microfibril orientations according to the theory of composite materials showed that during the initial growth phase, their anisotropy at the plasma membrane was sufficient to explain the anisotropy of subsequent growth. PMID:16404578

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

    PubMed

    Afitlhile, Meshack; Workman, Samantha; Duffield, Kayla; Sprout, Danielle; Berhow, Mark

    2013-12-01

    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 fatty acid composition and transcript levels of plastid-localized fatty acid desaturases in the wild type and ppi2 mutant. The objective was to evaluate whether the Toc159 receptor was critical in the import of lipid-synthesizing enzymes. The ppi2 mutant accumulated decreased levels of oleic acid (18:1) and α-linolenic acid (18:3). The mutant accumulated drastically reduced amounts of the chloroplast lipid monogalactosyldiacylglycerol (MGDG), which contains more than 80% of 18:3. The expression of genes that encode stearoyl-ACP desaturase and MGD1 synthase were down-regulated in the ppi2 mutant, and this corresponded to decreased levels of 18:1 and MGDG, respectively. We conclude that in the ppi2 mutant the impaired synthesis of MGDG resulted in decreased amounts of 18:3. The mutant however, had a 30-fold increase in fad5 transcript levels; this increase was mirrored by a 16- to 50-fold accumulation of hexadecatrienoic acid (16:3), a fatty acid found exclusively in MGDG. Taken together, these data suggest that the Toc159 receptor is required in the import of stearoyl-ACP desaturase and MGD1 synthase into the chloroplasts. Since the expression of fad5 gene was up-regulated in the ppi2 mutant, we propose that fad5 desaturase is imported into plastids through the atToc132/atToc120 protein import pathway. PMID:24184455

  17. 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. PMID:27217494

  18. Redirection of the phenylpropanoid pathway to feruloyl malate in Arabidopsis mutants deficient for cinnamoyl-CoA reductase 1.

    PubMed

    Mir Derikvand, Mohammad; Sierra, Jimmy Berrio; Ruel, Katia; Pollet, Brigitte; Do, Cao-Trung; Thévenin, Johanne; Buffard, Dominique; Jouanin, Lise; Lapierre, Catherine

    2008-04-01

    Cinnamoyl-CoA reductase 1 (CCR1, gene At1g15950) is the main CCR isoform implied in the constitutive lignification of Arabidopsis thaliana. In this work, we have identified and characterized two new knockout mutants for CCR1. Both have a dwarf phenotype and a delayed senescence. At complete maturity, their inflorescence stems display a 25-35% decreased lignin level, some alterations in lignin structure with a higher frequency of resistant interunit bonds and a higher content in cell wall-bound ferulic esters. Ferulic acid-coniferyl alcohol ether dimers were found for the first time in dicot cell walls and in similar levels in wild-type and mutant plants. The expression of CCR2, a CCR gene usually involved in plant defense, was increased in the mutants and could account for the biosynthesis of lignins in the CCR1-knockout plants. Mutant plantlets have three to four-times less sinapoyl malate (SM) than controls and accumulate some feruloyl malate. The same compositional changes occurred in the rosette leaves of greenhouse-grown plants. By contrast and relative to the control, their stems accumulated unusually high levels of both SM and feruloyl malate as well as more kaempferol glycosides. These findings suggest that, in their hypolignified stems, the mutant plants would avoid the feruloyl-CoA accumulation by its redirection to cell wall-bound ferulate esters, to feruloyl malate and to SM. The formation of feruloyl malate to an extent far exceeding the levels reported so far indicates that ferulic acid is a potential substrate for the enzymes involved in SM biosynthesis and emphasizes the remarkable plasticity of Arabidopsis phenylpropanoid metabolism. PMID:18046574

  19. Systematic Phenotypic Screen of Arabidopsis Peroxisomal Mutants Identifies Proteins Involved in β-Oxidation1[W][OPEN

    PubMed Central

    Cassin-Ross, Gaëlle; Hu, Jianping

    2014-01-01

    Peroxisomes are highly dynamic and multifunctional organelles essential to development. Plant peroxisomes accommodate a multitude of metabolic reactions, many of which are related to the β-oxidation of fatty acids or fatty acid-related metabolites. Recently, several dozens of novel peroxisomal proteins have been identified from Arabidopsis (Arabidopsis thaliana) through in silico and experimental proteomic analyses followed by in vivo protein targeting validations. To determine the functions of these proteins, we interrogated their transfer DNA insertion mutants with a series of physiological, cytological, and biochemical assays to reveal peroxisomal deficiencies. Sugar dependence and 2,4-dichlorophenoxybutyric acid and 12-oxo-phytodienoic acid response assays uncovered statistically significant phenotypes in β-oxidation-related processes in mutants for 20 of 27 genes tested. Additional investigations uncovered a subset of these mutants with abnormal seed germination, accumulation of oil bodies, and delayed degradation of long-chain fatty acids during early seedling development. Mutants for seven genes exhibited deficiencies in multiple assays, strongly suggesting the involvement of their gene products in peroxisomal β-oxidation and initial seedling growth. Proteins identified included isoforms of enzymes related to β-oxidation, such as acyl-CoA thioesterase2, acyl-activating enzyme isoform1, and acyl-activating enzyme isoform5, and proteins with functions previously unknown to be associated with β-oxidation, such as Indigoidine synthase A, Senescence-associated protein/B12D-related protein1, Betaine aldehyde dehydrogenase, and Unknown protein5. This multipronged phenotypic screen allowed us to reveal β-oxidation proteins that have not been discovered by single assay-based mutant screens and enabled the functional dissection of different isoforms of multigene families involved in β-oxidation. PMID:25253886

  20. Overaccumulation of γ-Glutamylcysteine in a Jasmonate-Hypersensitive Arabidopsis Mutant Causes Jasmonate-Dependent Growth Inhibition1[OPEN

    PubMed Central

    Wei, Hsin-Ho; Rowe, Martha; Riethoven, Jean-Jack M.; Grove, Ryan; Adamec, Jiri; Jikumaru, Yusuke; Staswick, Paul

    2015-01-01

    Glutathione (GSH) is essential for many aspects of plant biology and is associated with jasmonate signaling in stress responses. We characterized an Arabidopsis (Arabidopsis thaliana) jasmonate-hypersensitive mutant (jah2) with seedling root growth 100-fold more sensitive to inhibition by the hormone jasmonyl-isoleucine than the wild type. Genetic mapping and genome sequencing determined that the mutation is in intron 6 of GLUTATHIONE SYNTHETASE2, encoding the enzyme that converts γ-glutamylcysteine (γ-EC) to GSH. The level of GSH in jah2 was 71% of the wild type, while the phytoalexin-deficient2-1 (pad2-1) mutant, defective in GSH1 and having only 27% of wild-type GSH level, was not jasmonate hypersensitive. Growth defects for jah2, but not pad2, were also seen in plants grown to maturity. Surprisingly, all phenotypes in the jah2 pad2-1 double mutant were weaker than in jah2. Quantification of γ-EC indicated these defects result from hyperaccumulation of this GSH precursor by 294- and 65-fold in jah2 and the double mutant, respectively. γ-EC reportedly partially substitutes for loss of GSH, but growth inhibition seen here was likely not due to an excess of total glutathione plus γ-EC because their sum in jah2 pad2-1 was only 16% greater than in the wild type. Further, the jah2 phenotypes were lost in a jasmonic acid biosynthesis mutant background, indicating the effect of γ-EC is mediated through jasmonate signaling and not as a direct result of perturbed redox status. PMID:26282239

  1. Overaccumulation of γ-Glutamylcysteine in a Jasmonate-Hypersensitive Arabidopsis Mutant Causes Jasmonate-Dependent Growth Inhibition.

    PubMed

    Wei, Hsin-Ho; Rowe, Martha; Riethoven, Jean-Jack M; Grove, Ryan; Adamec, Jiri; Jikumaru, Yusuke; Staswick, Paul

    2015-10-01

    Glutathione (GSH) is essential for many aspects of plant biology and is associated with jasmonate signaling in stress responses. We characterized an Arabidopsis (Arabidopsis thaliana) jasmonate-hypersensitive mutant (jah2) with seedling root growth 100-fold more sensitive to inhibition by the hormone jasmonyl-isoleucine than the wild type. Genetic mapping and genome sequencing determined that the mutation is in intron 6 of GLUTATHIONE SYNTHETASE2, encoding the enzyme that converts γ-glutamylcysteine (γ-EC) to GSH. The level of GSH in jah2 was 71% of the wild type, while the phytoalexin-deficient2-1 (pad2-1) mutant, defective in GSH1 and having only 27% of wild-type GSH level, was not jasmonate hypersensitive. Growth defects for jah2, but not pad2, were also seen in plants grown to maturity. Surprisingly, all phenotypes in the jah2 pad2-1 double mutant were weaker than in jah2. Quantification of γ-EC indicated these defects result from hyperaccumulation of this GSH precursor by 294- and 65-fold in jah2 and the double mutant, respectively. γ-EC reportedly partially substitutes for loss of GSH, but growth inhibition seen here was likely not due to an excess of total glutathione plus γ-EC because their sum in jah2 pad2-1 was only 16% greater than in the wild type. Further, the jah2 phenotypes were lost in a jasmonic acid biosynthesis mutant background, indicating the effect of γ-EC is mediated through jasmonate signaling and not as a direct result of perturbed redox status. PMID:26282239

  2. 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

  3. A New Vertical Mesh Transfer Technique for Metal-Tolerance Studies in Arabidopsis (Ecotypic Variation and Copper-Sensitive Mutants).

    PubMed

    Murphy, A.; Taiz, L.

    1995-05-01

    A new vertical mesh transfer (VMT) technique has been developed to facilitate the rapid isolation of plant metal-tolerance mutants. The technique is quantitative, allowing comparisons of the growth responses of different strains or ecotypes. Using the VMT technique, we have characterized the dose responses of 10 ecotypes of Arabidopsis thaliana to Cu2+, Zn2+, Ni2+, Cr3+, Cd2+, and Al3+. Ecotypic variations in the highest concentration causing no inhibition and the lowest concentration causing complete inhibition for the six metals were observed. Two ecotypes, Ws and Enkheim, exhibited an inducible tolerance mechanism in response to copper. Pretreatment of Ws with the highest concentration causing no inhibition for copper resulted in a shifting of the lowest concentration causing complete inhibition to a higher value. Partial cross-induction and cross-tolerance between Cu2+ and Zn2+ were demonstrated. In addition, ethyl methanesulfonate-mutagenized Columbia seeds were screened for copper-sensitive (cus) mutants using the VMT procedure. Thus far, 59 putative cus mutants have survived retesting to the M4 or M5 generation. When grown on gellan gum supplemented with 30 [mu]M CuCl2, cus mutants develop marked toxicity symptoms. A copper dose-response curve of the cus1 mutant showed that the metal-sensitive phenotype is specific for the lower concentration range. PMID:12228451

  4. 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. PMID:25652756

  5. 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

  6. Analysis of leaf development in fugu mutants of Arabidopsis reveals three compensation modes that modulate cell expansion in determinate organs.

    PubMed

    Ferjani, Ali; Horiguchi, Gorou; Yano, Satoshi; Tsukaya, Hirokazu

    2007-06-01

    In multicellular organisms, the coordination of cell proliferation and expansion is fundamental for proper organogenesis, yet the molecular mechanisms involved in this coordination are largely unexplored. In plant leaves, the existence of this coordination is suggested by compensation, in which a decrease in cell number triggers an increase in mature cell size. To elucidate the mechanisms of compensation, we isolated five new Arabidopsis (Arabidopsis thaliana) mutants (fugu1-fugu5) that exhibit compensation. These mutants were characterized together with angustifolia3 (an3), erecta (er), and a KIP-RELATED PROTEIN2 (KRP2) overexpressor, which were previously reported to exhibit compensation. Time-course analyses of leaf development revealed that enhanced cell expansion in fugu2-1, fugu5-1, an3-4, and er-102 mutants is induced postmitotically, indicating that cell enlargement is not caused by the uncoupling of cell division from cell growth. In each of the mutants, either the rate or duration of cell expansion was selectively enhanced. In contrast, we found that enhanced cell expansion in KRP2 overexpressor occurs during cell proliferation. We further demonstrated that enhanced cell expansion occurs in cotyledons with dynamics similar to that in leaves. In contrast, cell expansion was not enhanced in roots even though they exhibit decreased cell numbers. Thus, compensation was confirmed to occur preferentially in determinate organs. Flow cytometric analyses revealed that increases in ploidy level are not always required to trigger compensation, suggesting that compensation is only partially mediated by ploidy-dependent processes. Our results suggest that compensation reflects an organ-wide coordination of cell proliferation and expansion in determinate organs, and involves at least three different expansion pathways. PMID:17468216

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

    PubMed Central

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

    2016-01-01

    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 Pst DC3000 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. PMID:26896849

  8. 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

  9. Photosynthetic redox imbalance governs leaf sectoring in the Arabidopsis thaliana variegation mutants immutans, spotty, var1, and var2.

    PubMed

    Rosso, Dominic; Bode, Rainer; Li, Wenze; Krol, Marianna; Saccon, Diego; Wang, Shelly; Schillaci, Lori A; Rodermel, Steven R; Maxwell, Denis P; Hüner, Norman P A

    2009-11-01

    We hypothesized that chloroplast energy imbalance sensed through alterations in the redox state of the photosynthetic electron transport chain, measured as excitation pressure, governs the extent of variegation in the immutans mutant of Arabidopsis thaliana. To test this hypothesis, we developed a nondestructive imaging technique and used it to quantify the extent of variegation in vivo as a function of growth temperature and irradiance. The extent of variegation was positively correlated (R(2) = 0.750) with an increase in excitation pressure irrespective of whether high light, low temperature, or continuous illumination was used to induce increased excitation pressure. Similar trends were observed with the variegated mutants spotty, var1, and var2. Measurements of greening of etiolated wild-type and immutans cotyledons indicated that the absence of IMMUTANS increased excitation pressure twofold during the first 6 to 12 h of greening, which led to impaired biogenesis of thylakoid membranes. In contrast with IMMUTANS, the expression of its mitochondrial analog, AOX1a, was transiently upregulated in the wild type but permanently upregulated in immutans, indicating that the effects of excitation pressure during greening were also detectable in mitochondria. We conclude that mutations involving components of the photosynthetic electron transport chain, such as those present in immutans, spotty, var1, and var2, predispose Arabidopsis chloroplasts to photooxidation under high excitation pressure, resulting in the variegated phenotype. PMID:19897671

  10. Photosynthetic Redox Imbalance Governs Leaf Sectoring in the Arabidopsis thaliana Variegation Mutants immutans, spotty, var1, and var2[W

    PubMed Central

    Rosso, Dominic; Bode, Rainer; Li, Wenze; Krol, Marianna; Saccon, Diego; Wang, Shelly; Schillaci, Lori A.; Rodermel, Steven R.; Maxwell, Denis P.; Hüner, Norman P.A.

    2009-01-01

    We hypothesized that chloroplast energy imbalance sensed through alterations in the redox state of the photosynthetic electron transport chain, measured as excitation pressure, governs the extent of variegation in the immutans mutant of Arabidopsis thaliana. To test this hypothesis, we developed a nondestructive imaging technique and used it to quantify the extent of variegation in vivo as a function of growth temperature and irradiance. The extent of variegation was positively correlated (R2 = 0.750) with an increase in excitation pressure irrespective of whether high light, low temperature, or continuous illumination was used to induce increased excitation pressure. Similar trends were observed with the variegated mutants spotty, var1, and var2. Measurements of greening of etiolated wild-type and immutans cotyledons indicated that the absence of IMMUTANS increased excitation pressure twofold during the first 6 to 12 h of greening, which led to impaired biogenesis of thylakoid membranes. In contrast with IMMUTANS, the expression of its mitochondrial analog, AOX1a, was transiently upregulated in the wild type but permanently upregulated in immutans, indicating that the effects of excitation pressure during greening were also detectable in mitochondria. We conclude that mutations involving components of the photosynthetic electron transport chain, such as those present in immutans, spotty, var1, and var2, predispose Arabidopsis chloroplasts to photooxidation under high excitation pressure, resulting in the variegated phenotype. PMID:19897671

  11. ß-amylase1 mutant Arabidopsis plants show improved drought tolerance due to reduced starch breakdown in guard cells

    PubMed Central

    Prasch, Christian Maximilian; Ott, Kirsten Verena; Bauer, Hubert; Ache, Peter; Hedrich, Rainer; Sonnewald, Uwe

    2015-01-01

    In plants, drought stress is a major growth limiting factor causing cell water loss through open stomata. In this study, guard cell-specific transcripts from drought-stressed Arabidopsis plants were analysed and a down-regulation of β-amylase 1 (BAM1) was found. In previous studies, BAM1 was shown to be involved in stomatal starch degradation under ambient conditions. Impaired starch breakdown of bam1 mutant plants was accompanied by decreased stomatal opening. Here, it is shown that drought tolerance of bam1 mutant plants is improved as compared with wild-type controls. Microarray analysis of stomata-specific transcripts from bam1 mutant plants revealed a significant down-regulation of genes encoding aquaporins, auxin- and ethylene-responsive factors, and cell-wall modifying enzymes. This expression pattern suggests that reduced water uptake and limited cell wall extension are associated with the closed state of stomata of bam1 mutant plants. Together these data suggest that regulation of stomata-specific starch turnover is important for adapting stomata opening to environmental needs and its breeding manipulation may result in drought tolerant crop plants. PMID:26139825

  12. ß-amylase1 mutant Arabidopsis plants show improved drought tolerance due to reduced starch breakdown in guard cells.

    PubMed

    Prasch, Christian Maximilian; Ott, Kirsten Verena; Bauer, Hubert; Ache, Peter; Hedrich, Rainer; Sonnewald, Uwe

    2015-09-01

    In plants, drought stress is a major growth limiting factor causing cell water loss through open stomata. In this study, guard cell-specific transcripts from drought-stressed Arabidopsis plants were analysed and a down-regulation of β-amylase 1 (BAM1) was found. In previous studies, BAM1 was shown to be involved in stomatal starch degradation under ambient conditions. Impaired starch breakdown of bam1 mutant plants was accompanied by decreased stomatal opening. Here, it is shown that drought tolerance of bam1 mutant plants is improved as compared with wild-type controls. Microarray analysis of stomata-specific transcripts from bam1 mutant plants revealed a significant down-regulation of genes encoding aquaporins, auxin- and ethylene-responsive factors, and cell-wall modifying enzymes. This expression pattern suggests that reduced water uptake and limited cell wall extension are associated with the closed state of stomata of bam1 mutant plants. Together these data suggest that regulation of stomata-specific starch turnover is important for adapting stomata opening to environmental needs and its breeding manipulation may result in drought tolerant crop plants. PMID:26139825

  13. 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

  14. 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.

  15. Identification of Coilin Mutants in a Screen for Enhanced Expression of an Alternatively Spliced GFP Reporter Gene in Arabidopsis thaliana

    PubMed Central

    Kanno, Tatsuo; Lin, Wen-Dar; Fu, Jason L.; Wu, Ming-Tsung; Yang, Ho-Wen; Lin, Shih-Shun; Matzke, Antonius J. M.; Matzke, Marjori

    2016-01-01

    Coilin is a marker protein for subnuclear organelles known as Cajal bodies, which are sites of various RNA metabolic processes including the biogenesis of spliceosomal small nuclear ribonucleoprotein particles. Through self-associations and interactions with other proteins and RNA, coilin provides a structural scaffold for Cajal body formation. However, despite a conspicuous presence in Cajal bodies, most coilin is dispersed in the nucleoplasm and expressed in cell types that lack these organelles. The molecular function of coilin, particularly of the substantial nucleoplasmic fraction, remains uncertain. We identified coilin loss-of-function mutations in a genetic screen for mutants showing either reduced or enhanced expression of an alternatively spliced GFP reporter gene in Arabidopsis thaliana. The coilin mutants feature enhanced GFP fluorescence and diminished Cajal bodies compared with wild-type plants. The amount of GFP protein is several-fold higher in the coilin mutants owing to elevated GFP transcript levels and more efficient splicing to produce a translatable GFP mRNA. Genome-wide RNA-sequencing data from two distinct coilin mutants revealed a small, shared subset of differentially expressed genes, many encoding stress-related proteins, and, unexpectedly, a trend toward increased splicing efficiency. These results suggest that coilin attenuates splicing and modulates transcription of a select group of genes. The transcriptional and splicing changes observed in coilin mutants are not accompanied by gross phenotypic abnormalities or dramatically altered stress responses, supporting a role for coilin in fine tuning gene expression. Our GFP reporter gene provides a sensitive monitor of coilin activity that will facilitate further investigations into the functions of this enigmatic protein. PMID:27317682

  16. Identification of Coilin Mutants in a Screen for Enhanced Expression of an Alternatively Spliced GFP Reporter Gene in Arabidopsis thaliana.

    PubMed

    Kanno, Tatsuo; Lin, Wen-Dar; Fu, Jason L; Wu, Ming-Tsung; Yang, Ho-Wen; Lin, Shih-Shun; Matzke, Antonius J M; Matzke, Marjori

    2016-08-01

    Coilin is a marker protein for subnuclear organelles known as Cajal bodies, which are sites of various RNA metabolic processes including the biogenesis of spliceosomal small nuclear ribonucleoprotein particles. Through self-associations and interactions with other proteins and RNA, coilin provides a structural scaffold for Cajal body formation. However, despite a conspicuous presence in Cajal bodies, most coilin is dispersed in the nucleoplasm and expressed in cell types that lack these organelles. The molecular function of coilin, particularly of the substantial nucleoplasmic fraction, remains uncertain. We identified coilin loss-of-function mutations in a genetic screen for mutants showing either reduced or enhanced expression of an alternatively spliced GFP reporter gene in Arabidopsis thaliana The coilin mutants feature enhanced GFP fluorescence and diminished Cajal bodies compared with wild-type plants. The amount of GFP protein is several-fold higher in the coilin mutants owing to elevated GFP transcript levels and more efficient splicing to produce a translatable GFP mRNA. Genome-wide RNA-sequencing data from two distinct coilin mutants revealed a small, shared subset of differentially expressed genes, many encoding stress-related proteins, and, unexpectedly, a trend toward increased splicing efficiency. These results suggest that coilin attenuates splicing and modulates transcription of a select group of genes. The transcriptional and splicing changes observed in coilin mutants are not accompanied by gross phenotypic abnormalities or dramatically altered stress responses, supporting a role for coilin in fine tuning gene expression. Our GFP reporter gene provides a sensitive monitor of coilin activity that will facilitate further investigations into the functions of this enigmatic protein. PMID:27317682

  17. 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.

  18. A SAL1 Loss-of-Function Arabidopsis Mutant Exhibits Enhanced Cadmium Tolerance in Association with Alleviation of Endoplasmic Reticulum Stress.

    PubMed

    Xi, Hongmei; Xu, Hua; Xu, Wenxiu; He, Zhenyan; Xu, Wenzhong; Ma, Mi

    2016-06-01

    SAL1, as a negative regulator of stress response signaling, has been studied extensively for its role in plant response to environmental stresses. However, the role of SAL1 in cadmium (Cd) stress response and the underlying mechanism is still unclear. Using an Arabidopsis thaliana loss-of-function mutant of SAL1, we assessed Cd resistance and further explored the Cd toxicity mechanism through analysis of the endoplasmic reticulum (ER) stress response. The loss of SAL1 function greatly improved Cd tolerance and significantly attenuated ER stress in Arabidopsis. Exposure to Cd induced an ER stress response in Arabidopsis as evidenced by unconventional splicing of AtbZIP60 and up-regulation of ER stress-responsive genes. Damage caused by Cd was markedly reduced in the ER stress response double mutant bzip28 bzip60 or by application of the ER stress-alleviating chemical agents, tauroursodeoxycholic acid (TUDCA) and 4-phenyl butyric acid (4-PBA), in wild-type plants. The Cd-induced ER stress in Arabidopsis was also alleviated by loss of function of SAL1. These results identified SAL1 as a new component mediating Cd toxicity and established the role of the ER stress response in Cd toxicity. Additionally, the attenuated ER stress in the sal1 mutant might also shed new light on the mechanism of diverse abiotic stress resistance in the SAL1 loss-of-function mutants. PMID:27044671

  19. 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…

  20. A putative soybean GmsSOS1 confers enhanced salt tolerance to transgenic Arabidopsis sos1-1 mutant.

    PubMed

    Nie, Wang-Xing; Xu, Lin; Yu, Bing-Jun

    2015-01-01

    The cDNA of GmsSOS1, a putative plasma membrane Na(+)/H(+) antiporter gene isolated from Glycine max, Glycine soja, and their hybrid, was constructed into plant expression vector pCAMBIA 1300 and then transformed with Agrobacterium tumefaciens under the control of CaMV 35S promoter to Arabidopsis thaliana wild-type (WT) and mutant (atsos1-1) plants. By hygromycin resistance detection and PCR analysis, transgenic plants (WT35S:GmsSOS1 and atsos1-1 35S:GmsSOS1) were obtained. Seed germination, seedling growth, and Na(+) contents in roots and shoots were analytically compared among WT, atsos1-1 mutant, and their transgenic lines under salt stress. The results showed that when GmsSOS1 was integrated into the genome of A. thaliana, the inhibitions of salt stress on seed germination and seedling growth were all significantly improved, and enhanced salt tolerance was displayed, which may be attributed to the decrease of Na(+) absorption in roots and transportation in shoots of the transgenic lines, especially for that of atsos1-1 mutant. PMID:24934653

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

    PubMed

    Schneider, Anja; Steinberger, Iris; Herdean, Andrei; Gandini, Chiara; Eisenhut, Marion; Kurz, Samantha; Morper, Anna; Hoecker, Natalie; Rühle, Thilo; Labs, Mathias; Flügge, Ulf-Ingo; Geimer, Stefan; Schmidt, Sidsel Birkelund; Husted, Søren; Weber, Andreas P M; Spetea, Cornelia; Leister, Dario

    2016-04-01

    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 Mn(2+) and Ca(2+) 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, Mn(2+) and Ca(2+) ions were differently sequestered in pam71, with Ca(2+) enriched in pam71 thylakoids relative to the wild type. The changes in Ca(2+) 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 Mn(2+), but not Ca(2+) Furthermore, PAM71 suppressed the Mn(2+)-sensitive phenotype of the yeast mutant Δpmr1 Therefore, PAM71 presumably functions in Mn(2+) uptake into thylakoids to ensure optimal PSII performance. PMID:27020959

  2. Effects of reduced amounts of lipid unsaturation on chloroplast ultrastructure and photosynthesis in a mutant of Arabidopsis

    SciTech Connect

    McCourt, P.; Kunst, L.; Browse, J.; Somerville, C.R.

    1987-06-01

    A mutant of Arabidopsis thaliana with reduced content of C/sub 18:3/ and C/sub 16:3/ fatty acids in membrane lipids exhibited a 45% reduction in the cross-sectional area of chloroplasts and had a decrease of similar magnitude in the amount of chloroplast lamellar membranes. The reduction in chloroplast size was partially compensated by a 45% increase in the number of chloroplasts per cell in the mutant. When expressed on a chlorophyll basis the rates of CO/sub 2/-fixation and photosynthetic electron transport were not affected by these changes. Fluorescence polarization measurements indicated that the fluidity of the thylakoid membranes was not significantly altered by the mutation. Similarly, on the basis of temperature-induced fluorescence yield enhancement measurements, there was no significant effect on the thermal stability of chlorophyll-protein complexes in the mutant. These observations suggest that the high content of trienoic fatty acids in chloroplast lipids may be an important factor regulating organelle biogenesis but is not required to support normal levels of the photosynthetic activities associated with the thylakoid membranes.

  3. Flowering responses to altered expression of phytochrome in mutants and transgenic lines of Arabidopsis thaliana (L.) Heynh.

    PubMed Central

    Bagnall, D J; King, R W; Whitelam, G C; Boylan, M T; Wagner, D; Quail, P H

    1995-01-01

    The long-day plant Arabidopsis thaliana (L.) Heynh. flowers early in response to brief end-of-day (EOD) exposures to far-red light (FR) following a fluorescent short day of 8 h. FR promotion of flowering was nullified by subsequent brief red light (R) EOD exposure, indicating phytochrome involvement. The EOD response to R or FR is a robust measure of phytochrome action. Along with their wild-type (WT) parents, mutants deficient in either phytochrome A or B responded similarly to the EOD treatments. Thus, neither phytochrome A nor B exclusively regulated flowering, although phytochrome B controlled hypocotyl elongation. Perhaps a third phytochrome species is important for the EOD responses of the mutants and/or their flowering is regulated by the amount of the FR-absorbing form of phytochrome, irrespective of the phytochrome species. Overexpression of phytochrome A or phytochrome B resulted in differing photoperiod and EOD responses among the genotypes. The day-neutral overexpressor of phytochrome A had an EOD response similar to all of the mutants and WTs, whereas R EOD exposure promoted flowering in the overexpressor of phytochrome B and FR EOD exposure inhibited this promotion. The comparisons between relative flowering times and leaf numbers at flowering of the over-expressors and their WTs were not consistent across photoperiods and light treatments, although both phytochromes A and B contributed to regulating flowering of the transgenic plants. PMID:7659750

  4. Insights into lignin primary structure and deconstruction from Arabidopsis thaliana COMT (caffeic acid O-methyl transferase) mutant Atomt1.

    PubMed

    Moinuddin, Syed G A; Jourdes, Michaël; Laskar, Dhrubojyoti D; Ki, Chanyoung; Cardenas, Claudia L; Kim, Kye-Won; Zhang, Dianzhong; Davin, Laurence B; Lewis, Norman G

    2010-09-01

    The Arabidopsis mutant Atomt1 lignin differs from native lignin in wild type plants, in terms of sinapyl (S) alcohol-derived substructures in fiber cell walls being substituted by 5-hydroxyconiferyl alcohol (5OHG)-derived moieties. During programmed lignin assembly, these engender formation of benzodioxane substructures due to intramolecular cyclization of their quinone methides that are transiently formed following 8-O-4' radical-radical coupling. Thioacidolytic cleavage of the 8-O-4' inter-unit linkages in the Atomt1 mutant, relative to the wild type, indicated that cleavable sinapyl (S) and coniferyl (G) alcohol-derived monomeric moieties were stoichiometrically reduced by a circa 2 : 1 ratio. Additionally, lignin degradative analysis resulted in release of a 5OHG-5OHG-G trimer from the Atomt1 mutant, which then underwent further cleavage. Significantly, the trimeric moiety released provides new insight into lignin primary structure: during polymer assembly, the first 5OHG moiety is linked via a C8-O-X inter-unit linkage, whereas subsequent addition of monomers apparently involves sequential addition of 5OHG and G moieties to the growing chain in a 2 : 1 overall stoichiometry. This quantification data thus provides further insight into how inter-unit linkage frequencies in native lignins are apparently conserved (or near conserved) during assembly in both instances, as well as providing additional impetus to resolve how the overall question of lignin macromolecular assembly is controlled in terms of both type of monomer addition and primary sequence. PMID:20652169

  5. 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.

  6. An Arabidopsis Mutant with High Cyclic Electron Flow around Photosystem I (hcef) Involving the NADPH Dehydrogenase Complex[W][OA

    PubMed Central

    Livingston, Aaron K.; Cruz, Jeffrey A.; Kohzuma, Kaori; Dhingra, Amit; Kramer, David M.

    2010-01-01

    Cyclic electron flow (CEFI) has been proposed to balance the chloroplast energy budget, but the pathway, mechanism, and physiological role remain unclear. We isolated a new class of mutant in Arabidopsis thaliana, hcef for high CEF1, which shows constitutively elevated CEF1. The first of these, hcef1, was mapped to chloroplast fructose-1,6-bisphosphatase. Crossing hcef1 with pgr5, which is deficient in the antimycin A–sensitive pathway for plastoquinone reduction, resulted in a double mutant that maintained the high CEF1 phenotype, implying that the PGR5-dependent pathway is not involved. By contrast, crossing hcef1 with crr2-2, deficient in thylakoid NADPH dehydrogenase (NDH) complex, results in a double mutant that is highly light sensitive and lacks elevated CEF1, suggesting that NDH plays a direct role in catalyzing or regulating CEF1. Additionally, the NdhI component of the NDH complex was highly expressed in hcef1, whereas other photosynthetic complexes, as well as PGR5, decreased. We propose that (1) NDH is specifically upregulated in hcef1, allowing for increased CEF1; (2) the hcef1 mutation imposes an elevated ATP demand that may trigger CEF1; and (3) alternative mechanisms for augmenting ATP cannot compensate for the loss of CEF1 through NDH. PMID:20081115

  7. An Arabidopsis pex10 Null Mutant Is Embryo Lethal, Implicating Peroxisomes in an Essential Role during Plant Embryogenesis1

    PubMed Central

    Sparkes, Imogen A.; Brandizzi, Federica; Slocombe, Stephen P.; El-Shami, Mahmoud; Hawes, Chris; Baker, Alison

    2003-01-01

    Peroxisomes participate in many important functions in plants, including seed reserve mobilization, photorespiration, defense against oxidative stress, and auxin and jasmonate signaling. In mammals, defects in peroxisome biogenesis result in multiple system abnormalities, severe developmental delay, and death, whereas in unicellular yeasts, peroxisomes are dispensable unless required for growth of specific substrates. PEX10 encodes an integral membrane protein required for peroxisome biogenesis in mammals and yeast. To investigate the importance of PEX10 in plants, we characterized a Ds insertion mutant in the PEX10 gene of Arabidopsis (AtPEX10). Heterozygous AtPEX10::dissociation element mutants show normal vegetative phenotypes under optimal growth conditions, but produce about 20% abnormal seeds. The embryos in the abnormal seeds are predominantly homozygous for the disruption allele. They show retarded development and some morphological abnormalities. No viable homozygous mutant plants were obtained. AtPEX10 fused to yellow fluorescent protein colocalized with green fluorescent protein-serine-lysine-leucine, a well-documented peroxisomal marker, suggesting that AtPEX10 encodes a peroxisomal protein that is essential for normal embryo development and viability. PMID:14576288

  8. 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. PMID:26428915

  9. Dominant negative suppression of arabidopsis photoresponses by mutant phytochrome A sequences identifies spatially discrete regulatory domains in the photoreceptor.

    PubMed Central

    Boylan, M; Douglas, N; Quail, P H

    1994-01-01

    We used the exaggerated short hypocotyl phenotype induced by oat phytochrome A overexpression in transgenic Arabidopsis to monitor the biological activity of mutant phytochrome A derivatives. Three different mutations, which were generated by removing 52 amino acids from the N terminus (delta N52), the entire C-terminal domain (delta C617), or amino acids 617-686 (delta 617-686) of the oat molecule, each caused striking dominant negative interference with the ability of endogenous Arabidopsis phytochrome A to inhibit hypocotyl growth in continuous far-red light ("far-red high irradiance response" conditions). By contrast, in continuous white or red light, delta N52 was as active as the unmutagenized oat phytochrome A protein in suppressing hypocotyl elongation, while delta C617 and delta 617-686 continued to exhibit dominant negative behavior under these conditions. These data suggest that at least three spatially discrete molecular domains coordinate the photoregulatory activities of phytochrome A in Arabidopsis seedlings. The first is the chromophore-bearing N-terminal domain between residues 53 and 616 that is apparently sufficient for the light-induced initiation but not the completion of productive interactions with transduction chain components. The second is the C-terminal domain between residues 617 and 1129 that is apparently necessary for completion of productive interactions under all irradiation conditions. The third is the N-terminal 52 amino acids that are apparently necessary for completion of productive interactions only under far-red high irradiance conditions and are completely dispensable under white and red light regimes. PMID:8180501

  10. Agrobacterium mediated transfer of a mutant Arabidopsis acetolactate synthase gene confers resistance to chlorsulfuron in chicory (Cichorium intybus L.).

    PubMed

    Vermeulen, A; Vaucheret, H; Pautot, V; Chupeau, Y

    1992-06-01

    Leaf discs of C. intybus were inoculated with an Agrobacterium tumefaciens strain harboring a neomycin phosphotransferase (neo) gene for kanamycin resistance and a mutant acetolactate synthase gene (csr1-1) from Arabidopsis thaliana conferring resistance to sulfonylurea herbicides. A regeneration medium was optimized which permitted an efficient shoot regeneration from leaf discs. Transgenic shoots were selected on rooting medium containing 100 mg/l kanamycin sulfate. Integration of the csr1-1 gene into genomic DNA of kanamycin resistant chicory plants was confirmed by Southern blot hybridizations. Analysis of the selfed progenies (S1 and S2) of two independent transformed clones showed that kanamycin and chlorsulfuron resistances were inherited as dominant Mendelian traits. The method described here for producing transformed plants will allow new opportunities for chicory breeding. PMID:24203132

  11. Root hair formation at the root-hypocotyl junction in CPC-LIKE MYB double and triple mutants of Arabidopsis

    PubMed Central

    Wada, Takuji; Hayashi, Naoto; Tominaga-Wada, Rumi

    2015-01-01

    In Arabidopsis thaliana, R3-type MYB genes, CAPRICE (CPC) and its family of genes including TRIPTYCHON (TRY), ENHANCER OF TRY AND CPC1 (ETC1), ETC2 and CPC-LIKE MYB3 cooperatively regulate epidermal cell differentiation. Root hair formation is greatly reduced by a mutation in CPC, and try and etc1 enhance this phenotype. In this study, we demonstrate that CPC, TRY and ETC1 are also involved in root hair formation at the root-hypocotyl junction. The cpc try and cpc etc1 double mutants showed a reduced number of root hairs in that area. Additionally, the expression of ETC1::GUS was higher near this area. These results suggest that CPC family of genes also cooperatively regulates root hair formation at the root-hypocotyl junction in unique ways. PMID:26339713

  12. Identification of activation-tag Arabidopsis mutants with altered production of germination stimulants for Phelipanche ramosa (L.)

    PubMed Central

    Kirilova, Ina; Denev, Iliya D.; Bineva, Rumyana; Gevezova, Maria; Alexandrova, Milena; Kostov, Kaloyan; Batchvarova, Rossitza

    2014-01-01

    Germination of seeds of root parasites like broomrapes (Orobanchaceae) is tightly regulated by chemical products exuded from the roots of the host plant, known as germination stimulants (GSs). Changes in the levels of synthesis and emission of GS can allow the development of practical measures for control of the crops-harming parasitic species. However, the genes encoding enzymes responsible for GS biosynthesis are still unknown. We performed a large-scale screening of 62,000 Arabidopsis activation-tag mutants for alteration in susceptibility to Phelipanche ramosa and to identify lines with altered GS production among them. After five successive screenings we identified 36 lines with altered susceptibility to P. ramosa. Seven of them displayed altered levels of GS production. By using a combination of Southern blot and thermal asymmetric interlaced polymerase chain reaction (TAIL-PCR), we pinpointed the location of activation-tag constructs in these lines. A combination of differential display and quantitative real-time PCR (qRT-PCR) allowed us to identify several affected genes. Two of them are directly involved in isoprenoid biosynthetic pathway in chloroplasts, and we believe that their activation led to increased levels of GS production. We believe that these genes are responsible for increased GS production in five of the Arabidopsis lines resistant to P. ramosa. PMID:26740753

  13. A small GTP-binding protein from Arabidopsis thaliana functionally complements the yeast YPT6 null mutant.

    PubMed Central

    Bednarek, S Y; Reynolds, T L; Schroeder, M; Grabowski, R; Hengst, L; Gallwitz, D; Raikhel, N V

    1994-01-01

    A clone designated A.t.RAB6 encoding a small GTP-binding protein was isolated from a cDNA library of Arabidopsis thaliana leaf tissue. The predicted amino acid sequence was highly homologous to the mammalian and yeast counterparts, H.Rab6 and Ryh1/Ypt6, respectively. Lesser homology was found between the predicted Arabidopsis protein sequence and two small GTP-binding proteins isolated from plant species (44% homology to Zea mays Ypt1 and 43% homology to Nicotiana tabacum Rab5). Conserved stretches in the deduced amino acid sequence of A.t.Rab6 include four regions involved in GTP-binding, an effector region, and C-terminal cysteine residues required for prenylation and subsequent membrane attachment. Northern blot analysis demonstrated that A.t.Rab6 mRNA was expressed in root, leaf, stem, and flower tissues from A. thaliana with the highest levels present in roots. Escherichia coli produced histidine-tagged A.t.Rab6 protein-bound GTP, whereas a mutation in one of the guanine nucleotide-binding sites (asparagine122 to isoleucine) rendered it incapable of binding GTP. Functionally, the A.t.RAB6 gene was able to complement the temperature-sensitive phenotype of the YPT6 null mutant in yeast. The isolation of this gene will aid in the dissection of the machinery involved in soluble protein sorting at the trans-Golgi network of plants. PMID:8159788

  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. PMID:23029671

  15. A double mutant allele, csr1-4, of Arabidopsis thaliana encodes an acetolactate synthase with altered kinetics.

    PubMed

    Mourad, G; Williams, D; King, J

    1995-01-01

    A comparison is made of the kinetic characteristics of acetolactate synthase (EC 4.1.3.18) in extracts from Columbia wild type and four near-isogenic, herbicide-resistant mutants of Arabidopsis thaliana (L.) Heynh. The mutants used were the chlorsulfuron-resistant GH50 (csr1-1), the imazapyr-resistant GH90 (csr1-2), the triazolopyrimidine-resistant Tzp5 (csr1-3) and the multiherbicide-resistant, double mutant GM4.8 (csr1-4), derived from csr1-1 and csr1-2 by intragenic recombination (G. Mourad et al. 1994, Mol. Gen. Genet. 243, 178-184). Kmapp and Vmax values for the substrate pyruvate were unaffected by any of the mutations giving rise to herbicide resistance. Feedback inhibition by L-valine (L-Val), L-leucine (L-Leu) and L-isoleucine (L-Ile) of acetolactate synthase extracted from wild type and mutants fitted a mixed competitive pattern most closely. Ki values for L-Val, L-Leu and L-Ile inhibition were not significantly different from wild type in extracts from csr1-1, csr1-2, and csr1-3. Ki values were significantly higher than wild type by two- and five-fold, respectively, for csr1-4 with L-Val and L-Leu but not L-Ile. GM4.8 (csr1-4) plants were also highly resistant in their growth to added L-Val and L-Leu.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7767237

  16. 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. PMID:27208253

  17. An Effective Strategy for Reliably Isolating Heritable and Cas9-Free Arabidopsis Mutants Generated by CRISPR/Cas9-Mediated Genome Editing1[OPEN

    PubMed Central

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

    2016-01-01

    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. PMID:27208253

  18. Functional characterization of mutants affected in the carbonic anhydrase domain of the respiratory complex I in Arabidopsis thaliana.

    PubMed

    Soto, Débora; Córdoba, Juan Pablo; Villarreal, Fernando; Bartoli, Carlos; Schmitz, Jessica; Maurino, Veronica G; Braun, Hans Peter; Pagnussat, Gabriela C; Zabaleta, Eduardo

    2015-09-01

    The NADH-ubiquinone oxidoreductase complex (complex I) (EC 1.6.5.3) is the main entrance site of electrons into the respiratory chain. In a variety of eukaryotic organisms, except animals and fungi (Opisthokonta), it contains an extra domain comprising trimers of putative γ-carbonic anhydrases, named the CA domain, which has been proposed to be essential for assembly of complex I. However, its physiological role in plants is not fully understood. Here, we report that Arabidopsis mutants defective in two CA subunits show an altered photorespiratory phenotype. Mutants grown in ambient air show growth retardation compared to wild-type plants, a feature that is reversed by cultivating plants in a high-CO2 atmosphere. Moreover, under photorespiratory conditions, carbon assimilation is diminished and glycine accumulates, suggesting an imbalance with respect to photorespiration. Additionally, transcript levels of specific CA subunits are reduced in plants grown under non-photorespiratory conditions. Taken together, these results suggest that the CA domain of plant complex I contributes to sustaining efficient photosynthesis under ambient (photorespiratory) conditions. PMID:26148112

  19. Hormone Autotrophic Growth and Differentiation Identifies Mutant Lines of Arabidopsis with Altered Cytokinin and Auxin Content or Signaling1

    PubMed Central

    Frank, Markus; Rupp, Hans-Michael; Prinsen, Els; Motyka, Václav; Van Onckelen, Harry; Schmülling, Thomas

    2000-01-01

    We describe mutant tissue lines of Arabidopsis that are able to grow in vitro as callus on hormone-free medium. The 14 lines presented here show different hormone autotrophic differentiation behaviors that can be classified into three categories: (a) forming roots (rooty callus), (b) forming shoots or shoot-like structures (shooty callus), or (c) growing without organ formation (callus). Three fast-growing lines showed altered steady-state mRNA levels of the Cdc2 and CycD3 cell cycle genes. Three of the six rooty callus lines contained about 20- to 30-fold higher levels of auxins than wild-type callus. These and two other lines with normal auxin content showed an increased steady-state level of IAA1 and IAA2 transcripts in the absence of exogenous auxin. Five of the six shooty callus lines had increased steady-state mRNA levels of the CKI1 gene and/or of the homeobox genes KNAT1 and STM, suggesting that the phenotype is linked to altered cytokinin signaling. Also, one cytokinin-overproducing line with only 5% of wild-type cytokinin oxidase activity was identified. These results indicate that screening for hormone-autonomous growth identifies mutants with altered hormone content or signaling. PMID:10712535

  20. The irregular xylem 2 mutant is an allele of korrigan that affects the secondary cell wall of Arabidopsis thaliana.

    PubMed

    Szyjanowicz, Pio M J; McKinnon, Iain; Taylor, Neil G; Gardiner, John; Jarvis, Mike C; Turner, Simon R

    2004-03-01

    The irregular xylem 2 (irx2) mutant of Arabidopsis thaliana exhibits a cellulose deficiency in the secondary cell wall, which is brought about by a point mutation in the KORRIGAN (KOR) beta,1-4 endoglucanase (beta,1-4 EGase) gene. Measurement of the total crystalline cellulose in the inflorescence stem indicates that the irx2 mutant contains approximately 30% of the level present in the wild type (WT). Fourier-Transform Infra Red (FTIR) analysis, however, indicates that there is no decrease in cellulose in primary cell walls of the cortical and epidermal cells of the stem. KOR expression is correlated with cellulose synthesis and is highly expressed in cells synthesising a secondary cell wall. Co-precipitation experiments, using either an epitope-tagged form of KOR or IRX3 (AtCesA7), suggest that KOR is not an integral part of the cellulose synthase complex. These data are supported by immunolocalisation of KOR that suggests that KOR does not localise to sites of secondary cell wall deposition in the developing xylem. The defect in irx2 plant is consistent with a role for KOR in the later stages of secondary cell wall formation, suggesting a role in processing of the growing microfibrils or release of the cellulose synthase complex. PMID:14871312

  1. The rib1 Mutant Is Resistant to Indole-3-Butyric Acid, an Endogenous Auxin in Arabidopsis1

    PubMed Central

    Poupart, Julie; Waddell, Candace S.

    2000-01-01

    The presence of indole-3-butyric acid (IBA) as an endogenous auxin in Arabidopsis has been recently demonstrated. However, the in vivo role of IBA remains to be elucidated. We present the characterization of a semi-dominant mutant that is affected in its response to IBA, but shows a wild-type response to indole-3-acetic acid (IAA), the predominant and most studied form of auxin. We have named this mutant rib1 for resistant to IBA. Root elongation assays show that rib1 is specifically resistant to IBA, to the synthetic auxin 2,4-dichlorophenoxyacetic acid, and to auxin transport inhibitors. rib1 does not display increased resistance to IAA, to the synthetic auxin naphthalene acetic acid, or to other classes of plant hormones. rib1 individuals also have other root specific phenotypes including a shortened primary root, an increased number of lateral roots, and a more variable response than wild type to a change in gravitational vector. Adult rib1 plants are morphologically indistinguishable from wild-type plants. These phenotypes suggest that rib1 alters IBA activity in the root, thereby affecting root development and response to environmental stimuli. We propose models in which RIB1 has a function in either IBA transport or response. Our experiments also suggest that IBA does not use the same mechanism to exit cells as does IAA and we propose a model for IBA transport. PMID:11115890

  2. Plasma membrane lipid–protein interactions affect signaling processes in sterol-biosynthesis mutants in Arabidopsis thaliana

    PubMed Central

    Zauber, Henrik; Burgos, Asdrubal; Garapati, Prashanth; Schulze, Waltraud X.

    2014-01-01

    The plasma membrane is an important organelle providing structure, signaling and transport as major biological functions. Being composed of lipids and proteins with different physicochemical properties, the biological functions of membranes depend on specific protein–protein and protein–lipid interactions. Interactions of proteins with their specific sterol and lipid environment were shown to be important factors for protein recruitment into sub-compartmental structures of the plasma membrane. System-wide implications of altered endogenous sterol levels for membrane functions in living cells were not studied in higher plant cells. In particular, little is known how alterations in membrane sterol composition affect protein and lipid organization and interaction within membranes. Here, we conducted a comparative analysis of the plasma membrane protein and lipid composition in Arabidopsis sterol-biosynthesis mutants smt1 and ugt80A2;B1. smt1 shows general alterations in sterol composition while ugt80A2;B1 is significantly impaired in sterol glycosylation. By systematically analyzing different cellular fractions and combining proteomic with lipidomic data we were able to reveal contrasting alterations in lipid–protein interactions in both mutants, with resulting differential changes in plasma membrane signaling status. PMID:24672530

  3. The Arabidopsis nox mutant lacking carotene hydroxylase activity reveals a critical role for xanthophylls in photosystem I biogenesis.

    PubMed

    Dall'Osto, Luca; Piques, Maria; Ronzani, Michela; Molesini, Barbara; Alboresi, Alessandro; Cazzaniga, Stefano; Bassi, Roberto

    2013-02-01

    Carotenes, and their oxygenated derivatives xanthophylls, are essential components of the photosynthetic apparatus. They contribute to the assembly of photosynthetic complexes and participate in light absorption and chloroplast photoprotection. Here, we studied the role of xanthophylls, as distinct from that of carotenes, by characterizing a no xanthophylls (nox) mutant of Arabidopsis thaliana, which was obtained by combining mutations targeting the four carotenoid hydroxylase genes. nox plants retained α- and β-carotenes but were devoid in xanthophylls. The phenotype included depletion of light-harvesting complex (LHC) subunits and impairment of nonphotochemical quenching, two effects consistent with the location of xanthophylls in photosystem II antenna, but also a decreased efficiency of photosynthetic electron transfer, photosensitivity, and lethality in soil. Biochemical analysis revealed that the nox mutant was specifically depleted in photosystem I function due to a severe deficiency in PsaA/B subunits. While the stationary level of psaA/B transcripts showed no major differences between genotypes, the stability of newly synthesized PsaA/B proteins was decreased and translation of psaA/B mRNA was impaired in nox with respect to wild-type plants. We conclude that xanthophylls, besides their role in photoprotection and LHC assembly, are also needed for photosystem I core translation and stability, thus making these compounds indispensable for autotrophic growth. PMID:23396829

  4. An Arabidopsis Mutant Tolerant to Lethal Ultraviolet-B Levels Shows Constitutively Elevated Accumulation of Flavonoids and Other Phenolics1

    PubMed Central

    Bieza, Kim; Lois, Rodrigo

    2001-01-01

    The isolation and characterization of mutants hypersensitive to ultraviolet (UV) radiation has been a powerful tool to learn about the mechanisms that protect plants against UV-induced damage. To increase our understanding of the various mechanisms of defense against UVB radiation, we searched for mutations that would increase the level of tolerance of Arabidopsis plants to UV radiation. We describe a single gene dominant mutation (uvt1) that leads to a remarkable tolerance to UVB radiation conditions that would kill wild-type plants. Pigment analyses show a constitutive increase in accumulation of UV-absorbing compounds in uvt1 that increases the capacity of the leaves to block UVB radiation and therefore is likely to be responsible for the elevated resistance of this mutant to UVB radiation. These increases in absorption in the UV region are due, at least in part, to increases in flavonoid and sinapate accumulation. Expression of chalcone synthase (CHS) mRNA was shown to be constitutively elevated in uvt1 plants, suggesting that the increases in absorption may be a consequence of changes in gene expression. Expression of CHS in uvt1 was shown to be still inducible by UV, indicating that the uvt1 lesion may not affect the UV-mediated regulation of CHS gene expression. Our data support an important role for UV screens in the overall protection of plants to UVB radiation. The uvt1 mutant could prove to be an important tool to elucidate further the exact role of UV-absorbing pigments in UV protection as well as the relative contribution of other mechanisms to the overall tolerance of plants to UV radiation. PMID:11457961

  5. 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

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

    PubMed

    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

  7. 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. PMID:27432873

  8. 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.

  9. Quenching in Arabidopsis thaliana mutants lacking monomeric antenna proteins of photosystem II.

    PubMed

    Miloslavina, Yuliya; de Bianchi, Silvia; Dall'Osto, Luca; Bassi, Roberto; Holzwarth, Alfred R

    2011-10-21

    The minor light-harvesting complexes CP24, CP26, and CP29 have been proposed to play a key role in the zeaxanthin (Zx)-dependent high light-induced regulation (NPQ) of excitation energy in higher plants. To characterize the detailed roles of these minor complexes in NPQ and to determine their specific quenching effects we have studied the ultrafast fluorescence kinetics in knockout (ko) mutants koCP26, koCP29, and the double mutant koCP24/CP26. The data provide detailed insight into the quenching processes and the reorganization of the Photosystem (PS) II supercomplex under quenching conditions. All genotypes showed two NPQ quenching sites. Quenching site Q1 is formed by a light-induced functional detachment of parts of the PSII supercomplex and a pronounced quenching of the detached antenna parts. The antenna remaining bound to the PSII core was also quenched substantially in all genotypes under NPQ conditions (quenching site Q2) as compared with the dark-adapted state. The latter quenching was about equally strong in koCP26 and the koCP24/CP26 mutants as in the WT. Q2 quenching was substantially reduced, however, in koCP29 mutants suggesting a key role for CP29 in the total NPQ. The observed quenching effects in the knockout mutants are complicated by the fact that other minor antenna complexes do compensate in part for the lack of the CP24 and/or CP29 complexes. Their lack also causes some LHCII dissociation already in the dark. PMID:21844190

  10. Quenching in Arabidopsis thaliana Mutants Lacking Monomeric Antenna Proteins of Photosystem II*

    PubMed Central

    Miloslavina, Yuliya; de Bianchi, Silvia; Dall'Osto, Luca; Bassi, Roberto; Holzwarth, Alfred R.

    2011-01-01

    The minor light-harvesting complexes CP24, CP26, and CP29 have been proposed to play a key role in the zeaxanthin (Zx)-dependent high light-induced regulation (NPQ) of excitation energy in higher plants. To characterize the detailed roles of these minor complexes in NPQ and to determine their specific quenching effects we have studied the ultrafast fluorescence kinetics in knockout (ko) mutants koCP26, koCP29, and the double mutant koCP24/CP26. The data provide detailed insight into the quenching processes and the reorganization of the Photosystem (PS) II supercomplex under quenching conditions. All genotypes showed two NPQ quenching sites. Quenching site Q1 is formed by a light-induced functional detachment of parts of the PSII supercomplex and a pronounced quenching of the detached antenna parts. The antenna remaining bound to the PSII core was also quenched substantially in all genotypes under NPQ conditions (quenching site Q2) as compared with the dark-adapted state. The latter quenching was about equally strong in koCP26 and the koCP24/CP26 mutants as in the WT. Q2 quenching was substantially reduced, however, in koCP29 mutants suggesting a key role for CP29 in the total NPQ. The observed quenching effects in the knockout mutants are complicated by the fact that other minor antenna complexes do compensate in part for the lack of the CP24 and/or CP29 complexes. Their lack also causes some LHCII dissociation already in the dark. PMID:21844190

  11. Leaf apoplastic proteome composition in UV-B treated Arabidopsis thaliana mutants impaired in extracellular glutathione degradation

    PubMed Central

    Masi, A.; Trentin, A.R.; Arrigoni, G.

    2015-01-01

    In plants, environmental perturbations often result in oxidative reactions in the apoplastic space, which are counteracted for by enzymatic and non-enzymatic antioxidative systems, including ascorbate and glutathione. However, the occurrence of the latter and its exact role in the extracellular space are not well documented. 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 UV-B radiation and studied in redox altered ggt1 mutants. Elevated UV-B has detrimental effects on plant metabolism, plasma membranes representing a major target for ROS generated by this harmful radiation. The response of ggt1 knockout Arabidopsis leaves to UV-B radiation was assessed by investigating changes in apoplastic protein composition. We then compared the expression changes resulting from the mutation and from the UV-B treatment. Rearrangements occurring in apoplastic protein composition suggest the involvement of hydrogen peroxide, which may ultimately act as a signal. Other important changes related to hormonal effects, cell wall remodeling, and redox activities are also reported. We argue that oxidative stress conditions imposed by UV-B and by disruption of the gamma-glutamyl cycle result in similar stress-induced responses, to some degree at least. Data shown here are associated with the article from Trentin et al. (2015) [1]; protein data have been deposited to the PRIDE database (Vizcaíno et al., 2014) [2] with identifier PXD001807. PMID:26862584

  12. 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

  13. The rib1 Mutant of Arabidopsis Has Alterations in Indole-3-Butyric Acid Transport, Hypocotyl Elongation, and Root Architecture1

    PubMed Central

    Poupart, Julie; Rashotte, Aaron M.; Muday, Gloria K.; Waddell, Candace S.

    2005-01-01

    Polar transport of the auxin indole-3-butyric acid (IBA) has recently been shown to occur in Arabidopsis (Arabidopis thaliana) seedlings, yet the physiological importance of this process has yet to be fully resolved. Here we describe the first demonstration of altered IBA transport in an Arabidopsis mutant, and show that the resistant to IBA (rib1) mutation results in alterations in growth, development, and response to exogenous auxin consistent with an important physiological role for IBA transport. Both hypocotyl and root IBA basipetal transport are decreased in rib1 and root acropetal IBA transport is increased. While indole-3-acetic acid (IAA) transport levels are not different in rib1 compared to wild type, root acropetal IAA transport is insensitive to the IAA efflux inhibitor naphthylphthalamic acid in rib1, as is the dependent physiological process of lateral root formation. These observed changes in IBA transport are accompanied by altered rib1 phenotypes. Previously, rib1 roots were shown to be less sensitive to growth inhibition by IBA, but to have a wild-type response to IAA in root elongation. rib1 is also less sensitive to IBA in stimulation of lateral root formation and in hypocotyl elongation under most, but not all, light and sucrose conditions. rib1 has wild-type responses to IAA, except under one set of conditions, low light and 1.5% sucrose, in which both hypocotyl elongation and lateral root formation show altered IAA response. Taken together, our results support a model in which endogenous IBA influences wild-type seedling morphology. Modifications in IBA distribution in seedlings affect hypocotyl and root elongation, as well as lateral root formation. PMID:16258013

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

    PubMed

    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

  15. Genetic screens for floral mutants in Arabidopsis thaliana: enhancers and suppressors.

    PubMed

    Dinh, Thanh Theresa; Luscher, Elizabeth; Li, Shaofang; Liu, Xigang; Won, So Youn; Chen, Xuemei

    2014-01-01

    The flower is a hallmark feature that has contributed to the evolutionary success of land plants. Diverse mutagenic agents have been employed as a tool to genetically perturb flower development and identify genes involved in floral patterning and morphogenesis. Since the initial studies to identify genes governing processes such as floral organ specification, mutagenesis in sensitized backgrounds has been used to isolate enhancers and suppressors to further probe the molecular basis of floral development. Here, we first describe two commonly employed methods for mutagenesis (using ethyl methanesulfonate (EMS) or T-DNAs as mutagens), and then describe three methods for identifying a mutation that leads to phenotypic alterations--traditional map-based cloning, TAIL-PCR, and deep sequencing in the plant model Arabidopsis thaliana. PMID:24395255

  16. Genetic Screens for Floral Mutants in Arabidopsis thaliana: Enhancers and Suppressors

    PubMed Central

    Dinh, Thanh Theresa; Luscher, Elizabeth; Li, Shaofang; Liu, Xigang; Won, So Youn; Chen, Xuemei

    2015-01-01

    The flower is a hallmark feature that has contributed to the evolutionary success of land plants. Diverse mutagenic agents have been employed as a tool to genetically perturb flower development and identify genes involved in floral patterning and morphogenesis. Since the initial studies to identify genes governing processes such as floral organ specification, mutagenesis in sensitized backgrounds has been used to isolate enhancers and suppressors to further probe the molecular basis of floral development. Here, we first describe two commonly employed methods for mutagenesis (using ethyl methanesulfonate (EMS) or T-DNAs as mutagens), and then describe three methods for identifying a mutation that leads to phenotypic alterations—traditional map-based cloning, TAIL-PCR, and deep sequencing in the plant model Arabidopsis thaliana. PMID:24395255

  17. A Protein-Based Genetic Screening Uncovers Mutants Involved in Phytochrome Signaling in Arabidopsis

    PubMed Central

    Zhu, Ling; Xin, Ruijiao; Huq, Enamul

    2016-01-01

    Plants perceive red and far-red region of the light spectrum to regulate photomorphogenesis through a family of photoreceptors called phytochromes. Phytochromes transduce the light signals to trigger a cascade of downstream gene regulation in part via a subfamily of bHLH transcription factors called Phytochrome Interacting Factors (PIFs). As the repressors of light signaling pathways, most PIFs are phosphorylated and degraded through the ubiquitin/26S proteasome pathway in response to light. The mechanisms involved in the phosphorylation and degradation of PIFs have not been fully understood yet. Here we used an EMS mutagenesis and luminescent imaging system to identify mutants defective in the degradation of one of the PIFs, called PIF1. We identified five mutants named stable PIF (spf) that showed reduced degradation of PIF1 under light treatment in both luminescent imaging and immunoblot assays. The amounts of PIF1 in spf3, spf4, and spf5 were similar to a PIF1 missense mutant (PIF1–3M) that lacks interactions between PIF1 and phyA/phyB under light. The hypocotyl lengths of spf1 and spf2 were slightly longer under red light compared to the LUC-PIF1 control, while only spf1 displayed weak phenotype under far-red light conditions. Interestingly, the spf3, spf4, and spf5 displayed high abundance of PIF1, yet the hypocotyl lengths were similar to the wild type under these conditions. Cloning and characterization of these mutants will help identify key players in the light signaling pathways including, the light-regulated kinase(s) and the E3 ligase(s) necessary for the light-induced degradation of PIFs. PMID:27499759

  18. 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.

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

    PubMed

    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

  20. Lutein Accumulation in the Absence of Zeaxanthin Restores Nonphotochemical Quenching in the Arabidopsis thaliana npq1 Mutant[W][OA

    PubMed Central

    Li, Zhirong; Ahn, Tae Kyu; Avenson, Thomas J.; Ballottari, Matteo; Cruz, Jeffrey A.; Kramer, David M.; Bassi, Roberto; Fleming, Graham R.; Keasling, Jay D.; Niyogi, Krishna K.

    2009-01-01

    Plants protect themselves from excess absorbed light energy through thermal dissipation, which is measured as nonphotochemical quenching of chlorophyll fluorescence (NPQ). The major component of NPQ, qE, is induced by high transthylakoid ΔpH in excess light and depends on the xanthophyll cycle, in which violaxanthin and antheraxanthin are deepoxidized to form zeaxanthin. To investigate the xanthophyll dependence of qE, we identified suppressor of zeaxanthin-less1 (szl1) as a suppressor of the Arabidopsis thaliana npq1 mutant, which lacks zeaxanthin. szl1 npq1 plants have a partially restored qE but lack zeaxanthin and have low levels of violaxanthin, antheraxanthin, and neoxanthin. However, they accumulate more lutein and α-carotene than the wild type. szl1 contains a point mutation in the lycopene β-cyclase (LCYB) gene. Based on the pigment analysis, LCYB appears to be the major lycopene β-cyclase and is not involved in neoxanthin synthesis. The Lhcb4 (CP29) and Lhcb5 (CP26) protein levels are reduced by 50% in szl1 npq1 relative to the wild type, whereas other Lhcb proteins are present at wild-type levels. Analysis of carotenoid radical cation formation and leaf absorbance changes strongly suggest that the higher amount of lutein substitutes for zeaxanthin in qE, implying a direct role in qE, as well as a mechanism that is weakly sensitive to carotenoid structural properties. PMID:19549928

  1. A vacuolar carboxypeptidase mutant of Arabidopsis thaliana is degraded by the ERAD pathway independently of its N-glycan

    SciTech Connect

    Yamamoto, Masaya; Kawanabe, Mitsuyoshi; Hayashi, Yoko; Endo, Toshiya; Nishikawa, Shuh-ichi

    2010-03-12

    Misfolded proteins produced in the endoplasmic reticulum (ER) are degraded by a mechanism, the ER-associated degradation (ERAD). Here we report establishment of the experimental system to analyze the ERAD in plant cells. Carboxypeptidase Y (CPY) is a vacuolar enzyme and its mutant CPY* is degraded by the ERAD in yeast. Since Arabidopsis thaliana has AtCPY, an ortholog of yeast CPY, we constructed and expressed fusion proteins consisting of AtCPY and GFP and of AtCPY*, which carries a mutation homologous to yeast CPY*, and GFP in A. thaliana cells. While AtCPY-GFP was efficiently transported to the vacuole, AtCPY*-GFP was retained in the ER to be degraded in proteasome- and Cdc48-dependent manners. We also found that AtCPY*-GFP was degraded by the ERAD in yeast cells, but that its single N-glycan did not function as a degradation signal in yeast or plant cells. Therefore, AtCPY*-GFP can be used as a marker protein to analyze the ERAD pathway, likely for nonglycosylated substrates, in plant cells.

  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. Intragenic Suppression of a Trafficking-Defective Brassinosteroid Receptor Mutant in Arabidopsis

    PubMed Central

    Belkhadir, Youssef; Durbak, Amanda; Wierzba, Michael; Schmitz, Robert J.; Aguirre, Andrea; Michel, Rene; Rowe, Scott; Fujioka, Shozo; Tax, Frans E.

    2010-01-01

    The cell surface receptor kinase BRASSINOSTEROID-INSENSITIVE-1 (BRI1) is the major receptor for steroid hormones in Arabidopsis. Plants homozygous for loss-of-function mutations in BRI1 display a reduction in the size of vegetative organs, resulting in dwarfism. The recessive bri1-5 mutation produces receptors that do not accumulate to wild-type levels and are retained mainly in the endoplasmic reticulum. We have isolated a dominant suppressor of the dwarf phenotype of bri1-5 plants. We show that this suppression is caused by a second-site mutation in BRI1, bri1-5R1. The bri1-5R1 mutation partially rescues the phenotypes of bri1-5 in many tissues and enhances bri1-5 phenotypes above wild-type levels in several other tissues. We demonstrate that the phenotypes of bri1-5R1 plants are due to both increased cell expansion and increased cell division. To test the mechanism of bri1-5 suppression, we assessed whether the phenotypic suppression in bri1-5R1 was dependent on ligand availability and the integrity of the signaling pathway. Our results indicate that the suppression of the dwarf phenotypes associated with bri1-5R1 requires both BR biosynthesis and the receptor kinase BRI1-ASSOCIATED KINASE-1 (BAK1). Finally, we show that bri1-5R1 partially restores the accumulation and plasma membrane localization of BRI1. Collectively, our results point toward a model in which bri1-R1 compensates for the protein-folding abnormalities caused by bri1-5, restoring accumulation of the receptor and its delivery to the cell surface. PMID:20457881

  4. 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

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

    PubMed

    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

  6. 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.

  7. 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

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

    DOE PAGESBeta

    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 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

  9. 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

  10. 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. PMID:27050746

  11. Accumulation of γ- Rather than α-Tocopherol Alters Ethylene Signaling Gene Expression in the vte4 Mutant of Arabidopsis thaliana

    PubMed Central

    Cela, Jana; Chang, Caren; Munné-Bosch, Sergi

    2011-01-01

    Tocopherols are antioxidants found in chloroplasts of leaves, and it is a matter of current debate whether or not they can affect signaling and gene expression in plant cells. For insight into the possible effects of altered tocopherol composition in chloroplasts on gene expression in the nucleus, the expression of ethylene biosynthesis, perception and signaling genes was investigated in vte1 and vte4 Arabidopsis thaliana mutants, which are impaired in tocopherol (vitamin E) biosynthesis. Changes in gene expression were measured in plants exposed to either salt or water stress, and in young and mature leaves of vte1 and vte4 mutants, which lack tocopherol cyclase and γ-tocopherol methyltransferase, respectively. While transcript levels of ethylene signaling genes in the vte1 mutant and the wild type were similar in all tested conditions, major changes in gene expression occurred in the vte4 mutant, particularly in mature leaves (compared with young leaves) and under salt stress. Accumulation of γ- instead of α-tocopherol in this mutant led to elevated transcript levels of ethylene signaling pathway genes (particularly CTR1, EIN2, EIN3 and ERF1) in mature leaves of control plants. However, with salt treatment, transcript levels of most of these genes remained constant or dropped in the vte4 mutant, while they were dramatically induced in the wild type and the vte1 mutant. Furthermore, under salt stress, leaf age-induced jasmonic acid accumulated in both the vte1 mutant and the wild type, but not in the vte4 mutant. It is concluded that jasmonic acid and ethylene signaling pathways are down-regulated in mature leaves of salt-stressed vte4 plants. PMID:21719428

  12. 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. PMID:24373011

  13. 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

  14. Arabidopsis phosphatase under-producer mutants pup1 and pup3 contain mutations in the AtPAP10 and AtPAP26 genes

    PubMed Central

    Zhang, Ye; Wang, Xiaoyue; Liu, Dong

    2015-01-01

    Production and secretion of acid phosphatases (APases) is a hallmark adaptive response of plants to phosphate (Pi) deprivation. Researchers have long hypothesized that Pi starvation-induced APases are involved in internal Pi recycling and remobilization as well as in external Pi utilization. Two phosphatase under-producer (pup) mutants, pup1 and pup3, were previously isolated in Arabidopsis. Characterization of these 2 pup mutants provided the first genetic evidence for the above hypothesis. To date, however, the molecular lesions in these 2 pup mutants remain unknown. In this work, we demonstrate that pup1 and pup3 contain point mutations in the Arabidopsis purple acid phosphatase gene AtPAP10 and AtPAP26, respectively. Our results answer a long-standing question about the molecular identity of the PUP1 and PUP3 genes and corroborate the conclusions from previous studies regarding the function of AtPAP10 and AtPAP26 in plant acclimation to Pi deprivation. PMID:26251878

  15. Desaturase mutants reveal that membrane rigidification acts as a cold perception mechanism upstream of the diacylglycerol kinase pathway in Arabidopsis cells.

    PubMed

    Vaultier, Marie-Noëlle; Cantrel, Catherine; Vergnolle, Chantal; Justin, Anne-Marie; Demandre, Chantal; Benhassaine-Kesri, Ghouziel; Ciçek, Dominique; Zachowski, Alain; Ruelland, Eric

    2006-07-24

    Membrane rigidification could be the first step of cold perception in poikilotherms. We have investigated its implication in diacylglycerol kinase (DAGK) activation by cold stress in suspension cells from Arabidopsis mutants altered in desaturase activities. By lateral diffusion assay, we showed that plasma membrane rigidification with temperature decrease was steeper in cells deficient in oleate desaturase than in wild type cells and in cells overexpressing linoleate desaturase. The threshold for the activation of the DAGK pathway in each type of cells correlated with this order of rigidification rate, suggesting that cold induced-membrane rigidification is upstream of DAGK pathway activation. PMID:16839551

  16. Proteomic and biochemical analyses show a functional network of proteins involved in antioxidant defense of the Arabidopsis anp2anp3 double mutant.

    PubMed

    Takáč, Tomáš; Šamajová, Olga; Vadovič, Pavol; Pechan, Tibor; Košútová, Petra; Ovečka, Miroslav; Husičková, Alexandra; Komis, George; Šamaj, Jozef

    2014-12-01

    Disentanglement of functional complexity associated with plant mitogen-activated protein kinase (MAPK) signaling has benefited from transcriptomic, proteomic, phosphoproteomic, and genetic studies. Published transcriptomic analysis of a double homozygous recessive anp2anp3 mutant of two MAPK kinase kinase (MAPKKK) genes called Arabidopsis thaliana Homologues of Nucleus- and Phragmoplast-localized Kinase 2 (ANP2) and 3 (ANP3) showed the upregulation of stress-related genes. In this study, a comparative proteomic analysis of anp2anp3 mutant against its respective Wassilevskaja ecotype (Ws) wild type background is provided. Such differential proteomic analysis revealed overabundance of core enzymes such as FeSOD1, MnSOD, DHAR1, and FeSOD1-associated regulatory protein CPN20, which are involved in the detoxification of reactive oxygen species in the anp2anp3 mutant. The proteomic results were validated at the level of single protein abundance by Western blot analyses and by quantitative biochemical determination of antioxidant enzymatic activities. Finally, the functional network of proteins involved in antioxidant defense in the anp2anp3 mutant was physiologically linked with the increased resistance of mutant seedlings against paraquat treatment. PMID:25325904

  17. 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

  18. Ultraviolet-B Radiation Impacts Light-Mediated Turnover of the Photosystem II Reaction Center Heterodimer in Arabidopsis Mutants Altered in Phenolic Metabolism

    PubMed Central

    Booij-James, Isabelle S.; Dube, Shyam K.; Jansen, Marcel A.K.; Edelman, Marvin; Mattoo, Autar K.

    2000-01-01

    Ultraviolet-B (UV-B) radiation can have a negative impact on the growth and development of plants. Plants tolerant to UV-B alleviate these effects using UV-screening pigments that reduce the penetration of UV-B into mesophyll tissue. Little is known about the relative contribution of specific phenolic compounds to the screening capacity of leaves. The D1 and D2 proteins constituting the photosystem (PS) II reaction center heterodimer are targets of UV-B radiation and can be used as an in situ sensor for UV penetration into photosynthetic tissue. Degradation of these proteins occurs under very low fluences of UV-B, and is strongly accelerated in the presence of visible light. Using the D1-D2 degradation assay, we characterized UV-B sensitivity of Arabidopsis mutants (tt4, tt5, and fah1) that are genetically altered in their composition of phenolic compounds. We found that changes in phenol metabolism result in altered rates of PSII reaction center heterodimer degradation under mixtures of photosynthetically active radiation and UV-B. A comparison of D2 degradation kinetics showed increased UV sensitivity of the Landsberg (Landsberg erecta) tt5 mutant relative to the Landsberg tt4 mutant and the Landsberg wild type. Despite a lack of flavonoid accumulation, the tt4 mutant is not particularly UV sensitive. However, the tolerance of this mutant to UV-B may reflect the increased accumulation of sinapate esters that strongly absorb in the UV range, and may thus protect the plant against environmentally relevant UV-B radiation. This sinapate-mediated protection is less obvious for the tt4 mutant of Columbia ecotype, indicating that the relative contribution of particular phenolics to the total screening capacity varies with the genetic background. The role of sinapate esters in UV screening is further substantiated by the results with the fah1 mutant where absence of most of the sinapate esters results in a significantly accelerated degradation of D2 under mixed light

  19. 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). PMID:19880215

  20. Gene expression profile of zeitlupe/lov kelch protein1 T-DNA insertion mutants in Arabidopsis thaliana: Downregulation of auxin-inducible genes in hypocotyls

    PubMed Central

    Saitoh, Aya; Takase, Tomoyuki; Kitaki, Hiroyuki; Miyazaki, Yuji; Kiyosue, Tomohiro

    2015-01-01

    Elongation of hypocotyl cells has been studied as a model for elucidating the contribution of cellular expansion to plant organ growth. ZEITLUPE (ZTL) or LOV KELCH PROTEIN1 (LKP1) is a positive regulator of warmth-induced hypocotyl elongation under white light in Arabidopsis, although the molecular mechanisms by which it promotes hypocotyl cell elongation remain unknown. Microarray analysis showed that 134 genes were upregulated and 204 genes including 15 auxin-inducible genes were downregulated in the seedlings of 2 ztl T-DNA insertion mutants grown under warm conditions with continuous white light. Application of a polar auxin transport inhibitor, an auxin antagonist or an auxin biosynthesis inhibitor inhibited hypocotyl elongation of control seedlings to the level observed with the ztl mutant. Our data suggest the involvement of auxin and auxin-inducible genes in ZTL-mediated hypocotyl elongation. PMID:26237185

  1. The effect of NaCl on stomatal opening in Arabidopsis wild type and agb1 heterotrimeric G-protein mutant plants.

    PubMed

    Yu, Yunqing; Assmann, Sarah M

    2016-01-01

    Salinity is a major agricultural problem that affects crop yield. Na(+) is transported to the shoot through the transpiration stream. The mutant of the sole Arabidopsis heterotrimeric G protein β subunit, agb1, is hypersensitive to salinity in part due to a higher transpiration rate. Here, we investigated the direct effect of Na(+) on stomatal opening using detached epidermal peels of wild type and agb1 plants. In both genotypes, NaCl is equally as effective as KCl in mediating stomatal opening at the concentrations tested. In both genotypes, ABA is less effective in inhibiting Na(+) mediated stomatal opening than K(+) mediated stomatal opening. The agb1 mutant is hyposensitive to ABA inhibition of K(+)-mediated but not Na(+)-mediated stomatal opening. These results suggest that the greater transpiration observed in agb1 plants grown in saline conditions is likely not mediated by differential genotypic direct effects of Na(+) on stomatal apertures. PMID:26431457

  2. The effect of NaCl on stomatal opening in Arabidopsis wild type and agb1 heterotrimeric G-protein mutant plants

    PubMed Central

    Yu, Yunqing; Assmann, Sarah M.

    2016-01-01

    Salinity is a major agricultural problem that affects crop yield. Na+ is transported to the shoot through the transpiration stream. The mutant of the sole Arabidopsis heterotrimeric G protein β subunit, agb1, is hypersensitive to salinity in part due to a higher transpiration rate. Here, we investigated the direct effect of Na+ on stomatal opening using detached epidermal peels of wild type and agb1 plants. In both genotypes, NaCl is equally as effective as KCl in mediating stomatal opening at the concentrations tested. In both genotypes, ABA is less effective in inhibiting Na+ mediated stomatal opening than K+ mediated stomatal opening. The agb1 mutant is hyposensitive to ABA inhibition of K+-mediated but not Na+-mediated stomatal opening. These results suggest that the greater transpiration observed in agb1 plants grown in saline conditions is likely not mediated by differential genotypic direct effects of Na+ on stomatal apertures. PMID:26431457

  3. Arabidopsis Cytokinin Receptor Mutants Reveal Functions in Shoot Growth, Leaf Senescence, Seed Size, Germination, Root Development, and Cytokinin MetabolismW⃞

    PubMed Central

    Riefler, Michael; Novak, Ondrej; Strnad, Miroslav; Schmülling, Thomas

    2006-01-01

    We used loss-of-function mutants to study three Arabidopsis thaliana sensor histidine kinases, AHK2, AHK3, and CRE1/AHK4, known to be cytokinin receptors. Mutant seeds had more rapid germination, reduced requirement for light, and decreased far-red light sensitivity, unraveling cytokinin functions in seed germination control. Triple mutant seeds were more than twice as large as wild-type seeds. Genetic analysis indicated a cytokinin-dependent endospermal and/or maternal control of embryo size. Unchanged red light sensitivity of mutant hypocotyl elongation suggests that previously reported modulation of red light signaling by A-type response regulators may not depend on cytokinin. Combined loss of AHK2 and AHK3 led to the most prominent changes during vegetative development. Leaves of ahk2 ahk3 mutants formed fewer cells, had reduced chlorophyll content, and lacked the cytokinin-dependent inhibition of dark-induced chlorophyll loss, indicating a prominent role of AHK2 and, particularly, AHK3 in the control of leaf development. ahk2 ahk3 double mutants developed a strongly enhanced root system through faster growth of the primary root and, more importantly, increased branching. This result supports a negative regulatory role for cytokinin in root growth regulation. Increased cytokinin content of receptor mutants indicates a homeostatic control of steady state cytokinin levels through signaling. Together, the analyses reveal partially redundant functions of the cytokinin receptors and prominent roles for the AHK2/AHK3 receptor combination in quantitative control of organ growth in plants, with opposite regulatory functions in roots and shoots. PMID:16361392

  4. Seed germination of GA-insensitive sleepy1 mutants does not require RGL2 protein disappearance in Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Seed germination is a complex developmental process regulated by phytohormones. The phytohormone abscisic acid (ABA) inhibits seed germination, whereas gibberellin (GA) stimulates seed germination. In tomato and Arabidopsis, GA is clearly required for seed germination. Recent evidence suggests tha...

  5. 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

  6. 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.

  7. Metabolome Analysis of Biosynthetic Mutants Reveals a Diversity of Metabolic Changes and Allows Identification of a Large Number of New Compounds in Arabidopsis1[W][OA

    PubMed Central

    Böttcher, Christoph; von Roepenack-Lahaye, Edda; Schmidt, Jürgen; Schmotz, Constanze; Neumann, Steffen; Scheel, Dierk; Clemens, Stephan

    2008-01-01

    Metabolomics is facing a major challenge: the lack of knowledge about metabolites present in a given biological system. Thus, large-scale discovery of metabolites is considered an essential step toward a better understanding of plant metabolism. We show here that the application of a metabolomics approach generating structural information for the analysis of Arabidopsis (Arabidopsis thaliana) mutants allows the efficient cataloging of metabolites. Fifty-six percent of the features that showed significant differences in abundance between seeds of wild-type, transparent testa4, and transparent testa5 plants could be annotated. Seventy-five compounds were structurally characterized, 21 of which could be identified. About 40 compounds had not been known from Arabidopsis before. Also, the high-resolution analysis revealed an unanticipated expansion of metabolic conversions upstream of biosynthetic blocks. Deficiency in chalcone synthase results in the increased seed-specific biosynthesis of a range of phenolic choline esters. Similarly, a lack of chalcone isomerase activity leads to the accumulation of various naringenin chalcone derivatives. Furthermore, our data provide insight into the connection between p-coumaroyl-coenzyme A-dependent pathways. Lack of flavonoid biosynthesis results in elevated synthesis not only of p-coumarate-derived choline esters but also of sinapate-derived metabolites. However, sinapoylcholine is not the only accumulating end product. Instead, we observed specific and sophisticated changes in the complex pattern of sinapate derivatives. PMID:18552234

  8. Deficiency in phylloquinone (vitamin K1) methylation affects prenyl quinone distribution, photosystem I abundance, and anthocyanin accumulation in the Arabidopsis AtmenG mutant.

    PubMed

    Lohmann, Antje; Schöttler, Mark Aurel; Bréhélin, Claire; Kessler, Felix; Bock, Ralph; Cahoon, Edgar B; Dörmann, Peter

    2006-12-29

    Phylloquinone (vitamin K(1)) is synthesized in cyanobacteria and in chloroplasts of plants, where it serves as electron carrier of photosystem I. The last step of phylloquinone synthesis in cyanobacteria is the methylation of 2-phytyl-1,4-naphthoquinone by the menG gene product. Here, we report that the uncharacterized Arabidopsis gene At1g23360, which shows sequence similarity to menG, functionally complements the Synechocystis menG mutant. An Arabidopsis mutant, AtmenG, carrying a T-DNA insertion in the gene At1g23360 is devoid of phylloquinone, but contains an increased amount of 2-phytyl-1,4-naphthoquinone. Phylloquinone and 2-phytyl-1,4-naphthoquinone in thylakoid membranes of wild type and AtmenG, respectively, predominantly localize to photosystem I, whereas excess amounts of prenyl quinones are stored in plastoglobules. Photosystem I reaction centers are decreased in AtmenG plants under high light, as revealed by immunoblot and spectroscopic measurements. Anthocyanin accumulation and chalcone synthase (CHS1) transcription are affected during high light exposure, indicating that alterations in photosynthesis in AtmenG affect gene expression in the nucleus. Photosystem II quantum yield is decreased under high light. Therefore, the loss of phylloquinone methylation affects photosystem I stability or turnover, and the limitation in functional photosystem I complexes results in overreduction of photosystem II under high light. PMID:17082184

  9. Isolation and characterization of mutants of Arabidopsis thaliana with increased resistance to growth inhibition by indoleacetic acid-amino acid conjugates.

    PubMed Central

    Campanella, J J; Ludwig-Mueller, J; Town, C D

    1996-01-01

    Two mutants of Arabidopsis thaliana that are resistant to growth inhibition by indole-3-acetic acid (IAA)-phenylalanine have been isolated. Both mutants were 2- to 3-fold more resistant than wild type to inhibition by IAA-phenylalanine, IAA-alanine, and IAA-glycine in root growth assays. The mutant icr1 (but not icr2) also shows some resistance to IAA-aspartate. Studies using 3H-labeled IAA-phenylalanine showed that the uptake of conjugate from the medium by icr1 was the same as wild type and was reduced by about 25% in icr2. No differences in hydrolysis of the exogenous conjugate were detected between the mutants and their wild-type parents. There was no significant metabolism of the IAA released from the [3H]IAA-phenylalanine, whereas exogenous [3H]IAA was rapidly metabolized to two unidentified products considerably more polar than IAA. Analysis of a cross between icr1 and icr2 indicated that these mutations were at distinct loci and that their effects were additive, and preliminary mapping data indicated that icr1 and icr2 were located at the top and bottom of chromosome V, respectively. PMID:8883385

  10. The phospholipid-deficient pho1 mutant of Arabidopsis thaliana is affected in the organization, but not in the light acclimation, of the thylakoid membrane.

    PubMed

    Härtel, H; Essigmann, B; Lokstein, H; Hoffmann-Benning, S; Peters-Kottig, M; Benning, C

    1998-12-01

    The pho1 mutant of Arabidopsis has been shown to respond to the phosphate deficiency in the leaves by decreasing the amount of phosphatidylglycerol (PG). PG is thought to be of crucial importance for the organization and function of the thylakoid membrane. This prompted us to ask what the consequences of the PG deficiency may be in the pho1 mutant when grown under low or high light. While in the wild-type, the lipid pattern was almost insensitive to changes in the growth light, PG was reduced to 45% under low light in the mutant, and it decreased further to 35% under high light. Concomitantly, sulfoquinovosyl diacylglycerol (SQDG) and to a lesser extent digalactosyl diacylglycerol (DGDG) increased. The SQDG increase correlated with increased amounts of the SQD1 protein, an indicator for an actively mediated process. Despite of alterations in the ultrastructure, mutant thylakoids showed virtually no effects on photosynthetic electron transfer, O2 evolution and excitation energy allocation to the reaction centers. Our results support the idea that PG deficiency can at least partially be compensated for by the anionic lipid SQDG and the not charged lipid DGDG. This seems to be an important strategy to maintain an optimal thylakoid lipid milieu for vital processes, such as photosynthesis, under a restricted phosphate availability. PMID:9858733

  11. The Arabidopsis root hair cell wall formation mutant lrx1 is suppressed by mutations in the RHM1 gene encoding a UDP-L-rhamnose synthase.

    PubMed

    Diet, Anouck; Link, Bruce; Seifert, Georg J; Schellenberg, Barbara; Wagner, Ulrich; Pauly, Markus; Reiter, Wolf-Dieter; Ringli, Christoph

    2006-07-01

    Cell and cell wall growth are mutually dependent processes that must be tightly coordinated and controlled. LRR-extensin1 (LRX1) of Arabidopsis thaliana is a potential regulator of cell wall development, consisting of an N-terminal leucine-rich repeat domain and a C-terminal extensin-like domain typical for structural cell wall proteins. LRX1 is expressed in root hairs, and lrx1 mutant plants develop distorted root hairs that often swell, branch, or collapse. The aberrant cell wall structures found in lrx1 mutants point toward a function of LRX1 during the establishment of the extracellular matrix. To identify genes that are involved in an LRX1-dependent developmental pathway, a suppressor screen was performed on the lrx1 mutant, and two independent rol1 (for repressor of lrx1) alleles were isolated. ROL1 is allelic to Rhamnose Biosynthesis1, which codes for a protein involved in the biosynthesis of rhamnose, a major monosaccharide component of pectin. The rol1 mutations modify the pectic polysaccharide rhamnogalacturonan I and, for one allele, rhamnogalacturonan II. Furthermore, the rol1 mutations cause a change in the expression of a number of cell wall-related genes. Thus, the lrx1 mutant phenotype is likely to be suppressed by changes in pectic polysaccharides or other cell wall components. PMID:16766693

  12. 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

  13. Isolation of a mutant Arabidopsis plant that lacks N-acetyl glucosaminyl transferase I and is unable to synthesize Golgi-modified complex N-linked glycans.

    PubMed Central

    von Schaewen, A; Sturm, A; O'Neill, J; Chrispeels, M J

    1993-01-01

    The complex asparagine-linked glycans of plant glycoproteins, characterized by the presence of beta 1-->2 xylose and alpha 1-->3 fucose residues, are derived from typical mannose9(N-acetylglucosamine)2 (Man9GlcNAc2) 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 Arabidopsis 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 Man5GlcNAc1 glycans. In the presence of deoxymannojirimycin, an inhibitor of mannosidase I, the mutant cells synthesize Man9GlcNAc2 and Man8GlcNAc2 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, we obtained a unique strain that synthesizes phytohemagglutinin with two high-mannose glycans, instead of one high-mannose and one complex glycan. PMID:8278542

  14. New Connections across Pathways and Cellular Processes: Industrialized Mutant Screening Reveals Novel Associations between Diverse Phenotypes in Arabidopsis1[W][OA

    PubMed Central

    Lu, Yan; Savage, Linda J.; Ajjawi, Imad; Imre, Kathleen M.; Yoder, David W.; Benning, Christoph; DellaPenna, Dean; Ohlrogge, John B.; Osteryoung, Katherine W.; Weber, Andreas P.; Wilkerson, Curtis G.; Last, Robert L.

    2008-01-01

    In traditional mutant screening approaches, genetic variants are tested for one or a small number of phenotypes. Once bona fide variants are identified, they are typically subjected to a limited number of secondary phenotypic screens. Although this approach is excellent at finding genes involved in specific biological processes, the lack of wide and systematic interrogation of phenotype limits the ability to detect broader syndromes and connections between genes and phenotypes. It could also prevent detection of the primary phenotype of a mutant. As part of a systems biology approach to understand plastid function, large numbers of Arabidopsis thaliana homozygous T-DNA lines are being screened with parallel morphological, physiological, and chemical phenotypic assays (www.plastid.msu.edu). To refine our approaches and validate the use of this high-throughput screening approach for understanding gene function and functional networks, approximately 100 wild-type plants and 13 known mutants representing a variety of phenotypes were analyzed by a broad range of assays including metabolite profiling, morphological analysis, and chlorophyll fluorescence kinetics. Data analysis using a variety of statistical approaches showed that such industrial approaches can reliably identify plant mutant phenotypes. More significantly, the study uncovered previously unreported phenotypes for these well-characterized mutants and unexpected associations between different physiological processes, demonstrating that this approach has strong advantages over traditional mutant screening approaches. Analysis of wild-type plants revealed hundreds of statistically robust phenotypic correlations, including metabolites that are not known to share direct biosynthetic origins, raising the possibility that these metabolic pathways have closer relationships than is commonly suspected. PMID:18263779

  15. Expression of a nuclear-encoded psbH gene complements the plastidic RNA processing defect in the PSII mutant hcf107 in Arabidopsis thaliana.

    PubMed

    Levey, Tatjana; Westhoff, Peter; Meierhoff, Karin

    2014-10-01

    The helical-repeat RNA-binding protein HCF107 is required for processing, stabilization and translation of plastid-encoded psbH mRNA. The psbH gene encodes a small, hydrophilic subunit of the PSII complex and is part of the plastidic psbB-psbT-psbH-petB-petD transcription unit. In Arabidopsis hcf107 mutants, only trace amounts of PSII proteins can be detected. Beside drastically reduced synthesis of PsbH, the synthesis of CP47 was also reduced in these mutants, although the corresponding psbB transcripts accumulate to wild type levels. This situation raises the question, whether the reduction of CP47 is a direct consequence of the mutation, based on targeting of HCF107 to the psbB mRNA, or a secondary affect due to the absent PsbH. To clarify this issue we introduced a chimeric psbH construct comprising a sequence encoding a chloroplast transit peptide into the hcf107-2 background. We found that the nucleus-localized psbH was able to complement the mutant defect resulting in photoautotrophic plants. The PSII proteins CP47 and D1 accumulated to barely half of the wild type level. Further experiments showed that cytosolically synthesized PsbH was imported into chloroplasts and assembled into PSII complexes. Using this approach, we showed that the tetratricopeptide repeat protein HCF107 of Arabidopsis is only responsible for expression of PsbH and not for synthesis of CP47. In addition the data suggest the necessity of the small, one-helix membrane spanning protein PsbH for the accumulation of CP47 in higher plants. PMID:25081859

  16. Reactive oxygen species and transcript analysis upon excess light treatment in wild-type Arabidopsis thaliana vs a photosensitive mutant lacking zeaxanthin and lutein

    PubMed Central

    2011-01-01

    Background Reactive oxygen species (ROS) are unavoidable by-products of oxygenic photosynthesis, causing progressive oxidative damage and ultimately cell death. Despite their destructive activity they are also signalling molecules, priming the acclimatory response to stress stimuli. Results To investigate this role further, we exposed wild type Arabidopsis thaliana plants and the double mutant npq1lut2 to excess light. The mutant does not produce the xanthophylls lutein and zeaxanthin, whose key roles include ROS scavenging and prevention of ROS synthesis. Biochemical analysis revealed that singlet oxygen (1O2) accumulated to higher levels in the mutant while other ROS were unaffected, allowing to define the transcriptomic signature of the acclimatory response mediated by 1O2 which is enhanced by the lack of these xanthophylls species. The group of genes differentially regulated in npq1lut2 is enriched in sequences encoding chloroplast proteins involved in cell protection against the damaging effect of ROS. Among the early fine-tuned components, are proteins involved in tetrapyrrole biosynthesis, chlorophyll catabolism, protein import, folding and turnover, synthesis and membrane insertion of photosynthetic subunits. Up to now, the flu mutant was the only biological system adopted to define the regulation of gene expression by 1O2. In this work, we propose the use of mutants accumulating 1O2 by mechanisms different from those activated in flu to better identify ROS signalling. Conclusions We propose that the lack of zeaxanthin and lutein leads to 1O2 accumulation and this represents a signalling pathway in the early stages of stress acclimation, beside the response to ADP/ATP ratio and to the redox state of both plastoquinone pool. Chloroplasts respond to 1O2 accumulation by undergoing a significant change in composition and function towards a fast acclimatory response. The physiological implications of this signalling specificity are discussed. PMID:21481232

  17. 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...

  18. A Collection of Ds Insertional Mutants Associated With Defects in Male Gametophyte Development and Function in Arabidopsis thaliana

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Functional analyses of the Arabidopsis genome require analysis of the gametophytic generation, since 10% of the genes are expressed in the male gametophyte and 9% in the female gametophyte. Here we describe the genetic and molecular characterization of 67 Ds insertion lines that show reduced transmi...

  19. Decrease in Leaf Sucrose Synthesis Leads to Increased Leaf Starch Turnover and Decreased RuBP-limited Photosynthesis But Not Rubisco-limited Photosynthesis in Arabidopsis Null Mutants of SPSA1

    Technology Transfer Automated Retrieval System (TEKTRAN)

    SPS (Sucrose phosphate synthase) isoforms from dicots cluster into families A, B and C. In this study, we investigated the individual effect of null mutations of each of the four SPS genes in Arabidopsis (spsa1, spsa2, spsb and spsc) on photosynthesis and carbon partitioning. Null mutants spsa1 and ...

  20. 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.

  1. Characterization of a NADH-Dependent Glutamate Dehydrogenase Mutant of Arabidopsis Demonstrates the Key Role of this Enzyme in Root Carbon and Nitrogen Metabolism[W

    PubMed Central

    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

    2012-01-01

    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. PMID:23054470

  2. 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. PMID:23054470

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

    PubMed Central

    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. PMID:9165751

  4. Expression of Escherichia coli glycogen branching enzyme in an Arabidopsis mutant devoid of endogenous starch branching enzymes induces the synthesis of starch-like polyglucans.

    PubMed

    Boyer, Laura; Roussel, Xavier; Courseaux, Adeline; Ndjindji, Ofilia M; Lancelon-Pin, Christine; Putaux, Jean-Luc; Tetlow, Ian J; Emes, Michael J; Pontoire, Bruno; D' Hulst, Christophe; Wattebled, Fabrice

    2016-07-01

    Starch synthesis requires several enzymatic activities including branching enzymes (BEs) responsible for the formation of α(1 → 6) linkages. Distribution and number of these linkages are further controlled by debranching enzymes that cleave some of them, rendering the polyglucan water-insoluble and semi-crystalline. Although the activity of BEs and debranching enzymes is mandatory to sustain normal starch synthesis, the relative importance of each in the establishment of the plant storage polyglucan (i.e. water insolubility, crystallinity and presence of amylose) is still debated. Here, we have substituted the activity of BEs in Arabidopsis with that of the Escherichia coli glycogen BE (GlgB). The latter is the BE counterpart in the metabolism of glycogen, a highly branched water-soluble and amorphous storage polyglucan. GlgB was expressed in the be2 be3 double mutant of Arabidopsis, which is devoid of BE activity and consequently free of starch. The synthesis of a water-insoluble, partly crystalline, amylose-containing starch-like polyglucan was restored in GlgB-expressing plants, suggesting that BEs' origin only has a limited impact on establishing essential characteristics of starch. Moreover, the balance between branching and debranching is crucial for the synthesis of starch, as an excess of branching activity results in the formation of highly branched, water-soluble, poorly crystalline polyglucan. PMID:26715025

  5. Ectopic expression of FaesAP3, a Fagopyrum esculentum (Polygonaceae) AP3 orthologous gene rescues stamen development in an Arabidopsis ap3 mutant.

    PubMed

    Fang, Zheng-wu; Qi, Rui; Li, Xiao-fang; Liu, Zhi-xiong

    2014-10-25

    Arabidopsis thaliana APETALA3 (AP3) and Antirrhinum majus DEFICIENS (DEF) MADS box genes are required to specify petal and stamen identity. AP3 and DEF are members of the euAP3 lineage, which arose by gene duplication coincident with radiation of the core eudicots. In order to investigate the molecular mechanisms underlying organ development in early diverging clades of core eudicots, we isolated and identified an AP3 homolog, FaesAP3, from Fagopyrum esculentum (buckwheat, Polygonaceae), a multi-food-use pseudocereal with healing benefits. Protein sequence alignment and phylogenetic analyses revealed that FaesAP3 grouped into the euAP3 lineage. Expression analysis showed that FaesAP3 was transcribed only in developing stamens, and differed from AP3 and DEF, which expressed in developing petals and stamens. Moreover, ectopic expression of FaesAP3 rescued stamen development without complementation of petal development in an Arabidopsis ap3 mutant. Our results suggest that FaesAP3 is involved in the development of stamens in buckwheat. These results also suggest that FaesAP3 holds some potential for biotechnical engineering to create a male sterile line of F. esculentum. PMID:25149019

  6. Expression of SofLAC, a new laccase in sugarcane, restores lignin content but not S:G ratio of Arabidopsis lac17 mutant.

    PubMed

    Cesarino, Igor; Araújo, Pedro; Sampaio Mayer, Juliana Lischka; Vicentini, Renato; Berthet, Serge; Demedts, Brecht; Vanholme, Bartel; Boerjan, Wout; Mazzafera, Paulo

    2013-04-01

    Lignin is a complex phenolic heteropolymer deposited in the secondarily thickened walls of specialized plant cells to provide strength for plants to stand upright and hydrophobicity to conducting cells for long-distance water transport. Although essential for plant growth and development, lignin is the major plant cell-wall component responsible for biomass recalcitrance to industrial processing. Peroxidases and laccases are generally thought to be responsible for lignin polymerization, but, given their broad substrate specificities and large gene families, specific isoforms involved in lignification are difficult to identify. This study used a combination of co-expression analysis, tissue/cell-type-specific expression analysis, and genetic complementation to correlate a sugarcane laccase gene, SofLAC, to the lignification process. A co-expression network constructed from 37 cDNA libraries showed that SofLAC was coordinately expressed with several phenylpropanoid biosynthesis genes. Tissue-specific expression analysis by quantitative RT-PCR showed that SofLAC was expressed preferentially in young internodes and that expression levels decrease with stem maturity. Cell-type-specific expression analysis by in situ hybridization demonstrated the localization of SofLAC mRNA in lignifying cell types, mainly in inner and outer portions of sclerenchymatic bundle sheaths. To investigate whether SofLAC is able to oxidize monolignols during lignification, the Arabidopsis lac17 mutant, which has reduced lignin levels, was complemented by expressing SofLAC under the control of the Arabidopsis AtLAC17 promoter. The expression of SofLAC restored the lignin content but not the lignin composition in complemented lac17 mutant lines. Taken together, these results suggest that SofLAC participates in lignification in sugarcane. PMID:23418623

  7. The Reaumuria trigyna leucoanthocyanidin dioxygenase (RtLDOX) gene complements anthocyanidin synthesis and increases the salt tolerance potential of a transgenic Arabidopsis LDOX mutant.

    PubMed

    Zhang, Huirong; Du, Chao; Wang, Yan; Wang, Jia; Zheng, Linlin; Wang, Yingchun

    2016-09-01

    Reaumuria trigyna is a typical, native desert halophyte that grows under extreme conditions in Inner Mongolia. In a previous transcriptomic profiling analysis, flavonoid pathway-related genes in R. trigyna showed significant differences in transcript abundance under salt stress. Leucoanthocyanidin dioxygenase (LDOX, EC 1.14.11.19) is one of three dioxygenases in the flavonoid pathway that catalyzes the formation of anthocyanidins from leucoanthocyanidins. In this study, we cloned the full-length cDNA of R. trigyna LDOX (RtLDOX), and found RtLDOX recombinant protein was able to replace flavanone-3-hydroxylase (F3H, EC 1.14.11.9), another dioxygenase in the flavonoid pathway, to convert naringenin to dihydrokaempferol in vitro. R. trigyna LDOX can complement the Arabidopsis LDOX mutant transparent testa11 (tt11-11), which has reduced proanthocyanin (PA) and anthocyanin levels in seeds, to accumulate these two compounds. Thus, RtLDOX acts as a multifunctional dioxygenase to effect the synthesis of PA and anthocyanins and can perform F3H dioxygenase activities in the flavonoid biosynthesis pathway. The RtLDOX promoter harbored many cis-acting elements that might be recognized and bound by transcription factors related to stress response. RtLDOX expression was strongly increased under salt stress, and RtLDOX transgenic Arabidopsis mutant under NaCl stress accumulated the content of flavonoids leading to an increased antioxidant activities and plant biomass. These results suggest that RtLDOX as a multifunctional dioxygenase in flavonoid biosynthesis involves in enhancing plant response to NaCl stress. PMID:27219053

  8. The Arabidopsis szl1 Mutant Reveals a Critical Role of β-Carotene in Photosystem I Photoprotection1[C][W

    PubMed Central

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

    2012-01-01

    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. PMID:23029671

  9. A genetic screen for mutants defective in IAA1-LUC degradation in Arabidopsis thaliana reveals an important requirement for TOPOISOMERASE6B in auxin physiology

    PubMed Central

    Gilkerson, Jonathan; Callis, Judy

    2014-01-01

    Many plant growth and developmental processes are modulated by the hormone auxin. Auxin-modulated proteolysis of Aux/IAAs, a family of transcriptional repressors, represents a major mode of auxin action. Auxin facilitates the interaction of Aux/IAAs with TIR1/AFB F-box proteins, promoting their ubiquitination by the SCFTIR1/AFB ubiquitin E3 ligase leading to subsequent degradation by the 26S proteasome. To identify new genes regulating Aux/IAA proteolysis in Arabidopsis thaliana, we took a genetic approach, identifying individuals with altered degradation of an IAA1-luciferase fusion protein (IAA1-LUC). A mutant with 2-fold slower IAA1-LUC degradation rate compared with wild-type was isolated. Positional cloning identified the mutant as an allele of TOPOISOMERASE6B, named top6b-7. TOP6B encodes a subunit of a plant and archea-specific enzyme regulating endoreduplication, DNA damage repair and transcription in plants. T-DNA insertion alleles (top6b-8 and top6b-9) were also analyzed. top6b-7 seedlings are less sensitive to exogenous auxin than wild-type siblings in primary root growth assays, and experiments with DR5:GUS. Additionally, top6b-7 seedlings have a 40% reduction in the amount of endogenous IAA. These data suggest that increased IAA1-LUC half-life in top6b-7 probably results from a combination of both lower endogenous IAA levels and reduced sensitivity to auxin. PMID:25482814

  10. 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

  11. Mutations in Ribosomal Proteins, RPL4 and RACK1, Suppress the Phenotype of a Thermospermine-Deficient Mutant of Arabidopsis thaliana

    PubMed Central

    Kakehi, Jun-Ichi; Kawano, Eri; Yoshimoto, Kaori; Cai, Qingqing; Imai, Akihiro; Takahashi, Taku

    2015-01-01

    Thermospermine acts in negative regulation of xylem differentiation and its deficient mutant of Arabidopsis thaliana, acaulis5 (acl5), shows excessive xylem formation and severe dwarfism. Studies of two dominant suppressors of acl5, sac51-d and sac52-d, have revealed that SAC51 and SAC52 encode a transcription factor and a ribosomal protein L10 (RPL10), respectively, and these mutations enhance translation of the SAC51 mRNA, which contains conserved upstream open reading frames in the 5’ leader. Here we report identification of SAC53 and SAC56 responsible for additional suppressors of acl5. sac53-d is a semi-dominant allele of the gene encoding a receptor for activated C kinase 1 (RACK1) homolog, a component of the 40S ribosomal subunit. sac56-d represents a semi-dominant allele of the gene for RPL4. We show that the GUS reporter activity driven by the CaMV 35S promoter plus the SAC51 5’ leader is reduced in acl5 and restored by sac52-d, sac53-d, and sac56-d as well as thermospermine. Furthermore, the SAC51 mRNA, which may be a target of nonsense-mediated mRNA decay, was found to be stabilized in these ribosomal mutants and by thermospermine. These ribosomal proteins are suggested to act in the control of uORF-mediated translation repression of SAC51, which is derepressed by thermospermine. PMID:25625317

  12. The Arabidopsis nox Mutant Lacking Carotene Hydroxylase Activity Reveals a Critical Role for Xanthophylls in Photosystem I Biogenesis[C][W

    PubMed Central

    Dall’Osto, Luca; Piques, Maria; Ronzani, Michela; Molesini, Barbara; Alboresi, Alessandro; Cazzaniga, Stefano; Bassi, Roberto

    2013-01-01

    Carotenes, and their oxygenated derivatives xanthophylls, are essential components of the photosynthetic apparatus. They contribute to the assembly of photosynthetic complexes and participate in light absorption and chloroplast photoprotection. Here, we studied the role of xanthophylls, as distinct from that of carotenes, by characterizing a no xanthophylls (nox) mutant of Arabidopsis thaliana, which was obtained by combining mutations targeting the four carotenoid hydroxylase genes. nox plants retained α- and β-carotenes but were devoid in xanthophylls. The phenotype included depletion of light-harvesting complex (LHC) subunits and impairment of nonphotochemical quenching, two effects consistent with the location of xanthophylls in photosystem II antenna, but also a decreased efficiency of photosynthetic electron transfer, photosensitivity, and lethality in soil. Biochemical analysis revealed that the nox mutant was specifically depleted in photosystem I function due to a severe deficiency in PsaA/B subunits. While the stationary level of psaA/B transcripts showed no major differences between genotypes, the stability of newly synthesized PsaA/B proteins was decreased and translation of psaA/B mRNA was impaired in nox with respect to wild-type plants. We conclude that xanthophylls, besides their role in photoprotection and LHC assembly, are also needed for photosystem I core translation and stability, thus making these compounds indispensable for autotrophic growth. PMID:23396829

  13. Arabidopsis Mutants Deleted in the Light-Harvesting Protein Lhcb4 Have a Disrupted Photosystem II Macrostructure and Are Defective in Photoprotection[C][W

    PubMed Central

    de Bianchi, Silvia; Betterle, Nico; Kouril, Roman; Cazzaniga, Stefano; Boekema, Egbert; Bassi, Roberto; Dall’Osto, Luca

    2011-01-01

    The role of the light-harvesting complex Lhcb4 (CP29) in photosynthesis was investigated in Arabidopsis thaliana by characterizing knockout lines for each of the three Lhcb4 isoforms (Lhcb4.1/4.2/4.3). Plants lacking all isoforms (koLhcb4) showed a compensatory increase of Lhcb1 and a slightly reduced photosystem II/I ratio with respect to the wild type. The absence of Lhcb4 did not result in alteration in electron transport rates. However, the kinetic of state transition was faster in the mutant, and nonphotochemical quenching activity was lower in koLhcb4 plants with respect to either wild type or mutants retaining a single Lhcb4 isoform. KoLhcb4 plants were more sensitive to photoinhibition, while this effect was not observed in knockout lines for any other photosystem II antenna subunit. Ultrastructural analysis of thylakoid grana membranes showed a lower density of photosystem II complexes in koLhcb4. Moreover, analysis of isolated supercomplexes showed a different overall shape of the C2S2 particles due to a different binding mode of the S-trimer to the core complex. An empty space was observed within the photosystem II supercomplex at the Lhcb4 position, implying that the missing Lhcb4 was not replaced by other Lhc subunits. This suggests that Lhcb4 is unique among photosystem II antenna proteins and determinant for photosystem II macro-organization and photoprotection. PMID:21803939

  14. Arabidopsis mutants deleted in the light-harvesting protein Lhcb4 have a disrupted photosystem II macrostructure and are defective in photoprotection.

    PubMed

    de Bianchi, Silvia; Betterle, Nico; Kouril, Roman; Cazzaniga, Stefano; Boekema, Egbert; Bassi, Roberto; Dall'Osto, Luca

    2011-07-01

    The role of the light-harvesting complex Lhcb4 (CP29) in photosynthesis was investigated in Arabidopsis thaliana by characterizing knockout lines for each of the three Lhcb4 isoforms (Lhcb4.1/4.2/4.3). Plants lacking all isoforms (koLhcb4) showed a compensatory increase of Lhcb1 and a slightly reduced photosystem II/I ratio with respect to the wild type. The absence of Lhcb4 did not result in alteration in electron transport rates. However, the kinetic of state transition was faster in the mutant, and nonphotochemical quenching activity was lower in koLhcb4 plants with respect to either wild type or mutants retaining a single Lhcb4 isoform. KoLhcb4 plants were more sensitive to photoinhibition, while this effect was not observed in knockout lines for any other photosystem II antenna subunit. Ultrastructural analysis of thylakoid grana membranes showed a lower density of photosystem II complexes in koLhcb4. Moreover, analysis of isolated supercomplexes showed a different overall shape of the C₂S₂ particles due to a different binding mode of the S-trimer to the core complex. An empty space was observed within the photosystem II supercomplex at the Lhcb4 position, implying that the missing Lhcb4 was not replaced by other Lhc subunits. This suggests that Lhcb4 is unique among photosystem II antenna proteins and determinant for photosystem II macro-organization and photoprotection. PMID:21803939

  15. Light-dependent gravitropism and negative phototropism of inflorescence stems in a dominant Aux/IAA mutant of Arabidopsis thaliana, axr2.

    PubMed

    Sato, Atsuko; Sasaki, Shu; Matsuzaki, Jun; Yamamoto, Kotaro T

    2014-09-01

    Gravitropism and phototropism of the primary inflorescence stems were examined in a dominant Aux/IAA mutant of Arabidopsis, axr2/iaa7, which did not display either tropism in hypocotyls. axr2-1 stems completely lacked gravitropism in the dark but slowly regained it in light condition. Though wild-type stems showed positive phototropism, axr2 stems displayed negative phototropism with essentially the same light fluence-response curve as the wild type (WT). Application of 1-naphthaleneacetic acid-containing lanolin to the stem tips enhanced the positive phototropism of WT, and reduced the negative phototropism of axr2. Decapitation of stems caused a small negative phototropism in WT, but did not affect the negative phototropism of axr2. p-glycoprotein 1 (pgp1) pgp19 double mutants showed no phototropism, while decapitated double mutants exhibited negative phototropism. Expression of auxin-responsive IAA14/SLR, IAA19/MSG2 and SAUR50 genes was reduced in axr2 and pgp1 pgp19 stems relative to that of WT. These suggest that the phototropic response of stem is proportional to the auxin supply from the shoot apex, and that negative phototropism may be a basal response to unilateral blue-light irradiation when the levels of auxin or auxin signaling are reduced to the minimal level in the primary stems. In contrast, all of these treatments reduced or did not affect gravitropism in wild-type or axr2 stems. Tropic responses of the transgenic lines that expressed axr2-1 protein by the endodermis-specific promoter suggest that AXR2-dependent auxin response in the endodermis plays a more crucial role in gravitropism than in phototropism in stems but no significant roles in either tropism in hypocotyls. PMID:24938853

  16. Phytoalexin-Deficient Mutants of Arabidopsis Reveal That Pad4 Encodes a Regulatory Factor and That Four Pad Genes Contribute to Downy Mildew Resistance

    PubMed Central

    Glazebrook, J.; Zook, M.; Mert, F.; Kagan, I.; Rogers, E. E.; Crute, I. R.; Holub, E. B.; Hammerschmidt, R.; Ausubel, F. M.

    1997-01-01

    We are working to determine the role of the Arabidopsis phytoalexin, camalexin, in protecting the plant from pathogen attack by isolating phytoalexin-deficient (pad) mutants in the accession Columbia (Col-0) and examining their response to pathogens. Mutations in PAD1, PAD2, and PAD4 caused enhanced susceptibility to the bacterial pathogen Pseudomonas syringae pv. maculicola strain ES4326 (PsmES4326), while mutations in PAD3 or PAD5 did not. Camalexin was not detected in any of the double mutants pad1-1 pad2-1, pad1-1 pad3-1 or pad2-1 pad3-1. Growth of PsmES4326 in pad1-1 pad2-1 was greater than that in pad1-1 or pad2-1 plants, while growth in pad1-1 pad3-1 and pad2-1 pad3-1 plants was similar to that in pad1-1 and pad2-1 plants, respectively. The pad4-1 mutation caused reduced camalexin synthesis in response to PsmES4326 infection, but not in response to Cochliobolus carbonum infection, indicating that PAD4 has a regulatory function. PAD1, PAD2, PAD3 and PAD4 are all required for resistance to the eukaryotic biotroph Peronospora parasitica. The pad4-1 mutation caused the most dramatic change, exhibiting full susceptibility to four of six Col-incompatible parasite isolates. Interestingly, each combination of double mutants between pad1-1, pad2-1 and pad3-1 exhibited additive shifts to moderate or full susceptibility to most of the isolates. PMID:9136026

  17. Leaves of the Arabidopsis maltose exporter1 Mutant Exhibit a Metabolic Profile with Features of Cold Acclimation in the Warm

    PubMed Central

    Purdy, Sarah J.; Bussell, John D.; Nunn, Christopher P.; Smith, Steven M.

    2013-01-01

    Background Arabidopsis plants accumulate maltose from starch breakdown during cold acclimation. The Arabidopsis mutant, maltose excess1-1, accumulates large amounts of maltose in the plastid even in the warm, due to a deficient plastid envelope maltose transporter. We therefore investigated whether the elevated maltose level in mex1-1 in the warm could result in changes in metabolism and physiology typical of WT plants grown in the cold. Principal Findings Grown at 21 °C, mex1-1 plants were much smaller, with fewer leaves, and elevated carbohydrates and amino acids compared to WT. However, after transfer to 4 °C the total soluble sugar pool and amino acid concentration was in equal abundance in both genotypes, although the most abundant sugar in mex1-1 was still maltose whereas sucrose was in greatest abundance in WT. The chlorophyll a/b ratio in WT was much lower in the cold than in the warm, but in mex1-1 it was low in both warm and cold. After prolonged growth at 4 °C, the shoot biomass, rosette diameter and number of leaves at bolting were similar in mex1-1 and WT. Conclusions The mex1-1 mutation in warm-grown plants confers aspects of cold acclimation, including elevated levels of sugars and amino acids and low chlorophyll a/b ratio. This may in turn compromise growth of mex1-1 in the warm relative to WT. We suggest that elevated maltose in the plastid could be responsible for key aspects of cold acclimation. PMID:24223944

  18. 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. PMID:27001427

  19. Abscisic Acid–Responsive Guard Cell Metabolomes of Arabidopsis Wild-Type and gpa1 G-Protein Mutants[C][W

    PubMed Central

    Jin, Xiaofen; Wang, Rui-Sheng; Zhu, Mengmeng; Jeon, Byeong Wook; Albert, Reka; Chen, Sixue; Assmann, Sarah M.

    2013-01-01

    Individual metabolites have been implicated in abscisic acid (ABA) signaling in guard cells, but a metabolite profile of this specialized cell type is lacking. We used liquid chromatography–multiple reaction monitoring mass spectrometry for targeted analysis of 85 signaling-related metabolites in Arabidopsis thaliana guard cell protoplasts over a time course of ABA treatment. The analysis utilized ∼350 million guard cell protoplasts from ∼30,000 plants of the Arabidopsis Columbia accession (Col) wild type and the heterotrimeric G-protein α subunit mutant, gpa1, which has ABA-hyposensitive stomata. These metabolomes revealed coordinated regulation of signaling metabolites in unrelated biochemical pathways. Metabolites clustered into different temporal modules in Col versus gpa1, with fewer metabolites showing ABA-altered profiles in gpa1. Ca2+-mobilizing agents sphingosine-1-phosphate and cyclic adenosine diphosphate ribose exhibited weaker ABA-stimulated increases in gpa1. Hormone metabolites were responsive to ABA, with generally greater responsiveness in Col than in gpa1. Most hormones also showed different ABA responses in guard cell versus mesophyll cell metabolomes. These findings suggest that ABA functions upstream to regulate other hormones, and are also consistent with G proteins modulating multiple hormonal signaling pathways. In particular, indole-3-acetic acid levels declined after ABA treatment in Col but not gpa1 guard cells. Consistent with this observation, the auxin antagonist α-(phenyl ethyl-2-one)-indole-3-acetic acid enhanced ABA-regulated stomatal movement and restored partial ABA sensitivity to gpa1. PMID:24368793

  20. Arabidopsis tic62 trol mutant lacking thylakoid-bound ferredoxin-NADP+ oxidoreductase shows distinct metabolic phenotype.

    PubMed

    Lintala, Minna; Schuck, Natalie; Thormählen, Ina; Jungfer, Andreas; Weber, Katrin L; Weber, Andreas P M; Geigenberger, Peter; Soll, Jürgen; Bölter, Bettina; Mulo, Paula

    2014-01-01

    Ferredoxin-NADP+ oxidoreductase (FNR), functioning in the last step of the photosynthetic electron transfer chain, exists both as a soluble protein in the chloroplast stroma and tightly attached to chloroplast membranes. Surface plasmon resonance assays showed that the two FNR isoforms, LFNR1 and LFNR2, are bound to the thylakoid membrane via the C-terminal domains of Tic62 and TROL proteins in a pH-dependent manner. The tic62 trol double mutants contained a reduced level of FNR, exclusively found in the soluble stroma. Although the mutant plants showed no visual phenotype or defects in the function of photosystems under any conditions studied, a low ratio of NADPH/NADP+ was detected. Since the CO₂ fixation capacity did not differ between the tic62 trol plants and wild-type, it seems that the plants are able to funnel reducing power to most crucial reactions to ensure survival and fitness of the plants. However, the activity of malate dehydrogenase was down-regulated in the mutant plants. Apparently, the plastid metabolism is able to cope with substantial changes in directing the electrons from the light reactions to stromal metabolism and thus only few differences are visible in steady-state metabolite pool sizes of the tic62 trol plants. PMID:24043709

  1. Isolation of the GA-response mutant sly1 as a suppressor of ABI1-1 in Arabidopsis thaliana.

    PubMed Central

    Steber, C M; Cooney, S E; McCourt, P

    1998-01-01

    Seed dormancy and germination in higher plants are partially controlled by the plant hormones abscisic acid (ABA) and gibberellic acid (GA). ABA establishes dormancy during embryo maturation, whereas GA breaks dormancy and induces germination. Previous attempts to identify GA response genes were confounded because GA mutants are not expected to germinate and, unlike GA auxotrophs, should fail to be rescued by exogenous GA. Here, we describe a screen for suppressors of the ABA-insensitive mutant ABI1-1 that enriches for GA auxotrophs and GA-insensitive mutants. The vast majority (76%) of the suppressors of ABI1-1 strongly resemble GA auxotrophs in that they are severely dwarfed and have dark green foliage and flowers with underdeveloped petals and stamen. Three isolates were alleles of the GA auxotroph ga1. The remaining severe dwarves were not rescued by GA and belong to a single complementation group that we designate sly1 (Sleepy 1). The alleles of sly1 identified are the first recessive GA-insensitive mutations to reflect the full spectrum of GA-associated phenotypes, including the failure to germinate in the absence of the ABI1-1 lesion. Thus, we postulate that SLY1 is a key factor in GA reception. PMID:9611170

  2. The Arabidopsis aba4-1 mutant reveals a specific function for neoxanthin in protection against photooxidative stress.

    PubMed

    Dall'Osto, Luca; Cazzaniga, Stefano; North, Helen; Marion-Poll, Annie; Bassi, Roberto

    2007-03-01

    The aba4-1 mutant completely lacks neoxanthin but retains all other xanthophyll species. The missing neoxanthin in light-harvesting complex (Lhc) proteins is compensated for by higher levels of violaxanthin, albeit with lower capacity for photoprotection compared with proteins with wild-type levels of neoxanthin. Detached leaves of aba4-1 were more sensitive to oxidative stress than the wild type when exposed to high light and incubated in a solution of photosensitizer agents. Both treatments caused more rapid pigment bleaching and lipid oxidation in aba4-1 than wild-type plants, suggesting that neoxanthin acts as an antioxidant within the photosystem II (PSII) supercomplex in thylakoids. While neoxanthin-depleted Lhc proteins and leaves had similar sensitivity as the wild type to hydrogen peroxide and singlet oxygen, they were more sensitive to superoxide anions. aba4-1 intact plants were not more sensitive than the wild type to high-light stress, indicating the existence of compensatory mechanisms of photoprotection involving the accumulation of zeaxanthin. However, the aba4-1 npq1 double mutant, lacking zeaxanthin and neoxanthin, underwent stronger PSII photoinhibition and more extensive oxidation of pigments than the npq1 mutant, which still contains neoxanthin. We conclude that neoxanthin preserves PSII from photoinactivation and protects membrane lipids from photooxidation by reactive oxygen species. Neoxanthin appears particularly active against superoxide anions produced by the Mehler's reaction, whose rate is known to be enhanced in abiotic stress conditions. PMID:17351115

  3. The Arabidopsis aba4-1 Mutant Reveals a Specific Function for Neoxanthin in Protection against Photooxidative Stress[W

    PubMed Central

    Dall'Osto, Luca; Cazzaniga, Stefano; North, Helen; Marion-Poll, Annie; Bassi, Roberto

    2007-01-01

    The aba4-1 mutant completely lacks neoxanthin but retains all other xanthophyll species. The missing neoxanthin in light-harvesting complex (Lhc) proteins is compensated for by higher levels of violaxanthin, albeit with lower capacity for photoprotection compared with proteins with wild-type levels of neoxanthin. Detached leaves of aba4-1 were more sensitive to oxidative stress than the wild type when exposed to high light and incubated in a solution of photosensitizer agents. Both treatments caused more rapid pigment bleaching and lipid oxidation in aba4-1 than wild-type plants, suggesting that neoxanthin acts as an antioxidant within the photosystem II (PSII) supercomplex in thylakoids. While neoxanthin-depleted Lhc proteins and leaves had similar sensitivity as the wild type to hydrogen peroxide and singlet oxygen, they were more sensitive to superoxide anions. aba4-1 intact plants were not more sensitive than the wild type to high-light stress, indicating the existence of compensatory mechanisms of photoprotection involving the accumulation of zeaxanthin. However, the aba4-1 npq1 double mutant, lacking zeaxanthin and neoxanthin, underwent stronger PSII photoinhibition and more extensive oxidation of pigments than the npq1 mutant, which still contains neoxanthin. We conclude that neoxanthin preserves PSII from photoinactivation and protects membrane lipids from photooxidation by reactive oxygen species. Neoxanthin appears particularly active against superoxide anions produced by the Mehler's reaction, whose rate is known to be enhanced in abiotic stress conditions. PMID:17351115

  4. 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

  5. RiceFOX: A Database of Arabidopsis Mutant Lines Overexpressing Rice Full-Length cDNA that Contains a Wide Range of Trait Information to Facilitate Analysis of Gene Function

    PubMed Central

    Sakurai, Tetsuya; Kondou, Youichi; Akiyama, Kenji; Kurotani, Atsushi; Higuchi, Mieko; Ichikawa, Takanari; Kuroda, Hirofumi; Kusano, Miyako; Mori, Masaki; Saitou, Tsutomu; Sakakibara, Hitoshi; Sugano, Shoji; Suzuki, Makoto; Takahashi, Hideki; Takahashi, Shinya; Takatsuji, Hiroshi; Yokotani, Naoki; Yoshizumi, Takeshi; Saito, Kazuki; Shinozaki, Kazuo; Oda, Kenji; Hirochika, Hirohiko; Matsui, Minami

    2011-01-01

    Identification of gene function is important not only for basic research but also for applied science, especially with regard to improvements in crop production. For rapid and efficient elucidation of useful traits, we developed a system named FOX hunting (Full-length cDNA Over-eXpressor gene hunting) using full-length cDNAs (fl-cDNAs). A heterologous expression approach provides a solution for the high-throughput characterization of gene functions in agricultural plant species. Since fl-cDNAs contain all the information of functional mRNAs and proteins, we introduced rice fl-cDNAs into Arabidopsis plants for systematic gain-of-function mutation. We generated >30,000 independent Arabidopsis transgenic lines expressing rice fl-cDNAs (rice FOX Arabidopsis mutant lines). These rice FOX Arabidopsis lines were screened systematically for various criteria such as morphology, photosynthesis, UV resistance, element composition, plant hormone profile, metabolite profile/fingerprinting, bacterial resistance, and heat and salt tolerance. The information obtained from these screenings was compiled into a database named ‘RiceFOX’. This database contains around 18,000 records of rice FOX Arabidopsis lines and allows users to search against all the observed results, ranging from morphological to invisible traits. The number of searchable items is approximately 100; moreover, the rice FOX Arabidopsis lines can be searched by rice and Arabidopsis gene/protein identifiers, sequence similarity to the introduced rice fl-cDNA and traits. The RiceFOX database is available at http://ricefox.psc.riken.jp/. PMID:21186176

  6. Suspensor-derived polyembryony caused by altered expression of valyl-tRNA synthetase in the twn2 mutant of Arabidopsis

    PubMed Central

    Zhang, James Z.; Somerville, Chris R.

    1997-01-01

    The twn2 mutant of Arabidopsis exhibits a defect in early embryogenesis where, following one or two divisions of the zygote, the decendents of the apical cell arrest. The basal cells that normally give rise to the suspensor proliferate abnormally, giving rise to multiple embryos. A high proportion of the seeds fail to develop viable embryos, and those that do, contain a high proportion of partially or completely duplicated embryos. The adult plants are smaller and less vigorous than the wild type and have a severely stunted root. The twn2-1 mutation, which is the only known allele, was caused by a T-DNA insertion in the 5′ untranslated region of a putative valyl-tRNA synthetase gene, valRS. The insertion causes reduced transcription of the valRS gene in reproductive tissues and developing seeds but increased expression in leaves. Analysis of transcript initiation sites and the expression of promoter–reporter fusions in transgenic plants indicated that enhancer elements inside the first two introns interact with the border of the T-DNA to cause the altered pattern of expression of the valRS gene in the twn2 mutant. The phenotypic consequences of this unique mutation are interpreted in the context of a model, suggested by Vernon and Meinke [Vernon, D. M. & Meinke, D. W. (1994) Dev. Biol. 165, 566–573], in which the apical cell and its decendents normally suppress the embryogenic potential of the basal cell and its decendents during early embryo development. PMID:9207094

  7. 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

  8. Action spectrum of photoinhibition in leaves of wild type and npq1-2 and npq4-1 mutants of Arabidopsis thaliana.

    PubMed

    Sarvikas, Päivi; Hakala, Marja; Pätsikkä, Eija; Tyystjärvi, Taina; Tyystjärvi, Esa

    2006-03-01

    Photoinhibition is light-induced inactivation of PSII. Hypotheses about the photoreceptor(s) of photoinhibition include the Chl antenna of PSII, manganese of the oxygen-evolving complex (OEC), uncoupled Chl and iron-sulfur centres. We measured the action spectrum of photoinhibition in vivo from wild-type Arabidopsis thaliana L. and from the npq1-2 and npq4-1 mutants defective in non-photochemical quenching (NPQ) of excitations of the PSII antenna. The in vivo action spectrum was found to resemble closely the in vitro action spectra published for photoinhibition. We compared the action spectrum with absorbance spectra of model compounds of the OEC complex and other potential photoreceptors of photoinhibition. The comparison suggests that both manganese and Chl function as photoreceptors in photoinhibition. In accordance with the function of two types of photoreceptors in photoinhibition, NPQ was found to offer only partial protection against photoinhibition at visible wavelengths. The low protective efficiency of NPQ supports the conclusion that the Chl antenna of PSII is not the only photoreceptor of photoinhibition. Comparison of the action spectrum of photoinhibition with the emission spectrum of sunlight shows that the UV part of sunlight is responsible for the major part of photoinhibition under natural conditions. PMID:16415063

  9. Isolation of an Arabidopsis thaliana gene encoding cycloartenol synthase by functional expression in a yeast mutant lacking lanosterol synthase by the use of a chromatographic screen.

    PubMed

    Corey, E J; Matsuda, S P; Bartel, B

    1993-12-15

    Whereas vertebrates and fungi synthesize sterols from epoxysqualene through the intermediate lanosterol, plants cyclize epoxysqualene to cycloartenol as the initial sterol. We report the cloning and characterization of CAS1, an Arabidopsis thaliana gene encoding cycloartenol synthase [(S)-2,3-epoxysqualene mutase (cyclizing, cycloartenol forming), EC 5.4.99.8]. A yeast mutant lacking lanosterol synthase [(S)-2,3-epoxysqualene mutase (cyclizing, lanosterol forming), EC 5.4.99.7] was transformed with an A. thaliana cDNA yeast expression library, and colonies were assayed for epoxysqualene mutase activity by thin-layer chromatography. One out of approximately 10,000 transformants produced a homogenate that cyclized 2,3-epoxysqualene to the plant sterol cycloartenol. This activity was shown to be plasmid dependent. The plasmid insert contains a 2277-bp open reading frame capable of encoding an 86-kDa protein with significant homology to lanosterol synthase from Candida albicans and squalene-hopene cyclase (EC 5.4.99.-) from Bacillus acidocalcarius. The method used to clone this gene should be generally applicable to genes responsible for secondary metabolite biosynthesis. PMID:7505443

  10. 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

    Anderson, Nickolas A; Tobimatsu, Yuki; Ciesielski, Peter N; Ximenes, Eduardo; Ralph, John; Donohoe, Bryon S; Ladisch, Michael; Chapple, Clint

    2015-08-01

    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

  11. The gain-of-function Arabidopsis acd6 mutant reveals novel regulation and function of the salicylic acid signaling pathway in controlling cell death, defenses, and cell growth.

    PubMed Central

    Rate, D N; Cuenca, J V; Bowman, G R; Guttman, D S; Greenberg, J T

    1999-01-01

    We isolated a dominant gain-of-function Arabidopsis mutant, accelerated cell death 6 (acd6), with elevated defenses, patches of dead and enlarged cells, reduced stature, and increased resistance to Pseudomonas syringae. The acd6-conferred phenotypes are suppressed by removing a key signaling molecule, salicylic acid (SA), by using the nahG transgene, which encodes SA hydroxylase. This suppression includes phenotypes that are not induced by application of SA to wild-type plants, indicating that SA acts with a second signal to cause many acd6-conferred phenotypes. acd6-nahG plants show hyperactivation of all acd6-conferred phenotypes after treatment with a synthetic inducer of the SA pathway, benzo(1,2, 3)thiadiazole-7-carbothioic acid (BTH), suggesting that SA acts with and also modulates the levels and/or activity of the second defense signal. acd6 acts partially through a NONEXPRESSOR OF PR 1 (NPR1) gene-independent pathway that activates defenses and confers resistance to P. syringae. Surprisingly, BTH-treated acd6-nahG plants develop many tumor-like abnormal growths, indicating a possible role for SA in modulating cell growth. PMID:10488236

  12. Overexpression of FAB1A-GFP recruits SNX2b on the endosome membrane in snx1–1 mutant in Arabidopsis

    PubMed Central

    Hirano, Tomoko; Sato, Masa H.

    2016-01-01

    Phosphatidylinositol 3,5-bisphosphate [PI(3,5)P2] is one of the phosphoinositides that controls endosomal trafficking events in eukaryotes. PtdIns(3,5)P2 is produced from PI(3)P by phosphatidylinositol 3-phosphate 5-kinase FAB1/PIKfyve. Recently, we reported that FAB1 predominantly localizes on the SNX1-residing late endosomes and a loss-of FAB1 function causes the release of late endosomal effector proteins, ARA7/RABF2b and SORTING NEXIN 1 from the endosome membrane, indicating that FAB1 or its product PtdIns(3,5)P2 mediates the maturation process of the late endosomes. Intriguingly, the ectopic expression of FAB1A could complement the sucrose-dependent seedling growth phenotype of snx1–1 mutant. Here, we demonstrated that the depletion of SNX1 causes the release of SNX2b-mRFP from the endosomal membrane. However, overexpression of FAB1A-GFP reassembles SNX2b-mRFP on the endosomal membrane despite the absence of SNX1. From these results, we proposed that SNX2b homodimer or SNX2a/SNX2b heterodimer might function as functional Sorting Nexin complex instead of SNX1 to attach the endosomal membrane by binding of overproduced PI(3,5)P2 in Arabidopsis. PMID:26515455

  13. 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

  14. Characterization of drr1, an Alkamide-Resistant Mutant of Arabidopsis, Reveals an Important Role for Small Lipid Amides in Lateral Root Development and Plant Senescence1[C][W][OA

    PubMed Central

    Morquecho-Contreras, Alina; Méndez-Bravo, Alfonso; Pelagio-Flores, Ramón; Raya-González, Javier; Ortíz-Castro, Randy; López-Bucio, José

    2010-01-01

    Alkamides belong to a class of small lipid signals of wide distribution in plants, which are structurally related to the bacterial quorum-sensing signals N-acyl-l-homoserine lactones. Arabidopsis (Arabidopsis thaliana) seedlings display a number of root developmental responses to alkamides, including primary root growth inhibition and greater formation of lateral roots. To gain insight into the regulatory mechanisms by which these compounds alter plant development, we performed a mutant screen for identifying Arabidopsis mutants that fail to inhibit primary root growth when grown under a high concentration of N-isobutyl decanamide. A recessive N-isobutyl decanamide-resistant mutant (decanamide resistant root [drr1]) was isolated because of its continued primary root growth and reduced lateral root formation in response to this alkamide. Detailed characterization of lateral root primordia development in the wild type and drr1 mutants revealed that DRR1 is required at an early stage of pericycle cell activation to form lateral root primordia in response to both N-isobutyl decanamide and N-decanoyl-l-homoserine lactone, a highly active bacterial quorum-sensing signal. Exogenously supplied auxin similarly inhibited primary root growth and promoted lateral root formation in wild-type and drr1 seedlings, suggesting that alkamides and auxin act by different mechanisms to alter root system architecture. When grown both in vitro and in soil, drr1 mutants showed dramatically increased longevity and reduced hormone- and age-dependent senescence, which were related to reduced lateral root formation when exposed to stimulatory concentrations of jasmonic acid. Taken together, our results provide genetic evidence indicating that alkamides and N-acyl-l-homoserine lactones can be perceived by plants to modulate root architecture and senescence-related processes possibly by interacting with jasmonic acid signaling. PMID:20107026

  15. Modified Clp Protease Complex in the ClpP3 Null Mutant and Consequences for Chloroplast Development and Function in Arabidopsis1[C][W][OA

    PubMed Central

    Kim, Jitae; Olinares, Paul Dominic; Oh, Soo-hyun; Ghisaura, Stefania; Poliakov, Anton; Ponnala, Lalit; van Wijk, Klaas J.

    2013-01-01

    The plastid ClpPRT protease consists of two heptameric rings of ClpP1/ClpR1/ClpR2/ClpR3/ClpR4 (the R-ring) and ClpP3/ClpP4/ClpP5/ClpP6 (the P-ring) and peripherally associated ClpT1/ClpT2 subunits. Here, we address the contributions of ClpP3 and ClpP4 to ClpPRT core organization and function in Arabidopsis (Arabidopsis thaliana). ClpP4 is strictly required for embryogenesis, similar to ClpP5. In contrast, loss of ClpP3 (clpp3-1) leads to arrest at the hypocotyl stage; this developmental arrest can be removed by supplementation with sucrose or glucose. Heterotrophically grown clpp3-1 can be transferred to soil and generate viable seed, which is surprising, since we previously showed that CLPR2 and CLPR4 null alleles are always sterile and die on soil. Based on native gels and mass spectrometry-based quantification, we show that despite the loss of ClpP3, modified ClpPR core(s) could be formed, albeit at strongly reduced levels. A large portion of ClpPR subunits accumulated in heptameric rings, with overaccumulation of ClpP1/ClpP5/ClpP6 and ClpR3. Remarkably, the association of ClpT1 to the modified Clp core was unchanged. Large-scale quantitative proteomics assays of clpp3-1 showed a 50% loss of photosynthetic capacity and the up-regulation of plastoglobules and all chloroplast stromal chaperone systems. Specific chloroplast proteases were significantly up-regulated, whereas the major thylakoid protease (FtsH1/FtsH2/FtsH5/FtsH8) was clearly unchanged, indicating a controlled protease network response. clpp3-1 showed a systematic decrease of chloroplast-encoded proteins that are part of the photosynthetic apparatus but not of chloroplast-encoded proteins with other functions. Candidate substrates and an explanation for the differential phenotypes between the CLPP3, CLPP4, and CLPP5 null mutants are discussed. PMID:23548781

  16. Membrane function in lipid mutants of Arabidopsis. Second year progress report, [June 15, 1992--June 14, 1994

    SciTech Connect

    Browse, J.A.

    1994-03-01

    The fad2 mutants are deficient in activity of the endoplasmic reticulum oleate desaturase that is the main enzyme responsible for polyunsaturated lipid synthesis in developing seeds of oil crops. A comparison of wild type and fad2 seeds developing on heterozygous (Fad2/-) plants was used as a model for genetically engineered high oleate oilseeds of species such as soybean and canola. When fad2 seeds developed at normal temperatures (22{degree}C), they showed high viability comparable to wild type seeds. When a portion of seed development took place at 6{degree}C, germination of the wild type siblings remained high but germination of fad2 segregants declined considerably. This was true even when exposure to low temperature was limited to the final stages of seed filing and maturation. Compared to wild type, fully viable fad2 seeds produced at 22{degree}C had reduced lipid contents and were slower to germinate at 10{degree}C and 6{degree}C. These results indicate that for some oilseed species at least, molecular genetic manipulation of oleate levels in the oil may result in plant lines with unacceptable performance in the field.

  17. Sensitivity of different ecotypes and mutants of Arabidopsis thaliana toward the bacterial elicitor flagellin correlates with the presence of receptor-binding sites.

    PubMed

    Bauer, Z; Gómez-Gómez, L; Boller, T; Felix, G

    2001-12-01

    Flagellin, the main building block of the bacterial flagellum, acts as potent elicitor of defense responses in different plant species. Genetic analysis in Arabidopsis thaliana identified two distinct loci, termed FLS1 and FLS2, that are essential for perception of flagellin-derived elicitors. FLS2 was found to encode a leucine-rich repeat transmembrane receptor-like kinase with similarities to Toll-like receptors involved in the innate immune system of mammals and insects. Here we used a radiolabeled derivative of flg22, a synthetic peptide representing the elicitor-active domain of flagellin, to probe the interaction of flagellin with its receptor in A. thaliana. The high affinity binding site detected in intact cells and membrane preparations exhibited specificity for flagellin-derived peptides with biological activity as agonists or antagonists of the elicitor responses. Specific binding activity was measurable in all ecotypes of A. thaliana that show sensitivity to flagellin but was barely detectable in the flagellin-insensitive ecotype Ws-0 affected in FLS1. A strongly impaired binding of flagellin was observed also in several independent flagellin-insensitive mutants isolated from the flagellin-sensitive ecotype La-er. In particular, no binding was found in plants carrying a mutation in the LRR domain of FLS2. These data indicate that the formation of functional receptor-binding sites depends on genes encoded by both loci, FLS1 and FLS2. The tight correlation between the presence of the binding site and elicitor response provides strong evidence that this binding site acts as the physiological receptor of flagellin. PMID:11564731

  18. Suppression of late-flowering and semi-dwarf phenotypes in the Arabidopsis clock mutant lhy-12;cca1-101 by phyB under continuous light.

    PubMed

    Miyata, Kana; Calviño, Martín; Oda, Atsushi Oda; Sugiyama, Haruna; Mizoguchi, Tsuyoshi

    2011-08-01

    Photoperiodic flowering in Arabidopsis is controlled not only by floral activators such as GI, CO, and FT, but also by repressors such as SVP and FLC. Double mutations in LHY and CCA1 (lhy;cca1) accelerated flowering under short days, mainly by the GI-CO dependent pathway. In contrast, lhy;cca1 showed delayed flowering under continuous light (LL), probably due to the GI-CO independent pathway. This late-flowering phenotype was suppressed by svp, flc, and elf3. However, how SVP, FLC, and ELF3 mediate LHY/CCA1 and flowering time is not fully understood. We found that lhy;cca1 exhibited short hypocotyls and petioles under LL, but the molecular mechanism for these effects has not been elucidated. To address these questions, we performed a screen for mutations that suppress either or both of the lhy;cca1 phenotypes under LL, using two different approaches. We identified two novel mutations, a dominant (del1) and a recessive (phyB-2511) allele of phyB. The flowering times of single mutants of three phyB alleles, hy3-1, del1, and phyB-2511, are almost the same and earlier than those of wild-type plants. A similar level of acceleration of flowering time was observed in all three phyB mutants tested when combined with the late-flowering mutations co-2 and SVPox. However, the effect of phyB-2511 on lhy;cca1 was different from those by hy3-1 or del1. svp-3 did not strongly enhance the early-flowering phenotypes of phyB-2511 or del1. These results suggest that light signaling via PhyB may affect factors downstream of the clock proteins, controlling flowering time and organ elongation. phyB mutations with different levels of effects on lhy;cca1-dependent late flowering would be useful to determine a specific role for PHYB in the flowering pathway controlled by lhy;cca1 under LL. PMID:21822060

  19. Suppression of late-flowering and semi-dwarf phenotypes in the arabidopsis clock mutant lhy-12;cca1-101 by phyB under continuous light

    PubMed Central

    Oda, Atsushi; Sugiyama, Haruna

    2011-01-01

    Photoperiodic flowering in Arabidopsis is controlled not only by floral activators such as GI, CO and FT, but also by repressors such as SVP and FLC. Double mutations in LHY and CCA1 (lhy;cca1) accelerated flowering under short days, mainly by the GI-CO dependent pathway. In contrast, lhy;cca1 showed delayed flowering under continuous light (LL), probably due to the GI-CO independent pathway. This late-flowering phenotype was suppressed by svp, flc and elf3. However, how SVP, FLC and ELF3 mediate LHY/CCA1 and flowering time is not fully understood. We found that lhy;cca1 exhibited short hypocotyls and petioles under LL, but the molecular mechanism for these effects has not been elucidated. To address these questions, we performed a screen for mutations that suppress either or both of the lhy;cca1 phenotypes under LL, using two different approaches. We identified two novel mutations, a dominant (del1) and a recessive (phyB-2511) allele of phyB. The flowering times of single mutants of three phyB alleles, hy3-1, del1 and phyB-2511, are almost the same and earlier than those of wild-type plants. A similar level of acceleration of flowering time was observed in all three phyB mutants tested when combined with the late-flowering mutations co-2 and SVPox. However, the effect of phyB-2511 on lhy;cca1 was different from those by hy3-1 or del1. svp-3 did not strongly enhance the early-flowering phenotypes of phyB-2511 or del1. These results suggest that light signaling via PhyB may affect factors downstream of the clock proteins, controlling flowering time and organ elongation. phyB mutations with different levels of effects on lhy;cca1-dependent late flowering would be useful to determine a specific role for PHYB in the flowering pathway controlled by lhy;cca1 under LL. PMID:21822060

  20. Dynamic mineral partitioning in diverse Arabidopsis thaliana accessions: the ysl1ysl3 mutant fails to distribute Cu, Fe, and Zn normally in vegetative tissues, fruits, and seeds

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Arabidopsis thaliana has been used as a model to elucidate mechanisms of mineral uptake and homeostasis. We and others are interested in using the information gained from arabidopsis studies to improve mineral concentration in edible portions of crop plants, a strategy known as biofortification. To ...

  1. Transcriptional silencing induced by Arabidopsis T-DNA mutants is associated with 35S promoter siRNAs and requires genes involved in siRNA-mediated chromatin silencing.

    PubMed

    Mlotshwa, Sizolwenkosi; Pruss, Gail J; Gao, Zhihuan; Mgutshini, Nomathamsanqa L; Li, Junjie; Chen, Xuemei; Bowman, Lewis H; Vance, Vicki

    2010-11-01

    The utility of many T-DNA insertion mutant lines of Arabidopsis is compromised by their propensity to trigger transcriptional silencing of transgenes expressed from the CaMV 35S promoter. To try to circumvent this problem, we characterized the genetic requirements for maintenance of 35S promoter homology-dependent transcriptional gene silencing induced by the dcl3-1 (SALK_005512) T-DNA insertion mutant line. Surprisingly, even though DCL3 and RDR2 are known components of the siRNA-dependent transcriptional gene silencing pathway, transcriptional gene silencing of a 35S promoter-driven GUS hairpin transgene did occur in plants homozygous for the dcl3-1 T-DNA insertion and was unaffected by loss of function of RDR2. However, the transcriptional gene silencing was alleviated in dcl2 dcl3 dcl4 triple mutant plants and also by mutations in AGO4, NRPD2, HEN1 and MOM1. Thus, some T-DNA insertion mutant lines induce 35S promoter homology-dependent transcriptional silencing that requires neither DCL3 nor RDR2, but involves other genes known to function in siRNA-dependent transcriptional silencing. Consistent with these results, we detected 35S promoter siRNAs in dcl3-1 SALK line plants, suggesting that the 35S promoter homology-dependent silencing induced by some T-DNA insertion mutant lines is siRNA-mediated. PMID:21070421

  2. Magnesium uptake of Arabidopsis transporters, AtMRS2-10 and AtMRS2-11, expressed in Escherichia coli mutants: Complementation and growth inhibition by aluminum.

    PubMed

    Ishijima, Sumio; Uda, Misaki; Hirata, Tomohiro; Shibata, Makiko; Kitagawa, Nozomi; Sagami, Ikuko

    2015-06-01

    Magnesium (Mg2+) plays a critical role in many physiological processes. Mg2+ transport systems in Salmonella have been well documented, but those in Escherichia coli have not been fully elucidated. We examined the effects of corA, mgtA, yhiD and corC gene deletion on Mg2+ transport in E. coli. We obtained every combination of double, triple and quadruple mutants. The corA and mgtA double mutant required addition of 10 mM Mg2+ to Luria-Bertani (LB) medium for growth, and the corA, mgtA and yhiD triple mutant TM2 required a higher Mg2+ concentration. The Mg2+ requirement of the quadruple mutant was similar to that of TM2. The results demonstrated that either CorA or MgtA is necessary for normal E. coli growth in LB medium and that YhiD plays a role in Mg2+ transport under high Mg2+ growth conditions in E. coli. The Arabidopsis Mg2+ transporters, AtMRS2-10 and AtMRS2-11, were heterologously expressed in TM2 cells. TM2 cells expressing AtMRS2-10 and AtMRS2-11 could grow in LB medium that had been supplemented with 1 mM Mg2+ and without Mg2+ supplementation, respectively, and cell growth was inhibited by 2 mM AlCl3. The results indicated that the growth of TM2 expressing AtMRS2-10 and AtMRS2-11 reflected these AtMRS2 function for Mg2+ and aluminum. The E. coli TM2 cells are useful for functional analysis of Arabidopsis MRS2 proteins. PMID:25772503

  3. A quadruple mutant of Arabidopsis reveals a β-carotene hydroxylation activity for LUT1/CYP97C1 and a regulatory role of xanthophylls on determination of the PSI/PSII ratio

    PubMed Central

    2012-01-01

    Background Xanthophylls are oxygenated carotenoids playing an essential role as structural components of the photosynthetic apparatus. Xanthophylls contribute to the assembly and stability of light-harvesting complex, to light absorbance and to photoprotection. The first step in xanthophyll biosynthesis from α- and β-carotene is the hydroxylation of ε- and β-rings, performed by both non-heme iron oxygenases (CHY1, CHY2) and P450 cytochromes (LUT1/CYP97C1, LUT5/CYP97A3). The Arabidopsis triple chy1chy2lut5 mutant is almost completely depleted in β-xanthophylls. Results Here we report on the quadruple chy1chy2lut2lut5 mutant, additionally carrying the lut2 mutation (affecting lycopene ε-cyclase). This genotype lacks lutein and yet it shows a compensatory increase in β-xanthophylls with respect to chy1chy2lut5 mutant. Mutant plants show an even stronger photosensitivity than chy1chy2lut5, a complete lack of qE, the rapidly reversible component of non-photochemical quenching, and a peculiar organization of the pigment binding complexes into thylakoids. Biochemical analysis reveals that the chy1chy2lut2lut5 mutant is depleted in Lhcb subunits and is specifically affected in Photosystem I function, showing a deficiency in PSI-LHCI supercomplexes. Moreover, by analyzing a series of single, double, triple and quadruple Arabidopsis mutants in xanthophyll biosynthesis, we show a hitherto undescribed correlation between xanthophyll levels and the PSI-PSII ratio. The decrease in the xanthophyll/carotenoid ratio causes a proportional decrease in the LHCII and PSI core levels with respect to PSII. Conclusions The physiological and biochemical phenotype of the chy1chy2lut2lut5 mutant shows that (i) LUT1/CYP97C1 protein reveals a major β-carotene hydroxylase activity in vivo when depleted in its preferred substrate α-carotene; (ii) xanthophylls are needed for normal level of Photosystem I and LHCII accumulation. PMID:22513258

  4. 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. PMID:26259182

  5. Effects of dicer-like endoribonucleases 2 and 4 on infection of Arabidopsis thaliana by cucumber mosaic virus and a mutant virus lacking the 2b counter-defence protein gene.

    PubMed

    Ziebell, Heiko; Carr, John P

    2009-09-01

    In tobacco and Nicotiana benthamiana, limited cross-protection against cucumber mosaic virus strain Fny (Fny-CMV) was provided by prior inoculation with a deletion mutant lacking the 2b silencing-suppressor protein gene (Fny-CMVDelta2b). Cross-protection by Fny-CMVDelta2b did not result from induction of systemic RNA silencing. We investigated whether protection occurs through induction of localized RNA silencing by using Arabidopsis thaliana plants harbouring mutations in genes encoding the dicer-like (DCL) endoribonucleases 2, 3 and 4 involved in antiviral silencing. In wild-type A. thaliana (Col-0) plants, Fny-CMVDelta2b was symptomless and cross-protected against Fny-CMV infection. Cross-protection by Fny-CMVDelta2b against Fny-CMV infection was not abolished in dcl2, dcl3 or dcl4 mutant plants and was strongest in dcl2/4 double mutants, although in these plants and in dcl4 mutants, Fny-CMVDelta2b replicated to high levels and induced strong symptoms. The results suggest that Fny-CMVDelta2b/Fny-CMV cross-protection is not completely dependent on RNA silencing and also involves competition between these viruses. PMID:19474248

  6. AtGRX4, an Arabidopsis chloroplastic monothiol glutaredoxin, is able to suppress yeast GRx5 mutant phenotypes and respond to oxidative stress

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Arabidopsis monothiol glutaredoxin (Grx), AtGRX4, was targeted to chloroplasts/plastids and had high similarity to yeast Grx5. In yeast expression assays, AtGRX4 localized to the mitochondria and suppressed the sensitivity of grx5 cells to oxidants. In addition, AtGRX4 reduced iron accumulation and ...

  7. The rate of Photosynthesis remains relatively high at moderately high temperatures in Arabidopsis thaliana rca mutant expressing thermostable chimeric Rubisco Activase

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The rate of photosynthesis declines at moderately high temperatures (30-42 deg C) in temperate plants like Arabidopsis. The decline is due to deactivation of Rubisco which in turn is due to a reduced ability of activase to activate Rubisco (Crafts-Brandner and Salvucci, PNAS 97:13430-13435, 2000). W...

  8. 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...

  9. 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...

  10. 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

  11. 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 Central

    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

  12. NO3- and ClO3- fluxes in the chl1-5 mutant of Arabidopsis thaliana. Does the CHL1-5 gene encode a low-affinity NO3- transporter?

    PubMed Central

    Touraine, B; Glass, A D

    1997-01-01

    The CHL1 gene is considered to encode a low-affinity transport system (LATS) for NO3- in Arabidopsis thaliana (Y.-F. Tsay, J.I. Schroeder, K.A. Feldmann, N.M. Crawford [1993] Cell 72: 705-713). However, the anticipated reduced NO3- uptake by the LATS associated with loss of CHL1 gene activity in chl1-5 deletion mutants was evident only when plants were grown on NH4NO3. When KNO3 was the sole N source, NO3- accumulation and short-term tracer influx (using 13NO3- and 15NO3-) in leaves and roots of wild-type and mutant plants were essentially identical. Nevertheless, root uptake of 36CIO3- by the LATS and CIO3- accumulation in roots and shoots of mutant plants were significantly lower than in wild-type plants when grown on KNO3. One explanation for these results is that a second LATS is able to compensate for the chl1-5 deficiency in KNO3-grown plants. Growth on NH4NO3 may down-regulate the second LATS enough that the anticipated difference in NO3- uptake becomes apparent. PMID:9159946

  13. 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. PMID:24520956

  14. A pqr2 mutant encodes a defective polyamine transporter and is negatively affected by ABA for paraquat resistance in Arabidopsis thaliana.

    PubMed

    Dong, Shuchao; Hu, Huizhen; Wang, Youmei; Xu, Zhengdan; Zha, Yi; Cai, Xiwen; Peng, Liangcai; Feng, Shengqiu

    2016-09-01

    Despite the paraquat-resistant mutants that have been reported in plants, this study identified a novel A. thaliana mutant (pqr2) from an XVE inducible activation library based on its resistance to 2 μM paraquat. The pqr2 mutant exhibited a termination mutation in the exon of AT1G31830/PAR1/PQR2, encoded a polyamine uptake transporter AtPUT2/PAR1/PQR2. The PQR2 mutation could largely reduce superoxide accumulation and cell death in the pqr2 plants under paraquat treatment. Moreover, compared with wild type, the pqr2 mutant exhibited much reduced tolerance to putrescine, a classic polyamine compound, which confirmed that PQR2 encoded a defective polyamine transporter. Notably, co-treated with ABA and paraquat, both pqr2 mutant and wild type exhibited a lethal phenotype from seed germination, but the wild type like pqr2 mutant, could remain paraquat-resistance while co-treated with high dosage of Na2WO4, an ABA synthesis inhibitor. Gene expression analysis suggested that ABA signaling should widely regulate paraquat-responsive genes distinctively in wild type and pqr2 mutant. Hence, this study has for the first time reported about ABA negative effect on paraquat-resistance in A. thaliana, providing insight into the ABA signaling involved in the oxidative stress responses induced by paraquat in plants. PMID:27229891

  15. Expression of HMA4 cDNAs of the zinc hyperaccumulator Noccaea caerulescens from endogenous NcHMA4 promoters does not complement the zinc-deficiency phenotype of the Arabidopsis thaliana hma2hma4 double mutant

    PubMed Central

    Iqbal, Mazhar; Nawaz, Ismat; Hassan, Zeshan; Hakvoort, Henk W. J.; Bliek, Mattijs; Aarts, Mark G.M.; Schat, Henk

    2013-01-01

    Noccaea caerulescens (Nc) exhibits a very high constitutive expression of the heavy metal transporting ATPase, HMA4, as compared to the non-hyperaccumulator Arabidopsis thaliana (At), due to copy number expansion and altered cis-regulation. We screened a BAC library for HMA4 and found that HMA4 is triplicated in the genome of a N. caerulescens accession from a former Zn mine near La Calamine (LC), Belgium. We amplified multiple HMA4 promoter sequences from three calamine N. caerulescens accessions, and expressed AtHMA4 and different NcHMA4 cDNAs under At and Nc HMA4 promoters in the A. thaliana (Col) hma2hma4 double mutant. Transgenic lines expressing HMA4 under the At promoter were always fully complemented for root-to-shoot Zn translocation and developed normally at a 2-μM Zn supply, whereas the lines expressing HMA4 under Nc promoters usually showed only slightly enhanced root to shoot Zn translocation rates in comparison with the double mutant, probably owing to ectopic expression in the roots, respectively. When expression of the Zn deficiency responsive marker gene ZIP4 was tested, the transgenic lines expressing AtHMA4 under an NcHMA4-1-LC promoter showed on average a 7-fold higher expression in the leaves, in comparison with the double hma2hma4 mutant, showing that this construct aggravated, rather than alleviated the severity of foliar Zn deficiency in the mutant, possible owing to expression in the leaf mesophyll. PMID:24187545

  16. 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.

  17. Expression of Caenorhabditis elegans PCS in the AtPCS1-deficient Arabidopsis thaliana cad1-3 mutant separates the metal tolerance and non-host resistance functions of phytochelatin synthases.

    PubMed

    Kühnlenz, Tanja; Westphal, Lore; Schmidt, Holger; Scheel, Dierk; Clemens, Stephan

    2015-11-01

    Phytochelatin synthases (PCS) play key roles in plant metal tolerance. They synthesize small metal-binding peptides, phytochelatins, under conditions of metal excess. Respective mutants are strongly cadmium and arsenic hypersensitive. However, their ubiquitous presence and constitutive expression had long suggested a more general function of PCS besides metal detoxification. Indeed, phytochelatin synthase1 from Arabidopsis thaliana (AtPCS1) was later implicated in non-host resistance. The two different physiological functions may be attributable to the two distinct catalytic activities demonstrated for AtPCS1, that is the dipeptidyl transfer onto an acceptor molecule in phytochelatin synthesis, and the proteolytic deglycylation of glutathione conjugates. In order to test this hypothesis and to possibly separate the two biological roles, we expressed a phylogenetically distant PCS from Caenorhabditis elegans in an AtPCS1 mutant. We confirmed the involvement of AtPCS1 in non-host resistance by showing that plants lacking the functional gene develop a strong cell death phenotype when inoculated with the potato pathogen Phytophthora infestans. Furthermore, we found that the C. elegans gene rescues phytochelatin synthesis and cadmium tolerance, but not the defect in non-host resistance. This strongly suggests that the second enzymatic function of AtPCS1, which remains to be defined in detail, is underlying the plant immunity function. PMID:25764348

  18. Reduced function of the RNA-binding protein FPA rescues a T-DNA insertion mutant in the Arabidopsis ZHOUPI gene by promoting transcriptional read-through.

    PubMed

    Zhang, Yaohua; Li, Xin; Goodrich, Justin; Wu, Chunxia; Wei, Haichao; Yang, Suxin; Feng, Xianzhong

    2016-07-01

    T-DNA insertion mutants have been widely used to investigate plant gene functions. Unexpectedly, in several reported cases, the phenotype of T-DNA insertion mutations can be suppressed because of trans T-DNA interactions associated with epigenetic modification, which indicates that caution is needed when T-DNA mutants are used. In the present study, we characterized a novel process suppressing a T-DNA mutation. The spz2 (suppressor of zou 2) mutant was isolated as a suppressor of the phenotype of the zou-4 mutant caused by a T-DNA insertion in the first intron. The spz2 mutation partially recovered the native ZOU gene expression in the zou-4 background, but not in two other zou alleles, zou-2 and zou-3, with T-DNAs inserted in the exon and intron, respectively. The suppressed phenotype was inherited in a Mendelian fashion and is not associated with epigenetic modification. The recovery of the native ZOU gene expression in the spz2 zou-4 double mutant is caused by transcriptional read-through of the intronic T-DNA as a result of decreased proximal polyadenylation. SPZ2 encodes an RNA-binding protein, FPA, which is known to regulate polyadenylation site selection. This is the first example of FPA rescuing a T-DNA insertion mutation by affecting the polyadenylation site selection. PMID:27164978

  19. 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.

  20. Ectopic Expression of Apple F3′H Genes Contributes to Anthocyanin Accumulation in the Arabidopsis tt7 Mutant Grown Under Nitrogen Stress1[C][W][OA

    PubMed Central

    Han, Yuepeng; Vimolmangkang, Sornkanok; Soria-Guerra, Ruth Elena; Rosales-Mendoza, Sergio; Zheng, Danman; Lygin, Anatoli V.; Korban, Schuyler S.

    2010-01-01

    Three genes encoding flavonoid 3′-hydroxylase (F3′H) in apple (Malus × domestica), designated MdF3′HI, MdF3′HIIa, and MdF3′HIIb, have been identified. MdF3′HIIa and MdF3′HIIb are almost identical in amino acid sequences, and they are allelic, whereas MdF3′HI has 91% nucleotide sequence identity in the coding region to both MdF3′HIIa and MdF3′HIIb. MdF3′HI and MdF3′HII genes are mapped onto linkage groups 14 and 6, respectively, of the apple genome. Throughout the development of apple fruit, transcriptional levels of MdF3′H genes along with other anthocyanin biosynthesis genes are higher in the red-skinned cv Red Delicious than that in the yellow-skinned cv Golden Delicious. Moreover, patterns of MdF3′H gene expression correspond to accumulation patterns of flavonoids in apple fruit. These findings suggest that MdF3′H genes are coordinately expressed with other genes in the anthocyanin biosynthetic pathway in apple. The functionality of these apple F3′H genes has been demonstrated via their ectopic expression in both the Arabidopsis (Arabidopsis thaliana) transparent testa7-1 (tt7) mutant and tobacco (Nicotiana tabacum). When grown under nitrogen-deficient conditions, transgenic Arabidopsis tt7 seedlings expressing apple F3′H regained red color pigmentation and significantly accumulated both 4′-hydrylated pelargonidin and 3′,4′-hydrylated cyanidin. When compared with wild-type plants, flowers of transgenic tobacco lines overexpressing apple F3′H genes exhibited enhanced red color pigmentation. This suggests that the F3′H enzyme may coordinately interact with other flavonoid enzymes in the anthocyanin biosynthesis pathway. PMID:20357139

  1. Stable Isotope Metabolic Labeling-based Quantitative Phosphoproteomic Analysis of Arabidopsis Mutants Reveals Ethylene-regulated Time-dependent Phosphoproteins and Putative Substrates of Constitutive Triple Response 1 Kinase*

    PubMed Central

    Yang, Zhu; Guo, Guangyu; Zhang, Manyu; Liu, Claire Y.; Hu, Qin; Lam, Henry; Cheng, Han; Xue, Yu; Li, Jiayang; Li, Ning

    2013-01-01

    Ethylene is an important plant hormone that regulates numerous cellular processes and stress responses. The mode of action of ethylene is both dose- and time-dependent. Protein phosphorylation plays a key role in ethylene signaling, which is mediated by the activities of ethylene receptors, constitutive triple response 1 (CTR1) kinase, and phosphatase. To address how ethylene alters the cellular protein phosphorylation profile in a time-dependent manner, differential and quantitative phosphoproteomics based on 15N stable isotope labeling in Arabidopsis was performed on both one-minute ethylene-treated Arabidopsis ethylene-overly-sensitive loss-of-function mutant rcn1-1, deficient in PP2A phosphatase activity, and a pair of long-term ethylene-treated wild-type and loss-of-function ethylene signaling ctr1-1 mutants, deficient in mitogen-activated kinase kinase kinase activity. In total, 1079 phosphopeptides were identified, among which 44 were novel. Several one-minute ethylene-regulated phosphoproteins were found from the rcn1-1. Bioinformatic analysis of the rcn1-1 phosphoproteome predicted nine phosphoproteins as the putative substrates for PP2A phosphatase. In addition, from CTR1 kinase-enhanced phosphosites, we also found putative CTR1 kinase substrates including plastid transcriptionally active protein and calcium-sensing receptor. These regulatory proteins are phosphorylated in the presence of ethylene. Analysis of ethylene-regulated phosphosites using the group-based prediction system with a protein–protein interaction filter revealed a total of 14 kinase–substrate relationships that may function in both CTR1 kinase- and PP2A phosphatase-mediated phosphor-relay pathways. Finally, several ethylene-regulated post-translational modification network models have been built using molecular systems biology tools. It is proposed that ethylene regulates the phosphorylation of arginine/serine-rich splicing factor 41, plasma membrane intrinsic protein 2A, light

  2. 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

  3. Loss-of-Function Mutants and Overexpression Lines of the Arabidopsis Cyclin CYCA1;2/TARDY ASYNCHRONOUS MEIOSIS Exhibit Different Defects in Prophase-I Meiocytes but Produce the Same Meiotic Products

    PubMed Central

    Wang, Yixing; Yang, Ming

    2014-01-01

    In Arabidopsis, loss-of-function mutations in the A-type cyclin CYCA1;2/TARDY ASYNCHRONOUS MEIOSIS (TAM) gene lead to the production of abnormal meiotic products including triads and dyads. Here we report that overexpression of TAM by the ASK1:TAM transgene also led to the production of triads and dyads in meiosis, as well as shriveled seeds, in a dominant fashion. However, the partial loss-of-function mutant tam-1, an ASK1:TAM line, and the wild type differed in dynamic changes in chromosome thread thickness from zygotene to diplotene. We also found that the pericentromeric heterochromatin regions in male meiocytes in tam-1 and tam-2 (a null allele) frequently formed a tight cluster at the pachytene and diplotene stages, in contrast to the infrequent occurrences of such clusters in the wild type and the ASK1:TAM line. Immunolocalization studies of the chromosome axial component ASY1 revealed that ASY1 was highly expressed at the appropriate male meiotic stages but not localized to the chromosomes in tam-2. The level of ASY1, however, was greatly reduced in another ASK1:TAM line with much overexpressed TAM. Our results indicate that the reduction and increase in the activity of TAM differentially affect chromosomal morphology and the action of ASY1 in prophase I. Based on these results, we propose that either the different meiotic defects or a common defect such as missing ASY1 on the chromosomal axes triggers a hitherto uncharacterized cell cycle checkpoint in the male meiocytes in the tam mutants and ASK1:TAM lines, leading to the production of the same abnormal meiotic products. PMID:25402453

  4. 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. PMID:26216646

  5. The pineapple AcMADS1 promoter confers high level expression in tomato and Arabidopsis flowering and fruiting tissues, but AcMADS1 does not complement the tomato LeMADS-RIN (rin) mutant.

    PubMed

    Moyle, Richard L; Koia, Jonni H; Vrebalov, Julia; Giovannoni, James; Botella, Jose R

    2014-11-01

    A previous EST study identified a MADS box transcription factor coding sequence, AcMADS1, that is strongly induced during non-climacteric pineapple fruit ripening. Phylogenetic analyses place the AcMADS1 protein in the same superclade as LeMADS-RIN, a master regulator of fruit ripening upstream of ethylene in climacteric tomato. LeMADS-RIN has been proposed to be a global ripening regulator shared among climacteric and non-climacteric species, although few functional homologs of LeMADS-RIN have been identified in non-climacteric species. AcMADS1 shares 67 % protein sequence similarity and a similar expression pattern in ripening fruits as LeMADS-RIN. However, in this study AcMADS1 was not able to complement the tomato rin mutant phenotype, indicating AcMADS1 may not be a functionally conserved homolog of LeMADS-RIN or has sufficiently diverged to be unable to act in the context of the tomato network of interacting proteins. The AcMADS1 promoter directed strong expression of the GUS reporter gene to fruits and developing floral organs in tomato and Arabidopsis thaliana, suggesting AcMADS1 may play a role in flower development as well as fruitlet ripening. The AcMADS1 promoter provides a useful molecular tool for directing transgene expression, particularly where up-regulation in developing flowers and fruits is desirable. PMID:25139231

  6. Activated expression of AtEDT1/HDG11 promotes lateral root formation in Arabidopsis mutant edt1 by upregulating jasmonate biosynthesis.

    PubMed

    Cai, Xiao-Teng; Xu, Ping; Wang, Yao; Xiang, Cheng-Bin

    2015-12-01

    Root architecture is crucial for plants to absorb water and nutrients. We previously reported edt1 (edt1D) mutant with altered root architecture that contributes significantly to drought resistance. However, the underlying molecular mechanisms are not well understood. Here we report one of the mechanisms underlying EDT1/HDG11-conferred altered root architecture. Root transcriptome comparison between the wild type and edt1D revealed that the upregulated genes involved in jasmonate biosynthesis and signaling pathway were enriched in edt1D root, which were confirmed by quantitative RT-PCR. Further analysis showed that EDT1/HDG11, as a transcription factor, bound directly to the HD binding sites in the promoters of AOS, AOC3, OPR3, and OPCL1, which encode four key enzymes in JA biosynthesis. We found that the jasmonic acid level was significantly elevated in edt1D root compared with that in the wild type subsequently. In addition, more auxin accumulation was observed in the lateral root primordium of edt1D compared with that of wild type. Genetic analysis of edt1D opcl1 double mutant also showed that HDG11 was partially dependent on JA in regulating LR formation. Taken together, overexpression of EDT1/HDG11 increases JA level in the root of edt1D by directly upregulating the expressions of several genes encoding JA biosynthesis enzymes to activate auxin signaling and promote lateral root formation. PMID:25752924

  7. Arabidopsis thaliana phytochelatin synthase 2 is constitutively active in vivo and can rescue the growth defect of the PCS1-deficient cad1-3 mutant on Cd-contaminated soil.

    PubMed

    Kühnlenz, Tanja; Schmidt, Holger; Uraguchi, Shimpei; Clemens, Stephan

    2014-08-01

    Phytochelatins play a key role in the detoxification of metals in plants and many other eukaryotes. Their formation is catalysed by phytochelatin synthases (PCS) in the presence of metal excess. It appears to be common among higher plants to possess two PCS genes, even though in Arabidopsis thaliana only AtPCS1 has been demonstrated to confer metal tolerance. Employing a highly sensitive quantification method based on ultraperformance electrospray ionization quadrupole time-of-flight mass spectrometry, we detected AtPCS2-dependent phytochelatin formation. Overexpression of AtPCS2 resulted in constitutive phytochelatin accumulation, i.e. in the absence of metal excess, both in planta and in a heterologous system. This indicates distinct enzymatic differences between AtPCS1 and AtPCS2. Furthermore, AtPCS2 was able to partially rescue the Cd hypersensitivity of the AtPCS1-deficient cad1-3 mutant in a liquid seedling assay, and, more importantly, when plants were grown on soil spiked with Cd to a level that is close to what can be found in agricultural soils. No rescue was found in vertical-plate assays, the most commonly used method to assess metal tolerance. Constitutive AtPCS2-dependent phytochelatin synthesis suggests a physiological role of AtPCS2 other than metal detoxification. The differences observed between wild-type plants and cad1-3 on Cd soil demonstrated: (i) the essentiality of phytochelatin synthesis for tolerating levels of Cd contamination that can naturally be encountered by plants outside of metal-rich habitats, and (ii) a contribution to Cd accumulation under these conditions. PMID:24821959

  8. Arabidopsis thaliana phytochelatin synthase 2 is constitutively active in vivo and can rescue the growth defect of the PCS1-deficient cad1-3 mutant on Cd-contaminated soil

    PubMed Central

    Kühnlenz, Tanja; Schmidt, Holger; Uraguchi, Shimpei; Clemens, Stephan

    2014-01-01

    Phytochelatins play a key role in the detoxification of metals in plants and many other eukaryotes. Their formation is catalysed by phytochelatin synthases (PCS) in the presence of metal excess. It appears to be common among higher plants to possess two PCS genes, even though in Arabidopsis thaliana only AtPCS1 has been demonstrated to confer metal tolerance. Employing a highly sensitive quantification method based on ultraperformance electrospray ionization quadrupole time-of-flight mass spectrometry, we detected AtPCS2-dependent phytochelatin formation. Overexpression of AtPCS2 resulted in constitutive phytochelatin accumulation, i.e. in the absence of metal excess, both in planta and in a heterologous system. This indicates distinct enzymatic differences between AtPCS1 and AtPCS2. Furthermore, AtPCS2 was able to partially rescue the Cd hypersensitivity of the AtPCS1-deficient cad1-3 mutant in a liquid seedling assay, and, more importantly, when plants were grown on soil spiked with Cd to a level that is close to what can be found in agricultural soils. No rescue was found in vertical-plate assays, the most commonly used method to assess metal tolerance. Constitutive AtPCS2-dependent phytochelatin synthesis suggests a physiological role of AtPCS2 other than metal detoxification. The differences observed between wild-type plants and cad1-3 on Cd soil demonstrated: (i) the essentiality of phytochelatin synthesis for tolerating levels of Cd contamination that can naturally be encountered by plants outside of metal-rich habitats, and (ii) a contribution to Cd accumulation under these conditions. PMID:24821959

  9. The jiaoyao1 Mutant Is an Allele of korrigan1 That Abolishes Endoglucanase Activity and Affects the Organization of Both Cellulose Microfibrils and Microtubules in Arabidopsis[C][W

    PubMed Central

    Lei, Lei; Zhang, Tian; Strasser, Richard; Lee, Christopher M.; Gonneau, Martine; Mach, Lukas; Vernhettes, Samantha; Kim, Seong H.; J. Cosgrove, Daniel; Li, Shundai; Gu, Ying

    2014-01-01

    In higher plants, cellulose is synthesized by plasma membrane–localized cellulose synthase complexes (CSCs). Arabidopsis thaliana GH9A1/KORRIGAN1 is a membrane-bound, family 9 glycosyl hydrolase that is important for cellulose synthesis in both primary and secondary cell walls. Most previously identified korrigan1 mutants show severe phenotypes such as embryo lethality; therefore, the role of GH9A1 in cellulose synthesis remains unclear. Here, we report a novel A577V missense mutation, designated jiaoyao1 (jia1), in the second of the glycosyl hydrolase family 9 active site signature motifs in GH9A1. jia1 is defective in cell expansion in dark-grown hypocotyls, roots, and adult plants. Consistent with its defect in cell expansion, this mutation in GH9A1 resulted in reduced cellulose content and reduced CSC velocity at the plasma membrane. Green fluorescent protein–GH9A1 is associated with CSCs at multiple locations, including the plasma membrane, Golgi, trans-Golgi network, and small CESA-containing compartments or microtubule-associated cellulose synthase compartments, indicating a tight association between GH9A1 and CSCs. GH9A1A577V abolishes the endoglucanase activity of GH9A1 in vitro but does not affect its interaction with CESAs in vitro, suggesting that endoglucanase activity is important for cellulose synthesis. Interestingly, jia1 results in both cellulose microfibril and microtubule disorganization. Our study establishes the important role of endoglucanase in cellulose synthesis and cellulose microfibril organization in plants. PMID:24963054

  10. 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

  11. Analysis of a Range of Catabolic Mutants Provides Evidence That Phytanoyl-Coenzyme A Does Not Act as a Substrate of the Electron-Transfer Flavoprotein/Electron-Transfer Flavoprotein:Ubiquinone Oxidoreductase Complex in Arabidopsis during Dark-Induced Senescence1[W][OA

    PubMed Central

    Araújo, Wagner L.; Ishizaki, Kimitsune; Nunes-Nesi, Adriano; Tohge, Takayuki; Larson, Tony R.; Krahnert, Ina; Balbo, Ilse; Witt, Sandra; Dörmann, Peter; Graham, Ian A.; Leaver, Christopher J.; Fernie, Alisdair R.

    2011-01-01

    The process of dark-induced senescence in plants is not fully understood, however, the functional involvement of an electron-transfer flavoprotein/electron-transfer flavoprotein:ubiquinone oxidoreductase (ETF/ETFQO), has been demonstrated. Recent studies have revealed that the enzymes isovaleryl-coenzyme A (CoA) dehydrogenase and 2-hydroxyglutarate dehydrogenase act as important electron donors to this complex. In addition both enzymes play a role in the breakdown of cellular carbon storage reserves with isovaleryl-CoA dehydrogenase being involved in degradation of the branched-chain amino acids, phytol, and lysine while 2-hydroxyglutarate dehydrogenase is exclusively involved in lysine degradation. Given that the chlorophyll breakdown intermediate phytanoyl-CoA accumulates dramatically both in knockout mutants of the ETF/ETFQO complex and of isovaleryl-CoA dehydrogenase following growth in extended dark periods we have investigated the direct importance of chlorophyll breakdown for the supply of carbon and electrons during this process. For this purpose we isolated three independent Arabidopsis (Arabidopsis thaliana) knockout mutants of phytanoyl-CoA 2-hydroxylase and grew them under the same extended darkness regime as previously used. Despite the fact that these mutants accumulated phytanoyl-CoA and also 2-hydroxyglutarate they exhibited no morphological changes in comparison to the other mutants previously characterized. These results are consistent with a single entry point of phytol breakdown into the ETF/ETFQO system and furthermore suggest that phytol is not primarily metabolized by this pathway. Furthermore analysis of isovaleryl-CoA dehydrogenase/2-hydroxyglutarate dehydrogenase double mutants generated here suggest that these two enzymes essentially account for the entire electron input via the ETF complex. PMID:21788362

  12. 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...

  13. An Arabidopsis callose synthase.

    PubMed

    Ostergaard, Lars; Petersen, Morten; Mattsson, Ole; Mundy, John

    2002-08-01

    Beta-1,3-glucan polymers are major structural components of fungal cell walls, while cellulosic beta-1,4-glucan is the predominant polysaccharide in plant cell walls. Plant beta-1,3-glucan, called callose, is produced in pollen and in response to pathogen attack and wounding, but it has been unclear whether callose synthases can also produce cellulose and whether plant cellulose synthases may also produce beta-1,3-glucans. We describe here an Arabidopsis gene, AtGsl5, encoding a plasma membrane-localized protein homologous to yeast beta-1,3-glucan synthase whose expression partially complements a yeast beta-1,3-glucan synthase mutant. AtGsl5 is developmentally expressed at highest levels in flowers, consistent with flowers having high beta-1,3-glucan synthase activities for deposition of callose in pollen. A role for AtGsl5 in callose synthesis is also indicated by AtGsl5 expression in the Arabidopsis mpk4 mutant which exhibits systemic acquired resistance (SAR), elevated beta-1,3-glucan synthase activity, and increased callose levels. In addition, AtGsl5 is a likely target of salicylic acid (SA)-dependent SAR, since AtGsl5 mRNA accumulation is induced by SA in wild-type plants, while expression of the nahG salicylate hydroxylase reduces AtGsl5 mRNA levels in the mpk4 mutant. These results indicate that AtGsl5 is likely involved in callose synthesis in flowering tissues and in the mpk4 mutant. PMID:12081364

  14. 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. PMID:26227242

  15. 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.

  16. Arabidopsis nucleolin affects plant development and patterning.

    PubMed

    Petricka, Jalean Joyanne; Nelson, Timothy Mark

    2007-05-01

    Nucleolin is a major nucleolar protein implicated in many aspects of ribosomal biogenesis, including early events such as processing of the large 35S preribosomal RNA. We found that the Arabidopsis (Arabidopsis thaliana) parallel1 (parl1) mutant, originally identified by its aberrant leaf venation, corresponds to the Arabidopsis nucleolin gene. parl1 mutants display parallel leaf venation, aberrant localization of the provascular marker Athb8:beta-glucuronidase, the auxin-sensitive reporter DR5:beta-glucuronidase, and auxin-dependent growth defects. PARL1 is highly similar to the yeast (Saccharomyces cerevisiae) nucleolin NUCLEAR SIGNAL RECOGNITION 1 (NSR1) multifunctional protein; the Arabidopsis PARL1 gene can rescue growth defects of yeast nsr1 null mutants. This suggests that PARL1 protein may have roles similar to those of the yeast nucleolin in nuclear signal recognition, ribosomal processing, and ribosomal subunit accumulation. Based on the range of auxin-related defects in parl1 mutants, we propose that auxin-dependent organ growth and patterning is highly sensitive to the efficiency of nucleolin-dependent ribosomal processing. PMID:17369435

  17. 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.

  18. The Phenylpropanoid Pathway in Arabidopsis

    PubMed Central

    Fraser, Christopher M.; Chapple, Clint

    2011-01-01

    The phenylpropanoid pathway serves as a rich source of metabolites in plants, being required for the biosynthesis of lignin, and serving as a starting point for the production of many other important compounds, such as the flavonoids, coumarins, and lignans. In spite of the fact that the phenylpropanoids and their derivatives are sometimes classified as secondary metabolites, their relevance to plant survival has been made clear via the study of Arabidopsis and other plant species. As a model system, Arabidopsis has helped to elucidate many details of the phenylpropanoid pathway, its enzymes and intermediates, and the interconnectedness of the pathway with plant metabolism as a whole. These advances in our understanding have been made possible in large part by the relative ease with which mutations can be generated, identified, and studied in Arabidopsis. Herein, we provide an overview of the research progress that has been made in recent years, emphasizing both the genes (and gene families) associated with the phenylpropanoid pathway in Arabidopsis, and the end products that have contributed to the identification of many mutants deficient in the phenylpropanoid metabolism: the sinapate esters. PMID:22303276

  19. Accumulation of endogenous salicylic acid confers drought tolerance to Arabidopsis.

    PubMed

    Okuma, Eiji; Nozawa, Rieko; Murata, Yoshiyuki; Miura, Kenji

    2014-01-01

    We investigated stomatal phenotype and drought tolerance of Arabidopsis salicylic acid-accumulating mutants, acd6 and cpr5. In these mutants, the light-induced stomatal opening was impaired and the impairment of stomatal opening was restored by peroxidase inhibitors, salicylhydroxamic acid, and azide. The acd6 and cpr5 mutant plants were more tolerant to drought stress than wild-type plants. Introduction of nahG gene into the acd6 and cpr5 mutants removed the inhibition of stomatal opening and reduced the drought tolerance. Drought tolerance-related genes were more highly expressed in the cpr5 and acd6 mutant plants than in the wild-type plants. These results suggest that accumulation of salicylic acid improves drought tolerance through inhibition of light-induced stomatal opening in Arabidopsis. PMID:24603484

  20. ERECTA contributes to non-host resistance to Magnaporthe oryzae in Arabidopsis.

    PubMed

    Takahashi, Toshiharu; Shibuya, Haruki; Ishikawa, Atsushi

    2016-07-01

    ERECTA controls both developmental processes and disease resistance in Arabidopsis. We investigated the function of ERECTA in non-host resistance to Magnaporthe oryzae in Arabidopsis. In the pen2 er mutant, penetration resistance and post-penetration resistance to M. oryzae were compromised. These results suggest that ERECTA is involved in both penetration and post-penetration resistance to M. oryzae in Arabidopsis. PMID:26924213

  1. Forward genetic screen for auxin-deficient mutants by cytokinin

    PubMed Central

    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

    2015-01-01

    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. PMID:26143750

  2. Abscisic acid deficiency increases defence responses against Myzus persicae in Arabidopsis.

    PubMed

    Hillwig, Melissa S; Chiozza, Mariana; Casteel, Clare L; Lau, Siau Ting; Hohenstein, Jessica; Hernández, Enrique; Jander, Georg; MacIntosh, Gustavo C

    2016-02-01

    Comparison of Arabidopsis thaliana (Arabidopsis) gene expression induced by Myzus persicae (green peach aphid) feeding, aphid saliva infiltration and abscisic acid (ABA) treatment showed a significant positive correlation. In particular, ABA-regulated genes are over-represented among genes that are induced by M. persicae saliva infiltration into Arabidopsis leaves. This suggests that the induction of ABA-related gene expression could be an important component of the Arabidopsis-aphid interaction. Consistent with this hypothesis, M. persicae populations induced ABA production in wild-type plants. Furthermore, aphid populations were smaller on Arabidopsis aba1-1 mutants, which cannot synthesize ABA, and showed a significant preference for wild-type plants compared with the mutant. Total free amino acids, which play an important role in aphid nutrition, were not altered in the aba1-1 mutant line, but the levels of isoleucine (Ile) and tryptophan (Trp) were differentially affected by aphids in wild-type and mutant plants. Recently, indole glucosinolates have been shown to promote aphid resistance in Arabidopsis. In this study, 4-methoxyindol-3-ylmethylglucosinolate was more abundant in the aba1-1 mutant than in wild-type Arabidopsis, suggesting that the induction of ABA signals that decrease the accumulation of defence compounds may be beneficial for aphids. PMID:25943308

  3. 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.

  4. 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

  5. Isolation of ABA hypersensitive mutants in allhexaploid breadwheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The plant hormone abscisic acid (ABA) stimulates seed dormancy during embryo maturation, inhibits germination of mature seed, and stimulates stress responses such as stomatal closure in response to drought stress. Arabidopsis mutants isolated for ABA hypersensitive (ABH) seed germination showed incr...

  6. 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. PMID:22173099

  7. 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...

  8. Arabidopsis cpSRP54 regulates carotenoid accumulation in Arabidopsis and Brassica napus

    PubMed Central

    Gruber, Margaret Y.; Hannoufa, Abdelali

    2012-01-01

    An Arabidopsis thaliana mutant, cbd (carotenoid biosynthesis deficient), was recovered from a mutant population based on its yellow cotyledons, yellow-first true leaves, and stunted growth. Seven-day-old seedlings and mature seeds of this mutant had lower chlorophyll and total carotenoids than the wild type (WT). Genetic and molecular characterization revealed that cbd was a recessive mutant caused by a T-DNA insertion in the gene cpSRP54 encoding the 54kDa subunit of the chloroplast signal recognition particle. Transcript levels of most of the main carotenoid biosynthetic genes in cbd were unchanged relative to WT, but expression increased in carotenoid and abscisic acid catabolic genes. The chloroplasts of cbd also had developmental defects that contributed to decreased carotenoid and chlorophyll contents. Transcription of AtGLK1 (Golden 2-like 1), AtGLK2, and GUN4 appeared to be disrupted in the cbd mutant suggesting that the plastid-to-nucleus retrograde signal may be affected, regulating the changes in chloroplast functional and developmental states and carotenoid content flux. Transformation of A. thaliana and Brassica napus with a gDNA encoding the Arabidopsis cpSRP54 showed the utility of this gene in enhancing levels of seed carotenoids without affecting growth or seed yield. PMID:22791829

  9. 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

  10. Arabidopsis thaliana glucuronosyltransferase in family GT14.

    PubMed

    Dilokpimol, Adiphol; Geshi, Naomi

    2014-01-01

    Arabinogalactan proteins are abundant cell-surface proteoglycans in plants and are involved in many cellular processes including somatic embryogenesis, cell-cell interactions, and cell elongation. We reported a glucuronosyltransferase encoded by Arabidopsis AtGlcAT14A, which catalyzes an addition of glucuronic acid residues to β-1,3- and β-1,6-linked galactans of arabinogalactan (Knoch et al. 2013). The knockout mutant of this gene resulted in the enhanced growth rate of hypocotyls and roots of seedlings, suggesting an involvement of AtGlcAT14A in cell elongation. AtGlcAt14A belongs to the family GT14 in the Carbohydrate Active Enzyme database (CAZy; www.cazy.org), in which a total of 11 proteins, including AtGLCAT14A, are classified from Arabidopsis thaliana. In this paper, we report the enzyme activities for the rest of the Arabidopsis GT14 isoforms, analyzed in the same way as for AtGlcAT14A. Evidently, two other Arabidopsis GT14 isoforms, At5g15050 and At2g37585, also possess the glucuronosyltransferase activity adding glucuronic acid residues to β-1,3- and β-1,6-linked galactans. Therefore, we named At5g15050 and At2g37585 as AtGlcAT14B and AtGlcAT14C, respectively. PMID:24739253

  11. 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

  12. 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. PMID:22323776

  13. 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.

  14. Role of a chalcone isomerase-like protein in flavonoid biosynthesis in Arabidopsis thaliana

    PubMed Central

    Jiang, Wenbo; Yin, Qinggang; Wu, Ranran; Zheng, Guangshun; Liu, Jinyue; Dixon, Richard A.; Pang, Yongzhen

    2015-01-01

    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. PMID:26347569

  15. 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

  16. A gain-of-function phenotype conferred by over-expression of functional subunits of the COP9 signalosome in Arabidopsis.

    PubMed

    Kang, D; Wang, X; Cao, K; Sun, C; Deng, X W; Wei, N

    2000-09-01

    The COP9 signalosome is a conserved cellular regulator present in diverse organisms. To understand the structural and functional relationship of the COP9 signalosome with its subunits, we expressed in wild-type and mutant Arabidopsis backgrounds two orthologues of subunit 1, rice FUS6 (rFUS6) and human GPS1, and Arabidopsis subunit 8 (COP9). In Arabidopsis, rFUS6 can functionally replace Arabidopsis endogenous FUS6 to form the COP9 signalosome complex and rescue the null fus6-1 mutant phenotype. Moreover, light-grown rFUS6 over-expression seedlings displayed longer hypocotyls and reduced anthocyanin accumulation in comparison to wild-type seedlings, which is opposite to the fus6/cop11 mutant phenotype. The long-hypocotyl phenotype was also observed in transgenic seedlings over-expressing Arabidopsis COP9. This finding indicates that over-expression of a functional subunit 1 or subunit 8 of the COP9 signalosome confers a gain-of-function phenotype relative to the complex. Human GPS1, when expressed in the fus6-1 null mutant of Arabidopsis, can assemble into a chimeric COP9 signalosome at low efficiency, demonstrating the structural conservation of the complexes between human and Arabidopsis. This low-abundancy chimeric complex is insufficient to fully rescue the mutant but is able to attenuate the mutant severity. PMID:10972886

  17. A FUSCA gene of Arabidopsis encodes a novel protein essential for plant development.

    PubMed Central

    Castle, L A; Meinke, D W

    1994-01-01

    Arabidopsis fusca mutants display striking purple coloration due to anthocyanin accumulation in their cotyledons. We describe six recessive fusca mutants isolated from Agrobacterium-transformed Arabidopsis families. These mutants first become defective during embryogenesis and exhibit limited seedling development. Double mutant constructs revealed that developmental defects were not simply a consequence of anthocyanin accumulation. fusca seedlings showed altered responses to several environmental and endogenous factors. Allelism tests established that three fusca loci are represented by mutants previously described as defective in light-regulated responses. To study the molecular basis of the fusca phenotype, we cloned the FUS6 gene. FUS6 encodes a novel protein that is hydrophilic, alpha-helical, and contains potential protein kinase C phosphorylation sites. The FUSCA proteins appear to act in a network of signal transduction pathways critical for plant development. PMID:8130643

  18. Singlet oxygen scavenging activity of tocopherol and plastochromanol in Arabidopsis thaliana: relevance to photooxidative stress.

    PubMed

    Rastogi, Anshu; Yadav, Deepak Kumar; Szymańska, Renata; Kruk, Jerzy; Sedlářová, Michaela; Pospíšil, Pavel

    2014-02-01

    In the present study, singlet oxygen (¹O₂) scavenging activity of tocopherol and plastochromanol was examined in tocopherol cyclase-deficient mutant (vte1) of Arabidopsis thaliana lacking both tocopherol and plastochromanol. It is demonstrated here that suppression of tocopherol and plastochromanol synthesis in chloroplasts isolated from vte1 Arabidopsis plants enhanced ¹O₂ formation under high light illumination as monitored by electron paramagnetic resonance spin-trapping spectroscopy. The exposure of vte1 Arabidopsis plants to high light resulted in the formation of secondary lipid peroxidation product malondialdehyde as determined by high-pressure liquid chromatography. Furthermore, it is shown here that the imaging of ultra-weak photon emission known to reflect oxidation of lipids was unambiguously higher in vte1 Arabidopsis plants. Our results indicate that tocopherol and plastochromanol act as efficient ¹O₂ scavengers and protect effectively lipids against photooxidative damage in Arabidopsis plants. PMID:23848570

  19. Farnesylcysteine Lyase is Involved in Negative Regulation of Abscisic Acid Signaling in Arabidopsis

    PubMed Central

    Huizinga, David H.; Denton, Ryan; Koehler, Kelly G.; Tomasello, Ashley; Wood, Lyndsay; Sen, Stephanie E.; Crowell, Dring N.

    2010-01-01

    The Arabidopsis FCLY gene encodes a specific farnesylcysteine (FC) lyase, which is responsible for the oxidative metabolism of FC to farnesal and cysteine. In addition, fcly mutants with quantitative decreases in FC lyase activity exhibit an enhanced response to ABA. However, the enzymological properties of the FCLY-encoded enzyme and its precise role in ABA signaling remain unclear. Here, we show that recombinant Arabidopsis FC lyase expressed in insect cells exhibits high selectivity for FC as a substrate and requires FAD and molecular oxygen for activity. Arabidopsis FC lyase is also shown to undergo post-translational N-glycosylation. FC, which is a competitive inhibitor of isoprenylcysteine methyltransferase (ICMT), accumulates in fcly mutants. Moreover, the enhanced response of fcly mutants to ABA is reversed by ICMT overexpression. These observations support the hypothesis that the ABA hypersensitive phenotype of fcly plants is the result of FC accumulation and inhibition of ICMT. PMID:19969520

  20. Ethylene Modulates Sphingolipid Synthesis in Arabidopsis

    PubMed Central

    Wu, Jian-xin; Wu, Jia-li; Yin, Jian; Zheng, Ping; Yao, Nan

    2015-01-01

    Sphingolipids have essential structural and bioactive functions in membranes and in signaling. However, how plants regulate sphingolipid biosynthesis in the response to stress remains unclear. Here, we reveal that the plant hormone ethylene can modulate sphingolipid synthesis. The fungal toxin Fumonisin B1 (FB1) inhibits the activity of ceramide synthases, perturbing sphingolipid homeostasis, and thus inducing cell death. We used FB1 to test the role of ethylene signaling in sphingolipid synthesis in Arabidopsis thaliana. The etr1-1 and ein2 mutants, which have disrupted ethylene signaling, exhibited hypersensitivity to FB1; by contrast, the eto1-1 and ctr1-1 mutants, which have enhanced ethylene signaling, exhibited increased tolerance to FB1. Gene expression analysis showed that during FB1 treatment, transcripts of genes involved in de novo sphingolipid biosynthesis were down-regulated in ctr1-1 mutants but up-regulated in ein2 mutants. Strikingly, under normal conditions, ctr1-1 mutants contained less ceramides and hydroxyceramides, compared with wild type. After FB1 treatment, ctr1-1 and ein2 mutants showed a significant improvement in sphingolipid contents, except the ctr1-1 mutants showed little change in hydroxyceramide levels. Treatment of wild-type seedlings with the ethylene precursor 1-aminocyclopropane carboxylic acid down-regulated genes involved in the sphingolipid de novo biosynthesis pathway, thus reducing sphingolipid contents and partially rescuing FB1-induced cell death. Taking these results together, we propose that ethylene modulates sphingolipids by regulating the expression of genes related to the de novo biosynthesis of sphingolipids. PMID:26734030

  1. Phosphatidylglycerol biosynthesis is required for the development of embryos and normal membrane structures of chloroplasts and mitochondria in Arabidopsis.

    PubMed

    Tanoue, Ryo; Kobayashi, Megumi; Katayama, Kenta; Nagata, Noriko; Wada, Hajime

    2014-05-01

    Phosphatidylglycerophosphate (PGP) synthase, encoded by PGP1 and PGP2 in Arabidopsis, catalyzes a committed step in the biosynthesis of phosphatidylglycerol (PG). In this study, we isolated a pgp1pgp2 double mutant of Arabidopsis to study the function of PG. In this mutant, embryo development was delayed and the majority of seeds did not germinate. Thylakoid membranes did not develop in plastids, mitochondrial membrane structures were abnormal in the mutant embryos, and radiolabeling of phospholipids showed that radioactivity was not significantly incorporated into PG. These results demonstrated that PG biosynthesis is essential for the development of embryos and normal membrane structures of chloroplasts and mitochondria. PMID:24632290

  2. Structural Characterization of Arabidopsis Leaf Arabinogalactan Polysaccharides1[W

    PubMed Central

    Tryfona, Theodora; Liang, Hui-Chung; Kotake, Toshihisa; Tsumuraya, Yoichi; Stephens, Elaine; Dupree, Paul

    2012-01-01

    Proteins decorated with arabinogalactan (AG) have important roles in cell wall structure and plant development, yet the structure and biosynthesis of this polysaccharide are poorly understood. To facilitate the analysis of biosynthetic mutants, water-extractable arabinogalactan proteins (AGPs) were isolated from the leaves of Arabidopsis (Arabidopsis thaliana) plants and the structure of the AG carbohydrate component was studied. Enzymes able to hydrolyze specifically AG were utilized to release AG oligosaccharides. The released oligosaccharides were characterized by high-energy matrix-assisted laser desorption ionization-collision-induced dissociation mass spectrometry and polysaccharide analysis by carbohydrate gel electrophoresis. The Arabidopsis AG is composed of a β-(1→3)-galactan backbone with β-(1→6)-d-galactan side chains. The β-(1→6)-galactan side chains vary in length from one to over 20 galactosyl residues, and they are partly substituted with single α-(1→3)-l-arabinofuranosyl residues. Additionally, a substantial proportion of the β-(1→6)-galactan side chain oligosaccharides are substituted at the nonreducing termini with single 4-O-methyl-glucuronosyl residues via β-(1→6)-linkages. The β-(1→6)-galactan side chains are occasionally substituted with α-l-fucosyl. In the fucose-deficient murus1 mutant, AGPs lack these fucose modifications. This work demonstrates that Arabidopsis mutants in AGP structure can be identified and characterized. The detailed structural elucidation of the AG polysaccharides from the leaves of Arabidopsis is essential for insights into the structure-function relationships of these molecules and will assist studies on their biosynthesis. PMID:22891237

  3. 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 ...

  4. Genetic control of polar cell expansion in Arabidopsis thaliana

    SciTech Connect

    Schiefelbein, J.; Ford, S. ); Somerville, C. )

    1990-05-01

    Certain plant cells, like root hairs and pollen tubes, exhibit polar cell growth, with expansion limited to the tip of the growing cell. In order to understand the mechanisms regulating polar cell expansion, we are studying the process of root hair elongation in Arabidopsis thaliana. By visually screening roots from 12,000 mutagenized Arabidopsis seedlings on Petri dishes, more than 40 root hair mutants have been identified. We have focused our attention on mutants that possess nuclear recessive mutations in three genes (RHD2, RHD3, and RDH4) that appear to be involved in controlling polar cell growth in root hairs. We are currently using cellular, genetic, and molecular approaches to understand these genes' normal roles in root hair elongation.

  5. Arabidopsis YAK1 regulates abscisic acid response and drought resistance.

    PubMed

    Kim, Dongjin; Ntui, Valentine Otang; Xiong, Liming

    2016-07-01

    Abscisic acid (ABA) is an important phytohormone that controls several plant processes such as seed germination, seedling growth, and abiotic stress response. Here, we report that AtYak1 plays an important role in ABA signaling and postgermination growth in Arabidopsis. AtYak1 knockout mutant plants were hyposensitive to ABA inhibition of seed germination, cotyledon greening, seedling growth, and stomatal movement. atyak1-1 mutant plants display reduced drought stress resistance, as evidenced by water loss rate and survival rate. Molecular genetic analysis revealed that AtYak1 deficiency led to elevated expression of stomatal-related gene, MYB60, and down-regulation of several stress-responsive genes. Altogether, these results indicate that AtYak1 plays a role as a positive regulator in ABA-mediated drought response in Arabidopsis. PMID:27264339

  6. Bacterial RNAs activate innate immunity in Arabidopsis.

    PubMed

    Lee, Boyoung; Park, Yong-Soon; Lee, Soohyun; Song, Geun Cheol; Ryu, Choong-Min

    2016-01-01

    The common molecular patterns of microbes play a critical role in the regulation of plant innate immunity. However, little is known about the role of nucleic acids in this process in plants. We pre-infiltrated Arabidopsis leaves with total RNAs from Pseudomonas syringae pv. tomato DC3000 (Pto DC3000) and subsequently inoculated these plants with the same bacterial cells. Total Pto DC3000 RNAs pre-infiltrated into Arabidopsis leaves elicited plant immune responses against Pto DC3000. However, sheared RNAs and RNase A application failed to induce immunity, suggesting that intact bacterial RNAs function in plant innate immunity. This notion was supported by the positive regulation of superoxide anion levels, callose deposition, two mitogen-activated protein kinases and defense-related genes observed in bacterial RNA-pre-treated leaves. Intriguingly, the Pto DC3000 population was not compromised in known pattern recognition receptor mutants for chitin, flagellin and elongation factor-Tu (EF-Tu). Plant defense-related mutant analyses further revealed that bacterial RNA-elicited innate immunity was normally required for salicylic and jasmonic acid signaling. Notably, among total RNAs, the abundant bacterial RNA species 16S and 23S ribosomal RNAs were the major determinants of this response. Our findings provide evidence that bacterial RNA serves as a microbe-associated molecular pattern in plants. PMID:26499893

  7. 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

  8. Adenosine-5'-phosphosulfate kinase is essential for Arabidopsis viability.

    PubMed

    Mugford, Sarah G; Matthewman, Colette A; Hill, Lionel; Kopriva, Stanislav

    2010-01-01

    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. PMID:19903478

  9. GOLDEN2-LIKE transcription factors coordinate the tolerance to Cucumber mosaic virus in Arabidopsis.

    PubMed

    Han, Xue-Ying; Li, Peng-Xu; Zou, Li-Juan; Tan, Wen-Rong; Zheng, Ting; Zhang, Da-Wei; Lin, Hong-Hui

    2016-09-01

    Arabidopsis thaliana GOLDEN2-LIKE (GLKs) transcription factors play important roles in regulation of photosynthesis-associated nuclear genes, as well as participate in chloroplast development. However, the involvement of GLKs in plants resistance to virus remains largely unknown. Here, the relationship between GLKs and Cucumber mosaic virus (CMV) stress response was investigated. Our results showed that the Arabidopsis glk1glk2 double-mutant was more susceptible to CMV infection and suffered more serious damages (such as higher oxidative damages, more compromised in PSII photochemistry and more reactive oxygen species accumulation) when compared with the wild-type plants. Interestingly, there was little difference between single mutant (glk1 or glk2) and wild-type plants in response to CMV infection, suggesting GLK1 and GLK2 might function redundant in virus resistance in Arabidopsis. Furthermore, the induction of antioxidant system and defense-associated genes expression in the double mutant were inhibited when compared with single mutant or wild-type plants after CMV infection. Further evidences showed that salicylic acid (SA) and jasmonic acid (JA) might be involved in GLKs-mediated virus resistance, as SA or JA level and synthesis-related genes transcription were impaired in glk1glk2 mutant. Taken together, our results indicated that GLKs played a positively role in virus resistance in Arabidopsis. PMID:27346129

  10. 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.

  11. 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. PMID:26990896

  12. Stress physiology functions of the Arabidopsis histidine kinase cytokinin receptors.

    PubMed

    Kumar, M Nagaraj; Verslues, Paul E

    2015-07-01

    Cytokinin signaling has complex effects on abiotic stress responses that remain to be fully elucidated. The Arabidopsis histidine kinases (AHKs), AHK2, AHK3 and CRE1 (cytokinin response1/AHK4) are the principle cytokinin receptors of Arabidopsis. Using a set of ahk mutants, we found dramatic differences in response to low water potential and salt stress among the AHKs. ahk3-3 mutants had increased root elongation after transfer to low water potential media. Conversely ahk2-2 was hypersensitive to salt stress in terms of root growth and fresh weight and accumulated higher than wild-type levels of proline specifically under salt stress. Strongly reduced proline accumulation in ahk double mutants after low water potential treatment indicated a more general role of cytokinin signaling in proline metabolism. Reduced P5CS1 (Δ(1) -pyrroline-5-carboxylate synthetase1) gene expression may have contributed to this reduced proline accumulation. Low water potential phenotypes of ahk mutants were not caused by altered abscisic acid (ABA) accumulation as all ahk mutants had wild-type ABA levels, despite the observation that ahk double mutants had reduced NCED3 (9-cis-epoxycartenoid dioxygenase3) expression when exposed to low water potential. No difference in osmoregulatory solute accumulation was detected in any of the ahk mutants indicating that they do not affect drought responsive osmotic adjustment. Overall, our examination of ahk mutants found specific phenotypes associated with AHK2 and AHK3 as well as a general function of cytokinin signaling in proline accumulation and low water potential induction of P5CS1 and NCED3 expression. These results show the stress physiology function of AHKs at a new level of detail. PMID:25263537

  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. Arabidopsis myosin XI: a motor rules the tracks.

    PubMed

    Cai, Chao; Henty-Ridilla, Jessica L; Szymanski, Daniel B; Staiger, Christopher J

    2014-11-01

    Plant cell expansion relies on intracellular trafficking of vesicles and macromolecules, which requires myosin motors and a dynamic actin network. Arabidopsis (Arabidopsis thaliana) myosin XI powers the motility of diverse cellular organelles, including endoplasmic reticulum, Golgi, endomembrane vesicles, peroxisomes, and mitochondria. Several recent studies show that there are changes in actin organization and dynamics in myosin xi mutants, indicating that motors influence the molecular tracks they use for transport. However, the mechanism by which actin organization and dynamics are regulated by myosin XI awaits further detailed investigation. Here, using high spatiotemporal imaging of living cells, we quantitatively assessed the architecture and dynamic behavior of cortical actin arrays in a mutant with three Myosin XI (XI-1, XI-2, and XI-K) genes knocked out (xi3KO). In addition to apparent reduction of organ and cell size, the mutant showed less dense and more bundled actin filament arrays in epidermal cells. Furthermore, the overall actin dynamicity was significantly inhibited in the xi3KO mutant. Because cytoskeletal remodeling is contributed mainly by filament assembly/disassembly and translocation/buckling, we also examined the dynamic behavior of individual actin filaments. We found that the xi3KO mutant had significantly decreased actin turnover, with a 2-fold reduction in filament severing frequency. Moreover, quantitative analysis of filament shape change over time revealed that myosin XI generates the force for buckling and straightening of both single actin filaments and actin bundles. Thus, our data provide genetic evidence that three Arabidopsis class XI myosins contribute to actin remodeling by stimulating turnover and generating the force for filament shape change. PMID:25237128

  15. Functional and Genetic Analysis Identify a Role for Arabidopsis ARGONAUTE5 in Antiviral RNA Silencing

    PubMed Central

    Brosseau, Chantal; Moffett, Peter

    2015-01-01

    RNA silencing functions as an antiviral defense through the action of DICER-like (DCL) and ARGONAUTE (AGO) proteins. In turn, plant viruses have evolved strategies to counteract this defense mechanism, including the expression of suppressors of RNA silencing. Potato virus X (PVX) does not systemically infect Arabidopsis thaliana Columbia-0, but is able to do so effectively in mutants lacking at least two of the four Arabidopsis DCL proteins. PVX can also infect Arabidopsis ago2 mutants, albeit less effectively than double DCL mutants, suggesting that additional AGO proteins may mediate anti-viral defenses. Here we show, using functional assays, that all Arabidopsis AGO proteins have the potential to target PVX lacking its viral suppressor of RNA silencing (VSR), P25, but that only AGO2 and AGO5 are able to target wild-type PVX. However, P25 directly affects only a small subset of AGO proteins, and we present evidence indicating that its protective effect is mediated by precluding AGO proteins from accessing viral RNA, as well as by directly inhibiting the RNA silencing machinery. In agreement with functional assays, we show that Potexvirus infection induces AGO5 expression and that both AGO2 and AGO5 are required for full restriction of PVX infection in systemic tissues of Arabidopsis. PMID:26023161

  16. Phosphate availability alters architecture and causes changes in hormone sensitivity in the Arabidopsis root system.

    PubMed

    López-Bucio, José; Hernández-Abreu, Esmeralda; Sánchez-Calderón, Lenin; Nieto-Jacobo, María Fernanda; Simpson, June; Herrera-Estrella, Luis

    2002-05-01

    The postembryonic developmental program of the plant root system is plastic and allows changes in root architecture to adapt to environmental conditions such as water and nutrient availability. Among essential nutrients, phosphorus (P) often limits plant productivity because of its low mobility in soil. Therefore, the architecture of the root system may determine the capacity of the plant to acquire this nutrient. We studied the effect of P availability on the development of the root system in Arabidopsis. We found that at P-limiting conditions (<50 microM), the Arabidopsis root system undergoes major architectural changes in terms of lateral root number, lateral root density, and primary root length. Treatment with auxins and auxin antagonists indicate that these changes are related to an increase in auxin sensitivity in the roots of P-deprived Arabidopsis seedlings. It was also found that the axr1-3, axr2-1, and axr4-1 Arabidopsis mutants have normal responses to low P availability conditions, whereas the iaa28-1 mutant shows resistance to the stimulatory effects of low P on root hair and lateral root formation. Analysis of ethylene signaling mutants and treatments with 1-aminocyclopropane-1-carboxylic acid showed that ethylene does not promote lateral root formation under P deprivation. These results suggest that in Arabidopsis, auxin sensitivity may play a fundamental role in the modifications of root architecture by P availability. PMID:12011355

  17. Functional and Genetic Analysis Identify a Role for Arabidopsis ARGONAUTE5 in Antiviral RNA Silencing.

    PubMed

    Brosseau, Chantal; Moffett, Peter

    2015-06-01

    RNA silencing functions as an antiviral defense through the action of DICER-like (DCL) and ARGONAUTE (AGO) proteins. In turn, plant viruses have evolved strategies to counteract this defense mechanism, including the expression of suppressors of RNA silencing. Potato virus X (PVX) does not systemically infect Arabidopsis thaliana Columbia-0, but is able to do so effectively in mutants lacking at least two of the four Arabidopsis DCL proteins. PVX can also infect Arabidopsis ago2 mutants, albeit less effectively than double DCL mutants, suggesting that additional AGO proteins may mediate anti-viral defenses. Here we show, using functional assays, that all Arabidopsis AGO proteins have the potential to target PVX lacking its viral suppressor of RNA silencing (VSR), P25, but that only AGO2 and AGO5 are able to target wild-type PVX. However, P25 directly affects only a small subset of AGO proteins, and we present evidence indicating that its protective effect is mediated by precluding AGO proteins from accessing viral RNA, as well as by directly inhibiting the RNA silencing machinery. In agreement with functional assays, we show that Potexvirus infection induces AGO5 expression and that both AGO2 and AGO5 are required for full restriction of PVX infection in systemic tissues of Arabidopsis. PMID:26023161

  18. 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. PMID:24057358

  19. P-proteins in Arabidopsis are heteromeric structures involved in rapid sieve tube sealing

    PubMed Central

    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. PMID:23840197

  20. Analysis of Arabidopsis glutathione-transferases in yeast.

    PubMed

    Krajewski, Matthias P; Kanawati, Basem; Fekete, Agnes; Kowalski, Natalie; Schmitt-Kopplin, Philippe; Grill, Erwin

    2013-07-01

    The genome of Arabidopsis thaliana encodes 54 functional glutathione transferases (GSTs), classified in seven clades. Although plant GSTs have been implicated in the detoxification of xenobiotics, such as herbicides, extensive redundancy within this large gene family impedes a functional analysis in planta. In this study, a GST-deficient yeast strain was established as a system for analyzing plant GSTs that allows screening for GST substrates and identifying substrate preferences within the plant GST family. To this end, five yeast genes encoding GSTs and GST-related proteins were simultaneously disrupted. The resulting yeast quintuple mutant showed a strongly reduced conjugation of the GST substrates 1-chloro-2,4-dinitrobenzene (CDNB) and 4-chloro-7-nitro-2,1,3-benzoxadiazole (NBD-Cl). Consistently, the quintuple mutant was hypersensitive to CDNB, and this phenotype was complemented by the inducible expression of Arabidopsis GSTs. The conjugating activity of the plant GSTs was assessed by in vitro enzymatic assays and via analysis of exposed yeast cells. The formation of glutathione adducts with dinitrobenzene was unequivocally verified by stable isotope labeling and subsequent accurate ultrahigh-resolution mass spectrometry (ICR-FTMS). Analysis of Arabidopsis GSTs encompassing six clades and 42 members demonstrated functional expression in yeast by using CDNB and NBD-Cl as model substrates. Subsequently, the established yeast system was explored for its potential to screen the Arabidopsis GST family for conjugation of the fungicide anilazine. Thirty Arabidopsis GSTs were identified that conferred increased levels of glutathionylated anilazine. Efficient anilazine conjugation was observed in the presence of the phi, tau, and theta clade GSTs including AtGSTF2, AtGSTF4, AtGSTF6, AtGSTF8, AtGSTF10, and AtGSTT2, none of which had previously been known to contribute to fungicide detoxification. ICR-FTMS analysis of yeast extracts allowed the simultaneous detection and

  1. Suspensor Length Determines Developmental Progression of the Embryo in Arabidopsis1[W][OA

    PubMed Central

    Babu, Yashodar; Musielak, Thomas; Henschen, Agnes; Bayer, Martin

    2013-01-01

    The first structure that differentiates during plant embryogenesis is the extra-embryonic suspensor that positions the embryo in the lumen of the seed. A central role in nutrient transport has been ascribed to the suspensor in species with prominent suspensor structures. Little is known, however, about what impact the size of the rather simple Arabidopsis (Arabidopsis thaliana) suspensor has on embryogenesis. Here, we describe mutations in the predicted exo-polygalacturonase gene NIMNA (NMA) that lead to cell elongation defects in the early embryo and markedly reduced suspensor length. Mutant nma embryos develop slower than wild-type embryos, and we could observe a similar developmental delay in another mutant with shorter suspensors. Interestingly, for both genes, the paternal allele has a stronger influence on the embryonic phenotype. We conclude that the length of the suspensor is crucial for fast developmental progression of the embryo in Arabidopsis. PMID:23709666

  2. 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...

  3. Arabidopsis Nucleolin Affects Plant Development and Patterning1[W][OA

    PubMed Central

    Petricka, Jalean Joyanne; Nelson, Timothy Mark

    2007-01-01

    Nucleolin is a major nucleolar protein implicated in many aspects of ribosomal biogenesis, including early events such as processing of the large 35S preribosomal RNA. We found that the Arabidopsis (Arabidopsis thaliana) parallel1 (parl1) mutant, originally identified by its aberrant leaf venation, corresponds to the Arabidopsis nucleolin gene. parl1 mutants display parallel leaf venation, aberrant localization of the provascular marker Athb8:β-glucuronidase, the auxin-sensitive reporter DR5:β-glucuronidase, and auxin-dependent growth defects. PARL1 is highly similar to the yeast (Saccharomyces cerevisiae) nucleolin NUCLEAR SIGNAL RECOGNITION 1 (NSR1) multifunctional protein; the Arabidopsis PARL1 gene can rescue growth defects of yeast nsr1 null mutants. This suggests that PARL1 protein may have roles similar to those of the yeast nucleolin in nuclear signal recognition, ribosomal processing, and ribosomal subunit accumulation. Based on the range of auxin-related defects in parl1 mutants, we propose that auxin-dependent organ growth and patterning is highly sensitive to the efficiency of nucleolin-dependent ribosomal processing. PMID:17369435

  4. AtHSPR may function in salt-induced cell death and ER stress in Arabidopsis.

    PubMed

    Yang, Tao; Zhang, Peng; Wang, Chongying

    2016-07-01

    Salt stress is a harmful and global abiotic stress to plants and has an adverse effect on all physiological processes of plants. Recently, we cloned and identified a novel AtHSPR (Arabidopsis thaliana Heat Shock Protein Related), which encodes a nuclear-localized protein with ATPase activity, participates in salt and drought tolerance in Arabidopsis. Transcript profiling analysis revealed a differential expression of genes involved in accumulation of reactive oxygen species (ROS), abscisic acid (ABA) signaling, stress response and photosynthesis between athspr mutant and WT under salt stress. Here, we provide further analysis of the data showing the regulation of salt-induced cell death and endoplasmic reticulum (ER) stress response in Arabidopsis and propose a hypothetical model for the role of AtHSPR in the regulation of the salt tolerance in Arabidopsis. PMID:27302034

  5. 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. PMID:19015126

  6. Requirement of Functional Ethylene-Insensitive 2 Gene for Efficient Resistance of Arabidopsis to Infection by Botrytis cinerea1

    PubMed Central

    Thomma, Bart P.H.J.; Eggermont, Kristel; Tierens, Koenraad F.M.-J.; Broekaert, Willem F.

    1999-01-01

    Inoculation of wild-type Arabidopsis plants with the fungus Alternaria brassicicola results in systemic induction of genes encoding a plant defensin (PDF1.2), a basic chitinase (PR-3), and an acidic hevein-like protein (PR-4). Pathogen-induced induction of these three genes is almost completely abolished in the ethylene-insensitive Arabidopsis mutant ein2-1. This indicates that a functional ethylene signal transduction component (EIN2) is required in this response. The ein2-1 mutants were found to be markedly more susceptible than wild-type plants to infection by two different strains of the gray mold fungus Botrytis cinerea. In contrast, no increased fungal colonization of ein2-1 mutants was observed after challenge with avirulent strains of either Peronospora parasitica or A. brassicicola. Our data support the conclusion that ethylene-controlled responses play a role in resistance of Arabidopsis to some but not all types of pathogens. PMID:10594097

  7. A new cyanogenic metabolite in Arabidopsis required for inducible pathogen defence.

    PubMed

    Rajniak, Jakub; Barco, Brenden; Clay, Nicole K; Sattely, Elizabeth S

    2015-09-17

    Thousands of putative biosynthetic genes in Arabidopsis thaliana have no known function, which suggests that there are numerous molecules contributing to plant fitness that have not yet been discovered. Prime among these uncharacterized genes are cytochromes P450 upregulated in response to pathogens. Here we start with a single pathogen-induced P450 (ref. 5), CYP82C2, and use a combination of untargeted metabolomics and coexpression analysis to uncover the complete biosynthetic pathway to 4-hydroxyindole-3-carbonyl nitrile (4-OH-ICN), a previously unknown Arabidopsis metabolite. This metabolite harbours cyanogenic functionality that is unprecedented in plants and exceedingly rare in nature; furthermore, the aryl cyanohydrin intermediate in the 4-OH-ICN pathway reveals a latent capacity for cyanogenic glucoside biosynthesis in Arabidopsis. By expressing 4-OH-ICN biosynthetic enzymes in Saccharomyces cerevisiae and Nicotiana benthamiana, we reconstitute the complete pathway in vitro and in vivo and validate the functions of its enzymes. Arabidopsis 4-OH-ICN pathway mutants show increased susceptibility to the bacterial pathogen Pseudomonas syringae, consistent with a role in inducible pathogen defence. Arabidopsis has been the pre-eminent model system for studying the role of small molecules in plant innate immunity; our results uncover a new branch of indole metabolism distinct from the canonical camalexin pathway, and support a role for this pathway in the Arabidopsis defence response. These results establish a more complete framework for understanding how the model plant Arabidopsis uses small molecules in pathogen defence. PMID:26352477

  8. Arabidopsis Rab Geranylgeranyltransferases Demonstrate Redundancy and Broad Substrate Specificity in Vitro.

    PubMed

    Shi, Wan; Zeng, Qin; Kunkel, Barbara N; Running, Mark P

    2016-01-15

    Posttranslational lipid modifications mediate the membrane attachment of Rab GTPases, facilitating their function in regulating intracellular vesicular trafficking. In Arabidopsis, most Rab GTPases have two C-terminal cysteines and potentially can be double-geranylgeranylated by heterodimeric Rab geranylgeranyltransferases (Rab-GGTs). Genes encoding two putative α subunits and two putative β subunits of Rab-GGTs have been annotated in the Arabidopsis thaliana genome, but little is known about Rab-GGT activity in Arabidopsis. In this study, we demonstrate that four different heterodimers can be formed between putative Arabidopsis Rab-GGT α subunits RGTA1/RGTA2 and β subunits RGTB1/RGTB2, but only RGTA1·RGTB1 and RGTA1·RGTB2 exhibit bona fide Rab-GGT activity, and they are biochemically redundant in vitro. We hypothesize that RGTA2 function might be disrupted by a 12-amino acid insertion in a conserved motif. We present evidence that Arabidopsis Rab-GGTs may have preference for prenylation of C-terminal cysteines in particular positions. We also demonstrate that Arabidopsis Rab-GGTs can not only prenylate a great variety of Rab GTPases in the presence of Rab escort protein but, unlike Rab-GGT in yeast and mammals, can also prenylate certain non-Rab GTPases independently of Rab escort protein. Our findings may help to explain some of the phenotypes of Arabidopsis protein prenyltransferase mutants. PMID:26589801

  9. Contribution of Structural Chromosome Mutants to the Study of Meiosis in Plants.

    PubMed

    Naranjo, Tomás

    2015-01-01

    Dissection of the molecular mechanisms underlying the transition through the complex events of the meiotic process requires the use of gene mutants or RNAi-mediated gene silencing. A considerable number of meiotic mutants have been isolated in plant species such as Arabidopsis thaliana, maize or rice. However, structural chromosome mutants are also important for the identification of the role developed by different chromosome domains in the meiotic process. This review summarizes the contribution of studies carried out in plants using structural chromosome variations. Meiotic events concerning the search of the homologous partner, the control of number and distribution of chiasmata, the mechanism of pairing correction, and chromosome segregation are considered. PMID:26658116

  10. Identification and characterization of Arabidopsis gibberellin receptors.

    PubMed

    Nakajima, Masatoshi; Shimada, Asako; Takashi, Yoshiyuki; Kim, Young-Cheon; Park, Seung-Hyun; Ueguchi-Tanaka, Miyako; Suzuki, Hiroyuki; Katoh, Etsuko; Iuchi, Satoshi; Kobayashi, Masatomo; Maeda, Tatsuya; Matsuoka, Makoto; Yamaguchi, Isomaro

    2006-06-01

    Three gibberellin (GA) receptor genes (AtGID1a, AtGID1b and AtGID1c), each an ortholog of the rice GA receptor gene (OsGID1), were cloned from Arabidopsis, and the characteristics of their recombinant proteins were examined. The GA-binding activities of the three recombinant proteins were confirmed by an in vitro assay. Biochemical analyses revealed similar ligand selectivity among the recombinants, and all recombinants showed higher affinity to GA(4) than to other GAs. AtGID1b was unique in its binding affinity to GA(4) and in its pH dependence when compared with the other two, by only showing binding in a narrow pH range (pH 6.4-7.5) with 10-fold higher affinity (apparent K(d) for GA(4) = 3 x 10(-8) m) than AtGID1a and AtGID1c. A two-hybrid yeast system only showed in vivo interaction in the presence of GA(4) between each AtGID1 and the Arabidopsis DELLA proteins (AtDELLAs), negative regulators of GA signaling. For this interaction with AtDELLAs, AtGID1b required only one-tenth of the amount of GA(4) that was necessary for interaction between the other AtGID1s and AtDELLAs, reflecting its lower K(d) value. AtDELLA boosted the GA-binding activity of AtGID1 in vitro, which suggests the formation of a complex between AtDELLA and AtGID1-GA that binds AtGID1 to GA more tightly. The expression of each AtGID1 clone in the rice gid1-1 mutant rescued the GA-insensitive dwarf phenotype. These results demonstrate that all three AtGID1s functioned as GA receptors in Arabidopsis. PMID:16709201

  11. The regulatory PII protein controls arginine biosynthesis in Arabidopsis.

    PubMed

    Ferrario-Méry, Sylvie; Besin, Evelyne; Pichon, Olivier; Meyer, Christian; Hodges, Michael

    2006-04-01

    In higher plants, PII is a nuclear-encoded plastid protein which is homologous to bacterial PII signalling proteins known to be involved in the regulation of nitrogen metabolism. A reduced ornithine, citrulline and arginine accumulation was observed in two Arabidopsis PII knock-out mutants in response to NH4+ resupply after N starvation. This difference could be explained by the regulation of a key enzyme of the arginine biosynthesis pathway, N-acetyl glutamate kinase (NAGK) by PII. In vitro assays using purified recombinant proteins showed the catalytic activation of Arabidopsis NAGK by PII giving the first evidence of a physiological role of the PII protein in higher plants. Using Arabidopsis transcriptome microarray (CATMA) and RT-PCR analyses, it was found that none of the genes involved in the arginine biosynthetic or catabolic pathways were differentially expressed in a PII knock-out mutant background. In conclusion, the observed changes in metabolite levels can be explained by the reduced activation of NAGK by PII. PMID:16545809

  12. Morphological and physiological traits of three major Arabidopsis thaliana accessions.

    PubMed

    Passardi, Filippo; Dobias, Jan; Valério, Luisa; Guimil, Sonia; Penel, Claude; Dunand, Christophe

    2007-08-01

    Arabidopsis is currently the most studied organism in plant biology. Its short life cycle and small genome size have rendered it one of the principal model systems. Additionally, numerous large T-DNA insertion mutant collections are available. The advent of molecular biology and the completion of the Arabidopsis genome sequence have contributed to helping researchers discover a large variety of mutants identified for their phenotypes. Yet, it is important to consider that natural phenotypic variations exist and appear in natural ecotypes, differing greatly in several traits. Although there are a vast number of ecotypes available, only a few have been extensively studied, and some have been created in laboratories. In order to identify new phenotypic differences, we chose to study the differences observed between three ecotypes: Columbia (Col-0), Landsberg erecta (Laer-0) and Wassilewskija (Ws-0). Our research focuses on observable morphological traits throughout plant growth and development along the entire plant life cycle. We then attempted to shed some light on phenotypic discrepancies through the study of the class III peroxidase protein family, which is involved in many aspects of plant growth and tissue differentiation. Both morphological and molecular aspects reveal that there are major variations between ecotypes, hence indicating a possibly interesting heterotic effect in the F1 from crosses between different Arabidopsis ecotypes. PMID:16904792

  13. Genetic analysis of growth-regulator-induced parthenocarpy in Arabidopsis.

    PubMed

    Vivian-Smith, A; Koltunow, A M

    1999-10-01

    In Arabidopsis, seedless silique development or parthenocarpy can be induced by the application of various plant growth regulators (PGRs) to unfertilized pistils. Ecotype-specific responses were observed in the Arabidopsis ecotypes Columbia and Landsberg relative to the type of PGR and level applied. The parthenocarpic response was greatest in ecotype Landsberg, and comparisons of fruit growth and morphology were studied primarily in this ecotype. Gibberellic acid application (10 micromol pistil(-1)) caused development similar to that in pollinated pistils, while benzyladenine (1 micromol pistil(-1)) and naphthylacetic acid (10 micromol pistil(-1)) treatment produced shorter siliques. Naphthylacetic acid primarily modified mesocarp cell expansion. Arabidopsis mutants were employed to examine potential dependencies on gibberellin biosynthesis (ga1-3, ga4-1, and ga5-1) and perception (spy-4 and gai) during parthenocarpic silique development. Emasculated spy-4 pistils were neither obviously parthenocarpic nor deficient in PGR perception. By contrast, emasculated gai mutants did not produce parthenocarpic siliques following gibberellic acid application, but silique development occurred following pollination or application of auxin and cytokinin. Pollinated gai siliques had decreased cell numbers and morphologically resembled auxin-induced parthenocarpic siliques. This shows that a number of independent and possibly redundant pathways can direct hormone-induced parthenocarpy, and that endogenous gibberellins play a role in regulating cell expansion and promoting cell division in carpels. PMID:10517835

  14. 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.

  15. Molecular Genetic Analysis of Phototropism in Arabidopsis

    PubMed Central

    Sakai, Tatsuya; Haga, Ken

    2012-01-01

    Plant life is strongly dependent on the environment, and plants regulate their growth and development in response to many different environmental stimuli. One of the regulatory mechanisms involved in these responses is phototropism, which allows plants to change their growth direction in response to the location of the light source. Since the study of phototropism by Darwin, many physiological studies of this phenomenon have been published. Recently, molecular genetic analyses of Arabidopsis have begun to shed light on the molecular mechanisms underlying this response system, including phototropin blue light photoreceptors, phototropin signaling components, auxin transporters, auxin action mechanisms and others. This review highlights some of the recent progress that has been made in further elucidating the phototropic response, with particular emphasis on mutant phenotypes. PMID:22864452

  16. 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. PMID:23834713

  17. 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. PMID:26586261

  18. A Molecular Portrait of Arabidopsis Meiosis

    PubMed Central

    Ma, Hong

    2006-01-01

    Meiosis is essential for eukaryotic sexual reproduction and important for genetic diversity among individuals. Efforts during the last decade in Arabidopsis have greatly expanded our understanding of the molecular basis of plant meiosis, which has traditionally provided much information about the cytological description of meiosis. Through both forward genetic analysis of mutants with reduced fertility and reverse genetic studies of homologs of known meiotic genes, we now have a basic knowledge about genes important for meiotic recombination and its relationship to pairing and synapsis, critical processes that ensure proper homolog segregation. In addition, several genes affecting meiotic progression, spindle assembly, chromosome separation, and meiotic cytokinesis have also been uncovered and characterized. It is worth noting that Arabidopsis molecular genetic studies are also revealing secrets of meiosis that have not yet been recognized elsewhere among eukaryotes, including gene functions that might be unique to plants and those that are potentially shared with animals and fungi. As we enter the post-genomics era of plant biology, there is no doubt that the next ten years will see an even greater number of discoveries in this important area of plant development and cell biology. Abbreviations: DAPI, 4′,6-diamidino-2-phenylindole; DSB, double strand break; DSBR, double strand break repair; SC, synaptonemal complex; TEM, transmission electron microscopy PMID:22303228

  19. 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

  20. MTHFD1 controls DNA methylation in Arabidopsis.

    PubMed

    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

  1. 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

  2. Novel aspects of COP9 signalosome functions revealed through analysis of hypomorphic csn mutants

    PubMed Central

    Parker, Jane E

    2009-01-01

    The COP9 signalosome (CSN) is a conserved eukaryotic protein complex implicated in the regulation of cullin-RING type E3 ubiquitin ligases by cleaving the small peptide RUB/Nedd8 from cullins. However, detailed analysis of CSN physiological functions in Arabidopsis has been hampered by the early seedling-lethality of csn null mutants. We and others have now identified a number of viable hypomorphic csn mutants which start to reveal novel CSN-dependent activities in adult Arabidopsis plants.1 Here, we present a detailed comparative analysis of the csn5a-1 and csn2-5 mutants as a mean to improve understanding of CSN functions in plant cells. Our observations point to CSN-independent activities of CSN5 and suggest a role of the CSN in cytoskeleton assembly/organization. PMID:19847120

  3. 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

  4. Strigolactones Suppress Adventitious Rooting in Arabidopsis and Pea1[C][W][OA

    PubMed Central

    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

    2012-01-01

    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. PMID:22323776

  5. Phenome Analysis in Plant Species Using Loss-of-Function and Gain-of-Function Mutants

    PubMed Central

    Kuromori, Takashi; Takahashi, Shinya; Kondou, Youichi; Shinozaki, Kazuo; Matsui, Minami

    2009-01-01

    Analysis of genetic mutations is one of the most effective ways to investigate gene function. We now have methods that allow for mass production of mutant lines and cells in a variety of model species. Recently, large numbers of mutant lines have been generated by both ‘loss-of-function’ and ‘gain-of-function’ techniques. In parallel, phenotypic information covering various mutant resources has been acquired and released in web-based databases. As a result, significant progress in comprehensive pheno-type analysis is being made through the use of these tools. Arabidopsis and rice are two major model plant species in which genome sequencing projects have been completed. Arabidopsis is the most widely used experimental plant, with a large number of mutant resources and several examples of systematic phenotype analysis. Rice is a major crop species and is used as a model plant, with an increasing number of mutant resources. Other plant species are also being employed in functional genetics research. In this review, the present status of mutant resources for large-scale studies of gene function in plant research and the current perspective on using loss-of-function and gain-of-function mutants in phenome research will be discussed. PMID:19502383

  6. Perspectives on Systematic Analyses of Gene Function in Arabidopsis thaliana: New Tools, Topics and Trends

    PubMed Central

    Bolle, C; Schneider, A; Leister, D

    2011-01-01

    Since the sequencing of the nuclear genome of Arabidopsis thaliana ten years ago, various large-scale analyses of gene function have been performed in this model species. In particular, the availability of collections of lines harbouring random T-DNA or transposon insertions, which include mutants for almost all of the ~27,000 A. thaliana genes, has been crucial for the success of forward and reverse genetic approaches. In the foreseeable future, genome-wide phenotypic data from mutant analyses will become available for Arabidopsis, and will stimulate a flood of novel in-depth gene-function analyses. In this review, we consider the present status of resources and concepts for systematic studies of gene function in A. thaliana. Current perspectives on the utility of loss-of-function and gain-of-function mutants will be discussed in light of the genetic and functional redundancy of many A. thaliana genes. PMID:21886450

  7. 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.

  8. 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

  9. Trichome morphogenesis in Arabidopsis.

    PubMed Central

    Schwab, B; Folkers, U; Ilgenfritz, H; Hülskamp, M

    2000-01-01

    Trichomes (plant hairs) in Arabidopsis thaliana are large non-secreting epidermal cells with a characteristic three-dimensional architecture. Because trichomes are easily accessible to a combination of genetic, cell biological and molecular methods they have become an ideal model system to study various aspects of plant cell morphogenesis. In this review we will summarize recent progress in the understanding of trichome morphogenesis. PMID:11128981

  10. Sphingolipids in the Root Play an Important Role in Regulating the Leaf Ionome in Arabidopsis thaliana[W][OA

    PubMed Central

    Chao, Dai-Yin; Gable, Kenneth; Chen, Ming; Baxter, Ivan; Dietrich, Charles R.; Cahoon, Edgar B.; Guerinot, Mary Lou; Lahner, Brett; Lü, Shiyou; Markham, Jonathan E.; Morrissey, Joe; Han, Gongshe; Gupta, Sita D.; Harmon, Jeffrey M.; Jaworski, Jan G.; Dunn, Teresa M.; Salt, David E.

    2011-01-01

    Sphingolipid synthesis is initiated by condensation of Ser with palmitoyl-CoA producing 3-ketodihydrosphinganine (3-KDS), which is reduced by a 3-KDS reductase to dihydrosphinganine. Ser palmitoyltransferase is essential for plant viability. Arabidopsis thaliana contains two genes (At3g06060/TSC10A and At5g19200/TSC10B) encoding proteins with significant similarity to the yeast 3-KDS reductase, Tsc10p. Heterologous expression in yeast of either Arabidopsis gene restored 3-KDS reductase activity to the yeast tsc10Δ mutant, confirming both as bona fide 3-KDS reductase genes. Consistent with sphingolipids having essential functions in plants, double mutant progeny lacking both genes were not recovered from crosses of single tsc10A and tsc10B mutants. Although the 3-KDS reductase genes are functionally redundant and ubiquitously expressed in Arabidopsis, 3-KDS reductase activity was reduced to 10% of wild-type levels in the loss-of-function tsc10a mutant, leading to an altered sphingolipid profile. This perturbation of sphingolipid biosynthesis in the Arabidopsis tsc10a mutant leads an altered leaf ionome, including increases in Na, K, and Rb and decreases in Mg, Ca, Fe, and Mo. Reciprocal grafting revealed that these changes in the leaf ionome are driven by the root and are associated with increases in root suberin and alterations in Fe homeostasis. PMID:21421810

  11. 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

  12. User guide for mapping-by-sequencing in Arabidopsis

    PubMed Central

    2013-01-01

    Mapping-by-sequencing combines genetic mapping with whole-genome sequencing in order to accelerate mutant identification. However, application of mapping-by-sequencing requires decisions on various practical settings on the experimental design that are not intuitively answered. Following an experimentally determined recombination landscape of Arabidopsis and next generation sequencing-specific biases, we simulated more than 400,000 mapping-by-sequencing experiments. This allowed us to evaluate a broad range of different types of experiments and to develop general rules for mapping-by-sequencing in Arabidopsis. Most importantly, this informs about the properties of different crossing scenarios, the number of recombinants and sequencing depth needed for successful mapping experiments. PMID:23773572

  13. Cinnamate-4-hydroxylase expression in arabidopsis. Regulation in response to development and the environment

    SciTech Connect

    Bell-Lelong, D.A.; Cusumano, J.C.; Meyer, K.; Chapple, C.

    1997-03-01

    Cinnamate-r-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. 77 refs., 5 figs.

  14. Azospirillum brasilense ameliorates the response of Arabidopsis thaliana to drought mainly via enhancement of ABA levels.

    PubMed

    Cohen, Ana C; Bottini, Rubén; Pontin, Mariela; Berli, Federico J; Moreno, Daniela; Boccanlandro, Hernán; Travaglia, Claudia N; Piccoli, Patricia N

    2015-01-01

    Production of phytohormones is one of the main mechanisms to explain the beneficial effects of plant growth-promoting rhizobacteria (PGPR) such as Azospirillum sp. The PGPRs induce plant growth and development, and reduce stress susceptibility. However, little is known regarding the stress-related phytohormone abscisic acid (ABA) produced by bacteria. We investigated the effects of Azospirillum brasilense Sp 245 strain on Arabidopsis thaliana Col-0 and aba2-1 mutant plants, evaluating the morphophysiological and biochemical responses when watered and in drought. We used an in vitro-grown system to study changes in the root volume and architecture after inoculation with Azospirillum in Arabidopsis wild-type Col-0 and on the mutant aba2-1, during early growth. To examine Arabidopsis development and reproductive success as affected by the bacteria, ABA and drought, a pot experiment using Arabidopsis Col-0 plants was also carried out. Azospirillum brasilense augmented plant biomass, altered root architecture by increasing lateral roots number, stimulated photosynthetic and photoprotective pigments and retarded water loss in correlation with incremented ABA levels. As well, inoculation improved plants seed yield, plants survival, proline levels and relative leaf water content; it also decreased stomatal conductance, malondialdehyde and relative soil water content in plants submitted to drought. Arabidopsis inoculation with A. brasilense improved plants performance, especially in drought. PMID:24796562

  15. Arabidopsis NATA1 Acetylates Putrescine and Decreases Defense-Related Hydrogen Peroxide Accumulation.

    PubMed

    Lou, Yann-Ru; Bor, Melike; Yan, Jian; Preuss, Aileen S; Jander, Georg

    2016-06-01

    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

  16. 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

  17. 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

  18. Arabidopsis heterotrimeric G-protein regulates cell wall defense and resistance to necrotrophic fungi.

    PubMed

    Delgado-Cerezo, Magdalena; Sánchez-Rodríguez, Clara; Escudero, Viviana; Miedes, Eva; Fernández, Paula Virginia; Jordá, Lucía; Hernández-Blanco, Camilo; Sánchez-Vallet, Andrea; Bednarek, Pawel; Schulze-Lefert, Paul; Somerville, Shauna; Estevez, José Manuel; Persson, Staffan; Molina, Antonio

    2012-01-01

    The Arabidopsis heterotrimeric G-protein controls defense responses to necrotrophic and vascular fungi. The agb1 mutant impaired in the Gβ subunit displays enhanced susceptibility to these pathogens. Gβ/AGB1 forms an obligate dimer with either one of the Arabidopsis Gγ subunits (γ1/AGG1 and γ2/AGG2). Accordingly, we now demonstrate that the agg1 agg2 double mutant is as susceptible as agb1 plants to the necrotrophic fungus Plectosphaerella cucumerina. To elucidate the molecular basis of heterotrimeric G-protein-mediated resistance, we performed a comparative transcriptomic analysis of agb1-1 mutant and wild-type plants upon inoculation with P. cucumerina. This analysis, together with metabolomic studies, demonstrated that G-protein-mediated resistance was independent of defensive pathways required for resistance to necrotrophic fungi, such as the salicylic acid, jasmonic acid, ethylene, abscisic acid, and tryptophan-derived metabolites signaling, as these pathways were not impaired in agb1 and agg1 agg2 mutants. Notably, many mis-regulated genes in agb1 plants were related with cell wall functions, which was also the case in agg1 agg2 mutant. Biochemical analyses and Fourier Transform InfraRed (FTIR) spectroscopy of cell walls from G-protein mutants revealed that the xylose content was lower in agb1 and agg1 agg2 mutants than in wild-type plants, and that mutant walls had similar FTIR spectratypes, which differed from that of wild-type plants. The data presented here suggest a canonical functionality of the Gβ and Gγ1/γ2 subunits in the control of Arabidopsis immune responses and the regulation of cell wall composition. PMID:21980142

  19. 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.

  20. Characterization and Biological Function of the ISOCHORISMATE SYNTHASE2 Gene of Arabidopsis1[OA

    PubMed Central

    Garcion, Christophe; Lohmann, Antje; Lamodière, Elisabeth; Catinot, Jérémy; Buchala, Antony; Doermann, Peter; Métraux, Jean-Pierre

    2008-01-01

    Salicylic acid (SA) is an important mediator of plant defense response. In Arabidopsis (Arabidopsis thaliana), this compound was proposed to derive mainly from isochorismate, itself produced from chorismate through the activity of ISOCHORISMATE SYNTHASE1 (ICS1). Null ics1 mutants still accumulate some SA, suggesting the existence of an enzymatic activity redundant with ICS1 or of an alternative ICS-independent SA biosynthetic route. Here, we studied the role of ICS2, a second ICS gene of the Arabidopsis genome, in the production of SA. We have shown that ICS2 encodes a functional ICS enzyme and that, similar to ICS1, ICS2 is targeted to the plastids. Comparison of SA accumulation in the ics1, ics2, and ics1 ics2 mutants indicates that ICS2 participates in the synthesis of SA, but in limited amounts that become clearly detectable only when ICS1 is lacking. This unequal redundancy relationship was also observed for phylloquinone, another isochorismate-derived end product. Furthermore, detection of SA in the double ics1 ics2 double mutant that is completely devoid of phylloquinone provides genetic evidence of the existence of an ICS-independent SA biosynthetic pathway in Arabidopsis. PMID:18451262

  1. The Arabidopsis thaliana proton transporters, AtNhx1 and Avp1, can function in cation detoxification in yeast

    PubMed Central

    Gaxiola, Roberto A.; Rao, Rajini; Sherman, Amir; Grisafi, Paula; Alper, Seth L.; Fink, Gerald R.

    1999-01-01

    Overexpression of the Arabidopsis thaliana vacuolar H+-pyrophosphatase (AVP1) confers salt tolerance to the salt-sensitive ena1 mutant of Saccharomyces cerevisiae. Suppression of salt sensitivity requires two ion transporters, the Gef1 Cl− channel and the Nhx1 Na+/H+ exchanger. These two proteins colocalize to the prevacuolar compartment of yeast and are thought to be required for optimal acidification of this compartment. Overexpression of AtNHX1, the plant homologue of the yeast Na+/H+ exchanger, suppresses some of the mutant phenotypes of the yeast nhx1 mutant. Moreover, the level of AtNHX1 mRNA in Arabidopsis is increased in the presence of NaCl. The regulation of AtNHX1 by NaCl and the ability of the plant gene to suppress the yeast nhx1 mutant suggest that the mechanism by which cations are detoxified in yeast and plants may be similar. PMID:9990049

  2. A Suite of Lotus japonicus Starch Mutants Reveals Both Conserved and Novel Features of Starch Metabolism1[W][OA

    PubMed Central

    Vriet, Cécile; Welham, Tracey; Brachmann, Andreas; Pike, Marilyn; Pike, Jodie; Perry, Jillian; Parniske, Martin; Sato, Shusei; Tabata, Satoshi; Smith, Alison M.; Wang, Trevor L.

    2010-01-01

    The metabolism of starch is of central importance for many aspects of plant growth and development. Information on leaf starch metabolism other than in Arabidopsis (Arabidopsis thaliana) is scarce. Furthermore, its importance in several agronomically important traits exemplified by legumes remains to be investigated. To address this issue, we have provided detailed information on the genes involved in starch metabolism in Lotus japonicus and have characterized a comprehensive collection of forward and TILLING (for Targeting Induced Local Lesions IN Genomes) reverse genetics mutants affecting five enzymes of starch synthesis and two enzymes of starch degradation. The mutants provide new insights into the structure-function relationships of ADP-glucose pyrophosphorylase and glucan, water dikinase1 in particular. Analyses of the mutant phenotypes indicate that the pathways of leaf starch metabolism in L. japonicus and Arabidopsis are largely conserved. However, the importance of these pathways for plant growth and development differs substantially between the two species. Whereas essentially starchless Arabidopsis plants lacking plastidial phosphoglucomutase grow slowly relative to wild-type plants, the equivalent mutant of L. japonicus grows normally even in a 12-h photoperiod. In contrast, the loss of GLUCAN, WATER DIKINASE1, required for starch degradation, has a far greater effect on plant growth and fertility in L. japonicus than in Arabidopsis. Moreover, we have also identified several mutants likely to be affected in new components or regulators of the pathways of starch metabolism. This suite of mutants provides a substantial new resource for further investigations of the partitioning of carbon and its importance for symbiotic nitrogen fixation, legume seed development, and perenniality and vegetative regrowth. PMID:20699404

  3. 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.

  4. 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

  5. Full genome re-sequencing reveals a novel circadian clock mutation in Arabidopsis

    PubMed Central

    2011-01-01

    Map based cloning in Arabidopsis thaliana can be a difficult and time-consuming process, specifically if the phenotype is subtle and scoring labour intensive. Here, we have re-sequenced the 120-Mb genome of a novel Arabidopsis clock mutant early bird (ebi-1) in Wassilewskija (Ws-2). We demonstrate the utility of sequencing a backcrossed line in limiting the number of SNPs considered. We identify a SNP in the gene AtNFXL-2 as the likely cause of the ebi-1 phenotype. PMID:21429190

  6. Altered etioplast development in phytochrome chromophore-deficient mutants.

    PubMed

    Terry, M J; Ryberg, M; Raitt, C E; Page, A M

    2001-12-01

    Inhibition of chromophore synthesis in the phytochrome-deficient aurea (au) and yellow-green-2 (yg-2) mutants of tomato (Solanum lycopersicum L.) results in a severe reduction of protochlorophyllide (Pchlide) accumulation in dark-grown hypocotyls. Experiments with apophytochrome-deficient mutants indicate that the inhibition of Pchlide accumulation results from two separate effects: one dependent on the activity of phytochromes A and B1 and one phytochrome-independent effect that is attributed to a feedback inhibition of the tetrapyrrole biosynthesis pathway. Cotyledons only show phytochrome-independent inhibition of Pchlide synthesis. Analysis of NADPH:protochlorophyllide oxidoreductase levels by western blotting showed that the reduction in Pchlide in au and yg-2 is accompanied by a correlative, but less substantial, decrease in NADPH:protochlorophyllide oxidoreductase. Consistent with this result, in vivo fluorescence spectra demonstrate that both mutants are primarily deficient in non-phototransformable Pchlide. Analysis of etioplast structure indicates that plastid development in au and yg-2 is retarded in hypocotyls and partially impaired in cotyledons, again correlating with the reduction in Pchlide. Since Pchlide synthesis is also reduced in chromophore-deficient mutants of pea (Pisum sativum L.) and Arabidopsis thaliana (L.) Heynh. (Landsberg erecta) these results may be significant for explaining aspects of the phenotype of this mutant class that are independent of the loss of phytochrome. PMID:11800397

  7. 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

  8. 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.

  9. Next-generation mapping of Arabidopsis genes.

    PubMed

    Austin, Ryan S; Vidaurre, Danielle; Stamatiou, George; Breit, Robert; Provart, Nicholas J; Bonetta, Dario; Zhang, Jianfeng; Fung, Pauline; Gong, Yunchen; Wang, Pauline W; McCourt, Peter; Guttman, David S

    2011-08-01

    Next-generation genomic sequencing technologies have made it possible to directly map mutations responsible for phenotypes of interest via direct sequencing. However, most mapping strategies proposed to date require some prior genetic analysis, which can be very time-consuming even in genetically tractable organisms. Here we present a de novo method for rapidly and robustly mapping the physical location of EMS mutations by sequencing a small pooled F₂ population. This method, called Next Generation Mapping (NGM), uses a chastity statistic to quantify the relative contribution of the parental mutant and mapping lines to each SNP in the pooled F₂ population. It then uses this information to objectively localize the candidate mutation based on its exclusive segregation with the mutant parental line. A user-friendly, web-based tool for performing NGM analysis is available at http://bar.utoronto.ca/NGM. We used NGM to identify three genes involved in cell-wall biology in Arabidopsis thaliana, and, in a power analysis, demonstrate success in test mappings using as few as ten F₂ lines and a single channel of Illumina Genome Analyzer data. This strategy can easily be applied to other model organisms, and we expect that it will also have utility in crops and any other eukaryote with a completed genome sequence. PMID:21518053

  10. An improved grafting technique for mature Arabidopsis plants demonstrates long-distance shoot-to-root transport of phytochelatins in Arabidopsis.

    PubMed

    Chen, Alice; Komives, Elizabeth A; Schroeder, Julian I

    2006-05-01

    Phytochelatins (PCs) are peptides that function in heavy-metal chelation and detoxification in plants and fungi. A recent study showed that PCs have the ability to undergo long-distance transport in a root-to-shoot direction in transgenic Arabidopsis (Arabidopsis thaliana). To determine whether long-distance transport of PCs can occur in the opposite direction, from shoots to roots, the wheat (Triticum aestivum) PC synthase (TaPCS1) gene was expressed under the control of a shoot-specific promoter (CAB2) in an Arabidopsis PC-deficient mutant, cad1-3 (CAB2TaPCS1/cad1-3). Analyses demonstrated that TaPCS1 is expressed only in shoots and that CAB2TaPCS1/cad1-3 lines complement the cadmium (Cd) and arsenic metal sensitivity of cad1-3 shoots. CAB2TaPCS1/cad1-3 plants exhibited higher Cd accumulation in roots and lower Cd accumulation in shoots compared to wild type. Fluorescence HPLC coupled to mass spectrometry analyses directly detected PC2 in the roots of CAB2:TaPCS1/cad1-3 but not in cad1-3 controls, suggesting that PC2 is transported over long distances in the shoot-to-root direction. In addition, wild-type shoot tissues were grafted onto PC synthase cad1-3 atpcs2-1 double loss-of-function mutant root tissues. An Arabidopsis grafting technique for mature plants was modified to obtain an 84% success rate, significantly greater than a previous rate of approximately 11%. Fluorescence HPLC-mass spectrometry showed the presence of PC2, PC3, and PC4 in the root tissue of grafts between wild-type shoots and cad1-3 atpcs2-1 double-mutant roots, demonstrating that PCs are transported over long distances from shoots to roots in Arabidopsis. PMID:16531489

  11. 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

  12. 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.

  13. Proteomics and transcriptomics analyses of Arabidopsis floral buds uncover important functions of ARABIDOPSIS SKP1-LIKE1

    DOE PAGESBeta

    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,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

  14. Regulation of Arabidopsis thaliana 5S rRNA Genes.

    PubMed

    Vaillant, Isabelle; Tutois, Sylvie; Cuvillier, Claudine; Schubert, Ingo; Tourmente, Sylvette

    2007-05-01

    The Arabidopsis thaliana genome comprises around 1,000 copies of 5S rRNA genes encoding both major and minor 5S rRNAs. In mature wild-type leaves, the minor 5S rRNA genes are silent. Using different mutants of DNA methyltransferases (met1, cmt3 and met1 cmt3), components of the RNAi pathway (ago4) or post-translational histone modifier (hda6/sil1), we show that the corresponding proteins are needed to maintain proper methylation patterns at heterochromatic 5S rDNA repeats. Using reverse transcription-PCR and cytological analyses, we report that a decrease of 5S rDNA methylation at CG or CNG sites in these mutants leads to the release of 5S rRNA gene silencing which occurred without detectable changes of the 5S rDNA chromatin structure. In spite of severely reduced DNA methylation, the met1 cmt3 double mutant revealed no increase in minor 5S rRNA transcripts. Furthermore, the release of silencing of minor 5S rDNAs can be achieved without increased formation of euchromatic loops by 5S rDNA, and is independent from the global heterochromatin content. Additionally, fluorescence in situ hybridization with centromeric 180 bp repeats confirmed that these highly repetitive sequences, in spite of their elevated transcriptional activity in the DNA methyltransferase mutants (met1, cmt3 and met1 cmt3), remain within chromocenters of the mutant nuclei. PMID:17412735

  15. Superroot, a recessive mutation in Arabidopsis, confers auxin overproduction.

    PubMed Central

    Boerjan, W; Cervera, M T; Delarue, M; Beeckman, T; Dewitte, W; Bellini, C; Caboche, M; Van Onckelen, H; Van Montagu, M; Inzé, D

    1995-01-01

    We have isolated seven allelic recessive Arabidopsis mutants, designated superroot (sur1-1 to sur1-7), displaying several abnormalities reminiscent of auxin effects. These characteristics include small and epinastic cotyledons, an elongated hypocotyl in which the connection between the stele and cortical and epidermal cells disintegrates, the development of excess adventitious and lateral roots, a reduced number of leaves, and the absence of an inflorescence. When germinated in the dark, sur1 mutants did not develop the apical hook characteristic of etiolated seedlings. We were able to phenocopy the Sur1- phenotype by supplying auxin to wild-type seedlings, to propagate sur1 explants on phytohormone-deficient medium, and to regenerate shoots from these explants by the addition of cytokinins alone to the culture medium. Analysis by gas chromatography coupled to mass spectrometry indicated increased levels of both free and conjugated indole-3-acetic acid. sur1 was crossed to the mutant axr2 and the altered-auxin response mutant ctr1. The phenotype of both double mutants was additive. The sur1 gene was mapped on chromosome 2 at 0.5 centimorgans from the gene encoding phytochrome B. PMID:8589625

  16. Disruption of the Arabidopsis Defense Regulator Genes SAG101, EDS1, and PAD4 Confers Enhanced Freezing Tolerance.

    PubMed

    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

    2015-10-01

    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 the 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 the 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 the wild-type leaves. These findings indicate that SAG101, EDS1, and PAD4 are involved in the freezing response in Arabidopsis, at least in part, by modulating the homeostasis of SA and DAG. PMID:26149542

  17. 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. PMID:21512130

  18. 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.

  19. Gibberellic Acid-Stimulated Arabidopsis6 Serves as an Integrator of Gibberellin, Abscisic Acid, and Glucose Signaling during Seed Germination in Arabidopsis1[OPEN

    PubMed Central

    Zhong, Chunmei; Xu, Hao; Ye, Siting; Wang, Shiyi; Li, Lingfei; Zhang, Shengchun; Wang, Xiaojing

    2015-01-01

    The DELLA protein REPRESSOR OF ga1-3-LIKE2 (RGL2) plays an important role in seed germination under different conditions through a number of transcription factors. However, the functions of the structural genes associated with RGL2-regulated germination are less defined. Here, we report the role of an Arabidopsis (Arabidopsis thaliana) cell wall-localized protein, Gibberellic Acid-Stimulated Arabidopsis6 (AtGASA6), in functionally linking RGL2 and a cell wall loosening expansin protein (Arabidopsis expansin A1 [AtEXPA1]), resulting in the control of embryonic axis elongation and seed germination. AtGASA6-overexpressing seeds showed precocious germination, whereas transfer DNA and RNA interference mutant seeds displayed delayed seed germination under abscisic acid, paclobutrazol, and glucose (Glc) stress conditions. The differences in germination rates resulted from corresponding variation in cell elongation in the hypocotyl-radicle transition region of the embryonic axis. AtGASA6 was down-regulated by RGL2, GLUCOSE INSENSITIVE2, and ABSCISIC ACID-INSENSITIVE5 genes, and loss of AtGASA6 expression in the gasa6 mutant reversed the insensitivity shown by the rgl2 mutant to paclobutrazol and the gin2 mutant to Glc-induced stress, suggesting that it is involved in regulating both the gibberellin and Glc signaling pathways. Furthermore, it was found that the promotion of seed germination and length of embryonic axis by AtGASA6 resulted from a promotion of cell elongation at the embryonic axis mediated by AtEXPA1. Taken together, the data indicate that AtGASA6 links RGL2 and AtEXPA1 functions and plays a role as an integrator of gibberellin, abscisic acid, and Glc signaling, resulting in the regulation of seed germination through a promotion of cell elongation. PMID:26400990

  20. An Arabidopsis peptide transporter is a member of a new class of membrane transport proteins.

    PubMed Central

    Steiner, H Y; Song, W; Zhang, L; Naider, F; Becker, J M; Stacey, G

    1994-01-01

    An Arabidopsis peptide transport gene was cloned from an Arabidopsis cDNA library by functionally complementing a yeast peptide transport mutant. The Arabidopsis plant peptide transporter (AtPTR2) allowed growth of yeast cells on dipeptides and tripeptides but not peptides four residues and higher. The plant peptide transporter also conferred sensitivity to a number of ethionine-containing, toxic peptides of chain length three or less and restored the ability to take up radiolabeled dileucine at levels similar to that of the wild type. Dileucine uptake was reduced by the addition of a variety of growth-promoting peptides. The sequence of a cDNA insert of 2.8 kb indicated an open reading frame encoding a 610-amino acid polypeptide (67.5 kD). Hydropathy analysis predicted a highly hydrophobic protein with a number of potential transmembrane segments. At the amino acid level, the Arabidopsis plant peptide transporter shows 24.6, 28.5, and 45.2% identity to the Arabidopsis nitrate-inducible nitrate transporter (CHL1), the rabbit small intestine oligopeptide transporter (PepT1), and the yeast peptide transporter (Ptr2p), respectively, but little identity to other proteins known to be involved in peptide transport. Root growth of Arabidopsis seedlings exposed to ethionine-containing toxic peptides was inhibited, and growth was restored by the addition of certain peptides shown to compete with dileucine uptake in yeast expressing the Arabidopsis transport gene. Consistent with the observed inhibition of root growth by toxic peptides, the peptide transporter is expressed in the roots of Arabidopsis seedlings. This study represents the characterization of a plant peptide transporter that is a member of a new class of related membrane transport proteins. PMID:7919993

  1. 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.

  2. Arabidopsis AtADF1 is functionally affected by mutations on actin binding sites.

    PubMed

    Dong, Chun-Hai; Tang, Wei-Ping; Liu, Jia-Yao

    2013-03-01

    The plant actin depolymerizing factor (ADF) binds to both monomeric and filamentous actin, and is directly involved in the depolymerization of actin filaments. To better understand the actin binding sites of the Arabidopsis thaliana L. AtADF1, we generated mutants of AtADF1 and investigated their functions in vitro and in vivo. Analysis of mutants harboring amino acid substitutions revealed that charged residues (Arg98 and Lys100) located at the α-helix 3 and forming an actin binding site together with the N-terminus are essential for both G- and F-actin binding. The basic residues on the β-strand 5 (K82/A) and the α-helix 4 (R135/A, R137/A) form another actin binding site that is important for F-actin binding. Using transient expression of CFP-tagged AtADF1 mutant proteins in onion (Allium cepa) peel epidermal cells and transgenic Arabidopsis thaliana L. plants overexpressing these mutants, we analyzed how these mutant proteins regulate actin organization and affect seedling growth. Our results show that the ADF mutants with a lower affinity for actin filament binding can still be functional, unless the affinity for actin monomers is also affected. The G-actin binding activity of the ADF plays an essential role in actin binding, depolymerization of actin polymers, and therefore in the control of actin organization. PMID:23190411

  3. LEAFY COTYLEDON1 Is an Essential Regulator of Late Embryogenesis and Cotyledon Identity in Arabidopsis.

    PubMed Central

    West, MAL.; Yee, K. M.; Danao, J.; Zimmerman, J. L.; Fischer, R. L.; Goldberg, R. B.; Harada, J. J.

    1994-01-01

    LEAFY COTYLEDON1 (LEC1) is an embryo defective mutation that affects cotyledon identity in Arabidopsis. Mutant cotyledons possess trichomes that are normally a leaf trait in Arabidopsis, and the cellular organization of these organs is intermediate between that of cotyledons and leaves from wild-type plants. We present several lines of evidence that indicate that the control of late embryogenesis is compromised by the mutation. First, mutant embryos are desiccation intolerant, yet embryos can be rescued before they dry to yield homozygous recessive plants that produce defective embryos exclusively. Second, although many genes normally expressed during embryonic development are active in the mutant, at least one maturation phase-specific gene is not activated. Third, the shoot apical meristem is activated precociously in mutant embryos. Fourth, in mutant embryos, several genes characteristic of postgerminative development are expressed at levels typical of wild-type seedlings rather than embryos. We conclude that postgerminative development is initiated prematurely and that embryonic and postgerminative programs operate simultaneously in mutant embryos. The pleiotropic effects of the mutation indicate that the LEC1 gene plays a fundamental role in regulating late embryogenesis. The role of LEC1 and its relationship to other genes involved in controlling late embryonic development are discussed. PMID:12244233

  4. The microRNA156 and microRNA172 gene regulation cascades at post-germinative stages in Arabidopsis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    MicroRNAs (miRNAs) are involved in developmental programs of plants including seed germination and post-germination. Here, we provide evidence that two different miRNA pathways, miR156 and miR172, interact during the post-germination stages in Arabidopsis. Mutant seedlings expressing miR156resistant...

  5. RABBIT EARS IS A SECOND-WHORL REPRESSOR OF AGAMOUS THAT MAINTAINS SPATIAL BOUNDARIES IN ARABIDOPSIS FLOWERS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The RABBIT EARS (RBE) gene has been identified as a regulator of petal development in Arabidopsis thaliana. We find that second-whorl petals in rbe mutants can be replaced with staminoid organs, stamens or filaments and that some rbe flowers have increased numbers of sepals and exhibit fusion of sep...

  6. THE ARABIDOPSIS GENE TARDY ASYNCHRONOUS MEIOSIS IS REQUIRED FOR THE NORMAL PACE AND SYNCHRONY OF CELL DIVISION DURING MALE MEIOSIS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Male meiosis in higher organisms features synchronous cell divisions in a large number of cells. It is not clear how this synchrony is achieved, nor is it known whether the synchrony is linked to the regulation of cell cycle progression. Here, we describe an Arabidopsis mutant, named tardy asynchron...

  7. PIRIN2 stabilizes cysteine protease XCP2 and increases susceptibility to the vascular pathogen Ralstonia solanacearum in Arabidopsis

    PubMed Central

    Zhang, Bo; Tremousaygue, Dominique; Denancé, Nicolas; van Esse, H Peter; Hörger, Anja C; Dabos, Patrick; Goffner, Deborah; Thomma, Bart P H J; van der Hoorn, Renier A L; Tuominen, Hannele

    2014-01-01

    PIRIN (PRN) is a member of the functionally diverse cupin protein superfamily. There are four members of the Arabidopsis thaliana PRN family, but the roles of these proteins are largely unknown. Here we describe a function of the Arabidopsis PIRIN2 (PRN2) that is related to susceptibility to the bacterial plant pathogen Ralstonia solanacearum. Two prn2 mutant alleles displayed decreased disease development and bacterial growth in response to R.  solanacearum infection. We elucidated the underlying molecular mechanism by analyzing PRN2 interactions with the papain-like cysteine proteases (PLCPs) XCP2, RD21A, and RD21B, all of which bound to PRN2 in yeast two-hybrid assays and in Arabidopsis protoplast co-immunoprecipitation assays. We show that XCP2 is stabilized by PRN2 through inhibition of its autolysis on the basis of PLCP activity profiling assays and enzymatic assays with recombinant protein. The stabilization of XCP2 by PRN2 was also confirmed in planta. Like prn2 mutants, an xcp2 single knockout mutant and xcp2 prn2 double knockout mutant displayed decreased susceptibility to R. solanacearum, suggesting that stabilization of XCP2 by PRN2 underlies susceptibility to R. solanacearum in Arabidopsis. PMID:24947605

  8. PIRIN2 stabilizes cysteine protease XCP2 and increases susceptibility to the vascular pathogen Ralstonia solanacearum in Arabidopsis.

    PubMed

    Zhang, Bo; Tremousaygue, Dominique; Denancé, Nicolas; van Esse, H Peter; Hörger, Anja C; Dabos, Patrick; Goffner, Deborah; Thomma, Bart P H J; van der Hoorn, Renier A L; Tuominen, Hannele

    2014-09-01

    PIRIN (PRN) is a member of the functionally diverse cupin protein superfamily. There are four members of the Arabidopsis thaliana PRN family, but the roles of these proteins are largely unknown. Here we describe a function of the Arabidopsis PIRIN2 (PRN2) that is related to susceptibility to the bacterial plant pathogen Ralstonia solanacearum. Two prn2 mutant alleles displayed decreased disease development and bacterial growth in response to R.  solanacearum infection. We elucidated the underlying molecular mechanism by analyzing PRN2 interactions with the papain-like cysteine proteases (PLCPs) XCP2, RD21A, and RD21B, all of which bound to PRN2 in yeast two-hybrid assays and in Arabidopsis protoplast co-immunoprecipitation assays. We show that XCP2 is stabilized by PRN2 through inhibition of its autolysis on the basis of PLCP activity profiling assays and enzymatic assays with recombinant protein. The stabilization of XCP2 by PRN2 was also confirmed in planta. Like prn2 mutants, an xcp2 single knockout mutant and xcp2 prn2 double knockout mutant displayed decreased susceptibility to R. solanacearum, suggesting that stabilization of XCP2 by PRN2 underlies susceptibility to R. solanacearum in Arabidopsis. PMID:24947605

  9. Genome-Wide Assessment of Efficiency and Specificity in CRISPR/Cas9 Mediated Multiple Site Targeting in Arabidopsis.

    PubMed

    Peterson, Brenda A; Haak, David C; Nishimura, Marc T; Teixeira, Paulo J P L; James, Sean R; Dangl, Jeffery L; Nimchuk, Zachary L

    2016-01-01

    Simultaneous multiplex mutation of large gene families using Cas9 has the potential to revolutionize agriculture and plant sciences. The targeting of multiple genomic sites at once raises concerns about the efficiency and specificity in targeting. The model Arabidopsis thaliana is widely used in basic plant research. Previous work has suggested that the Cas9 off-target rate in Arabidopsis is undetectable. Here we use deep sequencing on pooled plants simultaneously targeting 14 distinct genomic loci to demonstrate that multiplex targeting in Arabidopsis is highly specific to on-target sites with no detectable off-target events. In addition, chromosomal translocations are extremely rare. The high specificity of Cas9 in Arabidopsis makes this a reliable method for clean mutant generation with no need to enhance specificity or adopt alternate Cas9 variants. PMID:27622539

  10. Arabidopsis peroxisome proteomics

    PubMed Central

    Bussell, John D.; Behrens, Christof; Ecke, Wiebke; Eubel, Holger

    2013-01-01

    The analytical depth of investigation of the peroxisomal proteome of the model plant Arabidopsis thaliana has not yet reached that of other major cellular organelles such as chloroplasts or mitochondria. This is primarily due to the difficulties associated with isolating and obtaining purified samples of peroxisomes from Arabidopsis. So far only a handful of research groups have been successful in obtaining such fractions. To make things worse, enriched peroxisome fractions frequently suffer from significant organellar contamination, lowering confidence in localization assignment of the identified proteins. As with other cellular compartments, identification of peroxisomal proteins forms the basis for investigations of the dynamics of the peroxisomal proteome. It is therefore not surprising that, in terms of functional analyses by proteomic means, peroxisomes are lagging considerably behind chloroplasts or mitochondria. Alternative strategies are needed to overcome the obstacle of hard-to-obtain organellar fractions. This will help to close the knowledge gap between peroxisomes and other organelles and provide a full picture of the physiological pathways shared between organelles. In this review, we briefly summarize the status quo and discuss some of the methodological alternatives to classic organelle proteomic approaches. PMID:23630535

  11. Identification of an arsenic tolerant double mutant with a thiol-mediated component and increased arsenic tolerance in phyA mutants.

    PubMed

    Sung, Dong-Yul; Lee, David; Harris, Hugh; Raab, Andrea; Feldmann, Jörg; Meharg, Andrew; Kumabe, Bryan; Komives, Elizabeth A; Schroeder, Julian I

    2007-03-01

    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. PMID:17335514

  12. 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.

  13. Transgenic Arabidopsis Gene Expression System

    NASA Technical Reports Server (NTRS)

    Ferl, Robert; Paul, Anna-Lisa

    2009-01-01

    The Transgenic Arabidopsis Gene Expression System (TAGES) investigation is one in a pair of investigations that use the Advanced Biological Research System (ABRS) facility. TAGES uses Arabidopsis thaliana, thale cress, with sensor promoter-reporter gene constructs that render the plants as biomonitors (an organism used to determine the quality of the surrounding environment) of their environment using real-time nondestructive Green Fluorescent Protein (GFP) imagery and traditional postflight analyses.

  14. A glycolate dehydrogenase in the mitochondria of Arabidopsis thaliana.

    PubMed

    Bari, Rafijul; Kebeish, Rashad; Kalamajka, Rainer; Rademacher, Thomas; Peterhänsel, Christoph

    2004-03-01

    The fixation of molecular O2 by the oxygenase activity of Rubisco leads to the formation of phosphoglycolate in the chloroplast that is further metabolized in the process of photorespiration. The initial step of this pathway is the oxidation of glycolate to glyoxylate. Whereas in higher plants this reaction takes place in peroxisomes and is dependent on oxygen as a co-factor, most algae oxidize glycolate in the mitochondria using organic co-factors. The identification and characterization of a novel glycolate dehydrogenase in Arabidopsis thaliana is reported here. The enzyme is dependent on organic co-factors and resembles algal glycolate dehydrogenases in its enzymatic properties. Mutants of E. coli incapable of glycolate oxidation can be complemented by overexpression of the Arabidopsis open reading frame. The corresponding RNA accumulates preferentially in illuminated leaves, but was also found in other tissues investigated. A fusion of the N-terminal part of the Arabidopsis glycolate dehydrogenase to red fluorescent protein accumulates in mitochondria when overexpressed in the homologous system. Based on these results it is proposed that the basic photorespiratory system of algae is conserved in higher plants. PMID:14966218

  15. A hydroxycinnamoyltransferase responsible for synthesizing suberin aromatics in Arabidopsis

    SciTech Connect

    Gou, J.Y.; Liu, C.; Yu, X.-H.

    2009-11-03

    Suberin, a polyester polymer in the cell wall of terrestrial plants, controls the transport of water and nutrients and protects plant from pathogenic infections and environmental stresses. Structurally, suberin consists of aliphatic and aromatic domains; p-hydroxycinnamates, such as ferulate, p-coumarate, and/or sinapate, are the major phenolic constituents of the latter. By analyzing the 'wall-bound' phenolics of mutant lines of Arabidopsis deficient in a family of acyl-CoA dependent acyltransferase (BAHD) genes, we discovered that the formation of aromatic suberin in Arabidopsis, primarily in seed and root tissues, depends on a member of the BAHD superfamily of enzymes encoded by At5g41040. This enzyme exhibits an {omega}-hydroxyacid hydroxycinnamoyltransferase activity with an in vitro kinetic preference for feruloyl-CoA and 16-hydroxypalmitic acid. Knocking down or knocking out the At5g41040 gene in Arabidopsis reduces specifically the quantity of ferulate in suberin, but does not affect the accumulation of p-coumarate or sinapate. The loss of the suberin phenolic differentially affects the aliphatic monomer loads and alters the permeability and sensitivity of seeds and roots to salt stress. This highlights the importance of suberin aromatics in the polymer's function.

  16. The Significance of Hydrogen Sulfide for Arabidopsis Seed Germination

    PubMed Central

    Baudouin, Emmanuel; Poilevey, Aurélie; Hewage, Nishodi Indiketi; Cochet, Françoise; Puyaubert, Juliette; Bailly, Christophe

    2016-01-01

    Hydrogen sulfide (H2S) recently emerged as an important gaseous signaling molecule in plants. In this study, we investigated the possible functions of H2S in regulating Arabidopsis seed germination. NaHS treatments delayed seed germination in a dose-dependent manner and were ineffective in releasing seed dormancy. Interestingly, endogenous H2S content was enhanced in germinating seeds. This increase was correlated with higher activity of three enzymes (L-cysteine desulfhydrase, D-cysteine desulfhydrase, and β-cyanoalanine synthase) known as sources of H2S in plants. The H2S scavenger hypotaurine and the D/L cysteine desulfhydrase inhibitor propargylglycine significantly delayed seed germination. We analyzed the germinative capacity of des1 seeds mutated in Arabidopsis cytosolic L-cysteine desulfhydrase. Although the mutant seeds do not exhibit germination-evoked H2S formation, they retained similar germination capacity as the wild-type seeds. In addition, des1 seeds responded similarly to temperature and were as sensitive to ABA as wild type seeds. Taken together, these data suggest that, although its metabolism is stimulated upon seed imbibition, H2S plays, if any, a marginal role in regulating Arabidopsis seed germination under standard conditions. PMID:27446159

  17. Small Glycosylated Lignin Oligomers Are Stored in Arabidopsis Leaf Vacuoles

    PubMed Central

    Dima, Oana; Morreel, Kris; Vanholme, Bartel; Kim, Hoon; Ralph, John; Boerjan, Wout

    2015-01-01

    Lignin is an aromatic polymer derived from the combinatorial coupling of monolignol radicals in the cell wall. Recently, various glycosylated lignin oligomers have been revealed in Arabidopsis thaliana. Given that monolignol oxidation and monolignol radical coupling are known to occur in the apoplast, and glycosylation in the cytoplasm, it raises questions about the subcellular localization of glycosylated lignin oligomer biosynthesis and their storage. By metabolite profiling of Arabidopsis leaf vacuoles, we show that the leaf vacuole stores a large number of these small glycosylated lignin oligomers. Their structural variety and the incorporation of alternative monomers, as observed in Arabidopsis mutants with altered monolignol biosynthesis, indicate that they are all formed by combinatorial radical coupling. In contrast to the common believe that combinatorial coupling is restricted to the apoplast, we hypothesized that the aglycones of these compounds are made within the cell. To investigate this, leaf protoplast cultures were cofed with 13C6-labeled coniferyl alcohol and a 13C4-labeled dimer of coniferyl alcohol. Metabolite profiling of the cofed protoplasts provided strong support for the occurrence of intracellular monolignol coupling. We therefore propose a metabolic pathway involving intracellular combinatorial coupling of monolignol radicals, followed by oligomer glycosylation and vacuolar import, which shares characteristics with both lignin and lignan biosynthesis. PMID:25700483

  18. Connexin Mutants and Cataracts

    PubMed Central

    Beyer, Eric C.; Ebihara, Lisa; Berthoud, Viviana M.

    2013-01-01

    The lens is a multicellular, but avascular tissue that must stay transparent to allow normal transmission of light and focusing of it on the retina. Damage to lens cells and/or proteins can cause cataracts, opacities that disrupt these processes. The normal survival of the lens is facilitated by an extensive network of gap junctions formed predominantly of connexin46 and connexin50. Mutations of the genes that encode these connexins (GJA3 and GJA8) have been identified and linked to inheritance of cataracts in human families and mouse lines. In vitro expression studies of several of these mutants have shown that they exhibit abnormalities that may lead to disease. Many of the mutants reduce or modify intercellular communication due to channel alterations (including loss of function or altered gating) or due to impaired cellular trafficking which reduces the number of gap junction channels within the plasma membrane. However, the abnormalities detected in studies of other mutants suggest that they cause cataracts through other mechanisms including gain of hemichannel function (leading to cell injury and death) and formation of cytoplasmic accumulations (that may act as light scattering particles). These observations and the anticipated results of ongoing studies should elucidate the mechanisms of cataract development due to mutations of lens connexins and abnormalities of other lens proteins. They may also contribute to our understanding of the mechanisms of disease due to connexin mutations in other tissues. PMID:23596416

  19. Phosphoenolpyruvate Carboxylase in Arabidopsis Leaves Plays a Crucial Role in Carbon and Nitrogen Metabolism1

    PubMed Central

    Shi, Jianghua; Yi, Keke; Liu, Yu; Xie, Li; Zhou, Zhongjing; Chen, Yue; Hu, Zhanghua; Zheng, Tao; Liu, Renhu; Chen, Yunlong; Chen, Jinqing

    2015-01-01

    Phosphoenolpyruvate carboxylase (PEPC) is a crucial enzyme that catalyzes an irreversible primary metabolic reaction in plants. Previous studies have used transgenic plants expressing ectopic PEPC forms with diminished feedback inhibition to examine the role of PEPC in carbon and nitrogen metabolism. To date, the in vivo role of PEPC in carbon and nitrogen metabolism has not been analyzed in plants. In this study, we examined the role of PEPC in plants, demonstrating that PPC1 and PPC2 were highly expressed genes encoding PEPC in Arabidopsis (Arabidopsis thaliana) leaves and that PPC1 and PPC2 accounted for approximately 93% of total PEPC activity in the leaves. A double mutant, ppc1/ppc2, was constructed that exhibited a severe growth-arrest phenotype. The ppc1/ppc2 mutant accumulated more starch and sucrose than wild-type plants when seedlings were grown under normal conditions. Physiological and metabolic analysis revealed that decreased PEPC activity in the ppc1/ppc2 mutant greatly reduced the synthesis of malate and citrate and severely suppressed ammonium assimilation. Furthermore, nitrate levels in the ppc1/ppc2 mutant were significantly lower than those in wild-type plants due to the suppression of ammonium assimilation. Interestingly, starch and sucrose accumulation could be prevented and nitrate levels could be maintained by supplying the ppc1/ppc2 mutant with exogenous malate and glutamate, suggesting that low nitrogen status resulted in the alteration of carbon metabolism and prompted the accumulation of starch and sucrose in the ppc1/ppc2 mutant. Our results demonstrate that PEPC in leaves plays a crucial role in modulating the balance of carbon and nitrogen metabolism in Arabidopsis. PMID:25588735

  20. Arabidopsis ribosomal proteins control developmental programs through translational regulation of auxin response factors

    PubMed Central

    Rosado, Abel; Li, Ruixi; van de Ven, Wilhelmina; Hsu, Emily; Raikhel, Natasha V.

    2012-01-01

    Upstream ORFs are elements found in the 5′-leader sequences of specific mRNAs that modulate the translation of downstream ORFs encoding major gene products. In Arabidopsis, the translational control of auxin response factors (ARFs) by upstream ORFs has been proposed as a regulatory mechanism required to respond properly to complex auxin-signaling inputs. In this study, we identify and characterize the aberrant auxin responses in specific ribosomal protein mutants in which multiple ARF transcription factors are simultaneously repressed at the translational level. This characteristic lends itself to the use of these mutants as genetic tools to bypass the genetic redundancy among members of the ARF family in Arabidopsis. Using this approach, we were able to assign unique functions for ARF2, ARF3, and ARF6 in plant development. PMID:23144218

  1. An Arabidopsis flavonoid transporter is required for anther dehiscence and pollen development

    PubMed Central

    Thompson, Elinor P.; Wilkins, Christopher; Demidchik, Vadim; Davies, Julia M.; Glover, Beverley J.

    2010-01-01

    FLOWER FLAVONOID TRANSPORTER (FFT) encodes a multidrug and toxin efflux family transporter in Arabidopsis thaliana. FFT (AtDTX35) is highly transcribed in floral tissues, the transcript being localized to epidermal guard cells, including those of the anthers, stigma, siliques and nectaries. Mutant analysis demonstrates that the absence of FFT transcript affects flavonoid levels in the plant and that the altered flavonoid metabolism has wide-ranging consequences. Root growth, seed development and germination, and pollen development, release and viability are all affected. Spectrometry of mutant versus wild-type flowers shows altered levels of a glycosylated flavonol whereas anthocyanin seems unlikely to be the substrate as previously speculated. Thus, as well as adding FFT to the incompletely described flavonoid transport network, it is found that correct reproductive development in Arabidopsis is perturbed when this particular transporter is missing. PMID:19995827

  2. Enhanced Formaldehyde Detoxification by Overexpression of Glutathione-Dependent Formaldehyde Dehydrogenase from Arabidopsis1

    PubMed Central

    Achkor, Hakima; Díaz, Maykelis; Fernández, M. Rosario; Biosca, Josep Antoni; Parés, Xavier; Martínez, M. Carmen

    2003-01-01

    The ADH2 gene codes for the Arabidopsis glutathione-dependent formaldehyde dehydrogenase (FALDH), an enzyme involved in formaldehyde metabolism in eukaryotes. In the present work, we have investigated the potential role of FALDH in detoxification of exogenous formaldehyde. We have generated a yeast (Saccharomyces cerevisiae) mutant strain (sfa1Δ) by in vivo deletion of the SFA1 gene that codes for the endogenous FALDH. Overexpression of Arabidopsis FALDH in this mutant confers high resistance to formaldehyde added exogenously, which demonstrates the functional conservation of the enzyme through evolution and supports its essential role in formaldehyde metabolism. To investigate the role of the enzyme in plants, we have generated Arabidopsis transgenic lines with modified levels of FALDH. Plants overexpressing the enzyme show a 25% increase in their efficiency to take up exogenous formaldehyde, whereas plants with reduced levels of FALDH (due to either a cosuppression phenotype or to the expression of an antisense construct) show a marked slower rate and reduced ability for formaldehyde detoxification as compared with the wild-type Arabidopsis. These results show that the capacity to take up and detoxify high concentrations of formaldehyde is proportionally related to the FALDH activity in the plant, revealing the essential role of this enzyme in formaldehyde detoxification. PMID:12913179

  3. Biochemical characterization of Arabidopsis APYRASE family reveals their roles in regulating endomembrane NDP/NMP homoeostasis.

    PubMed

    Chiu, Tsan-Yu; Lao, Jeemeng; Manalansan, Bianca; Loqué, Dominique; Roux, Stanley J; Heazlewood, Joshua L

    2015-11-15

    Plant apyrases are nucleoside triphosphate (NTP) diphosphohydrolases (NTPDases) and have been implicated in an array of functions within the plant including the regulation of extracellular ATP. Arabidopsis encodes a family of seven membrane bound apyrases (AtAPY1-7) that comprise three distinct clades, all of which contain the five conserved apyrase domains. With the exception of AtAPY1 and AtAPY2, the biochemical and the sub-cellular characterization of the other members are currently unavailable. In this research, we have shown all seven Arabidopsis apyrases localize to internal membranes comprising the cis-Golgi, endoplasmic reticulum (ER) and endosome, indicating an endo-apyrase classification for the entire family. In addition, all members, with the exception of AtAPY7, can function as endo-apyrases by complementing a yeast double mutant (Δynd1Δgda1) which lacks apyrase activity. Interestingly, complementation of the mutant yeast using well characterized human apyrases could only be accomplished by using a functional ER endo-apyrase (NTPDase6), but not the ecto-apyrase (NTPDase1). Furthermore, the substrate specificity analysis for the Arabidopsis apyrases AtAPY1-6 indicated that each member has a distinct set of preferred substrates covering various NDPs (nucleoside diphosphates) and NTPs. Combining the biochemical analysis and sub-cellular localization of the Arabidopsis apyrases family, the data suggest their possible roles in regulating endomembrane NDP/NMP (nucleoside monophosphate) homoeostasis. PMID:26338998

  4. The COP9 signalosome interacts with SCF UFO and participates in Arabidopsis flower development.

    PubMed

    Wang, Xiping; Feng, Suhua; Nakayama, Naomi; Crosby, W L; Irish, Vivian; Deng, Xing Wang; Wei, Ning

    2003-05-01

    The COP9 signalosome (CSN) is involved in multiple developmental processes. It interacts with SCF ubiquitin ligases and deconjugates Nedd8/Rub1 from cullins (deneddylation). CSN is highly expressed in Arabidopsis floral tissues. To investigate the role of CSN in flower development, we examined the expression pattern of CSN in developing flowers. We report here that two csn1 partially deficient Arabidopsis strains exhibit aberrant development of floral organs, decline of APETALA3 (AP3) expression, and low fertility in addition to defects in shoot and inflorescence meristems. We show that UNUSUAL FLORAL ORGANS (UFO) forms a SCF(UFO) complex, which is associated with CSN in vivo. Genetic interaction analysis indicates that CSN is necessary for the gain-of-function activity of the F-box protein UFO in AP3 activation and in floral organ transformation. Compared with the previously reported csn5 antisense and csn1 null mutants, partial deficiency of CSN1 causes a reduction in the level of CUL1 in the mutant flowers without an obvious defect in CUL1 deneddylation. We conclude that CSN is an essential regulator of Arabidopsis flower development and suggest that CSN regulates Arabidopsis flower development in part by modulating SCF(UFO)-mediated AP3 activation. PMID:12724534

  5. Interactions of light and ethylene in hypocotyl hook maintenance in Arabidopsis thaliana seedlings

    NASA Technical Reports Server (NTRS)

    Knee, E. M.; Hangarter, R. P.; Knee, M.

    2000-01-01

    Etiolated seedlings frequently display a hypocotyl or epicotyl hook which opens on exposure to light. Etylene has been shown to be necessary for maintenance of the hook in a number of plants in darkness. We investigated the interaction of ethylene and light in the regulation of hypocotyl hook opening in Arabidopsis thaliana. We found that hooks of Arabidopsis open in response to continuous red, far-red or blue light in the presence of up to 100 microliters l-1 ethylene. Thus a change in sensitivity to ethylene is likely to be responsible for hook opening in Arabidopsis, rather than a decrease in ethylene production in hook tissues. We used photomorphogenic mutants of Arabidopsis to demonstrate the involvement of both blue light and phytochrome photosensory systems in light-induced hook opening in the presence of ethylene. In addition we used ethylene mutants and inhibitors of ethylene action to investigate the role of ethylene in hook maintenance in seedlings grown in light and darkness.

  6. Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic1

    PubMed Central

    Ventriglia, Tiziana; Kuhn, Misty L.; Ruiz, Ma Teresa; Ribeiro-Pedro, Marina; Valverde, Federico; Ballicora, Miguel A.; Preiss, Jack; Romero, José M.

    2008-01-01

    ADP-glucose (Glc) pyrophosphorylase (ADP-Glc PPase) catalyzes the first committed step in starch biosynthesis. Higher plant ADP-Glc PPase is a heterotetramer (α2β2) consisting of two small and two large subunits. There is increasing evidence that suggests that catalytic and regulatory properties of the enzyme from higher plants result from the synergy of both types of subunits. In Arabidopsis (Arabidopsis thaliana), two genes encode small subunits (APS1 and APS2) and four large subunits (APL1–APL4). Here, we show that in Arabidopsis, APL1 and APL2, besides their regulatory role, have catalytic activity. Heterotetramers formed by combinations of a noncatalytic APS1 and the four large subunits showed that APL1 and APL2 exhibited ADP-Glc PPase activity with distinctive sensitivities to the allosteric activator (3-phosphoglycerate). Mutation of the Glc-1-P binding site of Arabidopsis and potato (Solanum tuberosum) isoforms confirmed these observations. To determine the relevance of these activities in planta, a T-DNA mutant of APS1 (aps1) was characterized. aps1 is starchless, lacks ADP-Glc PPase activity, APS1 mRNA, and APS1 protein, and is late flowering in long days. Transgenic lines of the aps1 mutant, expressing an inactivated form of APS1, recovered the wild-type phenotype, indicating that APL1 and APL2 have catalytic activity and may contribute to ADP-Glc synthesis in planta. PMID:18614708

  7. Two Arabidopsis ADP-glucose pyrophosphorylase large subunits (APL1 and APL2) are catalytic.

    PubMed

    Ventriglia, Tiziana; Kuhn, Misty L; Ruiz, Ma Teresa; Ribeiro-Pedro, Marina; Valverde, Federico; Ballicora, Miguel A; Preiss, Jack; Romero, José M

    2008-09-01

    ADP-glucose (Glc) pyrophosphorylase (ADP-Glc PPase) catalyzes the first committed step in starch biosynthesis. Higher plant ADP-Glc PPase is a heterotetramer (alpha(2)beta(2)) consisting of two small and two large subunits. There is increasing evidence that suggests that catalytic and regulatory properties of the enzyme from higher plants result from the synergy of both types of subunits. In Arabidopsis (Arabidopsis thaliana), two genes encode small subunits (APS1 and APS2) and four large subunits (APL1-APL4). Here, we show that in Arabidopsis, APL1 and APL2, besides their regulatory role, have catalytic activity. Heterotetramers formed by combinations of a noncatalytic APS1 and the four large subunits showed that APL1 and APL2 exhibited ADP-Glc PPase activity with distinctive sensitivities to the allosteric activator (3-phosphoglycerate). Mutation of the Glc-1-P binding site of Arabidopsis and potato (Solanum tuberosum) isoforms confirmed these observations. To determine the relevance of these activities in planta, a T-DNA mutant of APS1 (aps1) was characterized. aps1 is starchless, lacks ADP-Glc PPase activity, APS1 mRNA, and APS1 protein, and is late flowering in long days. Transgenic lines of the aps1 mutant, expressing an inactivated form of APS1, recovered the wild-type phenotype, indicating that APL1 and APL2 have catalytic activity and may contribute to ADP-Glc synthesis in planta. PMID:18614708

  8. Cytokinins can act as suppressors of nitric oxide in Arabidopsis

    PubMed Central

    Liu, Wei-Zhong; Kong, Dong-Dong; Gu, Xue-Xin; Gao, Hong-Bo; Wang, Jin-Zheng; Xia, Min; Gao, Qian; Tian, Li-Li; Xu, Zhang-Hong; Bao, Fang; Hu, Yong; Ye, Neng-Sheng; Pei, Zhen-Ming; He, Yi-Kun

    2013-01-01

    Maintaining nitric oxide (NO) homeostasis is essential for normal plant physiological processes. However, very little is known about the mechanisms of NO modulation in plants. Here, we report a unique mechanism for the catabolism of NO based on the reaction with the plant hormone cytokinin. We screened for NO-insensitive mutants in Arabidopsis and isolated two allelic lines, cnu1-1 and 1–2 (continuous NO-unstressed 1), that were identified as the previously reported altered meristem program 1 (amp1) and as having elevated levels of cytokinins. A double mutant of cnu1-2 and nitric oxide overexpression 1 (nox1) reduced the severity of the phenotypes ascribed to excess NO levels as did treating the nox1 line with trans-zeatin, the predominant form of cytokinin in Arabidopsis. We further showed that peroxinitrite, an active NO derivative, can react with zeatin in vitro, which together with the results in vivo suggests that cytokinins suppress the action of NO most likely through direct interaction between them, leading to the reduction of endogenous NO levels. These results provide insights into NO signaling and regulation of its bioactivity in plants. PMID:23319631

  9. Indole Glucosinolate Biosynthesis Limits Phenylpropanoid Accumulation in Arabidopsis thaliana

    PubMed Central

    2015-01-01

    Plants produce an array of metabolites (including lignin monomers and soluble UV-protective metabolites) from phenylalanine through the phenylpropanoid biosynthetic pathway. A subset of plants, including many related to Arabidopsis thaliana, synthesizes glucosinolates, nitrogen- and sulfur-containing secondary metabolites that serve as components of a plant defense system that deters herbivores and pathogens. Here, we report that the Arabidopsis thaliana reduced epidermal fluorescence5 (ref5-1) mutant, identified in a screen for plants with defects in soluble phenylpropanoid accumulation, has a missense mutation in CYP83B1 and displays defects in glucosinolate biosynthesis and in phenylpropanoid accumulation. CYP79B2 and CYP79B3 are responsible for the production of the CYP83B1 substrate indole-3-acetaldoxime (IAOx), and we found that the phenylpropanoid content of cyp79b2 cyp79b3 and ref5-1 cyp79b2 cyp79b3 plants is increased compared with the wild type. These data suggest that levels of IAOx or a subsequent metabolite negatively influence phenylpropanoid accumulation in ref5 and more importantly that this crosstalk is relevant in the wild type. Additional biochemical and genetic evidence indicates that this inhibition impacts the early steps of the phenylpropanoid biosynthetic pathway and restoration of phenylpropanoid accumulation in a ref5-1 med5a/b triple mutant suggests that the function of the Mediator complex is required for the crosstalk. PMID:25944103

  10. Telomere Rapid Deletion Regulates Telomere Length in Arabidopsis thaliana▿

    PubMed Central

    Watson, J. Matthew; Shippen, Dorothy E.

    2007-01-01

    Telomere length is maintained in species-specific equilibrium primarily through a competition between telomerase-mediated elongation and the loss of terminal DNA through the end-replication problem. Recombinational activ