Showing papers in "Plant and Cell Physiology in 2007"
TL;DR: Results suggest that MdMYBA is a key regulatory gene in anthocyanin biosynthesis in apple skin.
Abstract: Red coloration of apple (Malus x domestica) skin is an important determinant of consumer preference and marketability. Anthocyanins are responsible for this coloration, and their accumulation is positively correlated with the expression level of anthocyanin biosynthetic genes. Regulation of expression of these genes is believed to be controlled by MYB transcription factors, and the MYB transcription factors involved in the activation of anthocyanin biosynthetic genes have been isolated in various plants. In the present study, we isolated and characterized a MYB transcription factor gene (MdMYBA) from apple skin. Characterization of MdMYBA demonstrated that (i) MdMYBA expression was specifically regulated depending on the tissue and cultivar/species; (ii) its expression level was much higher in a deep-red cultivar ('Jonathan') than in a pale-red cultivar ('Tsugaru'); (iii) when cauliflower mosaic virus 35S::MdMYBA was introduced into the cotyledons of apple seedlings by means of a transient assay, reddish-purple spots were induced, and MdMYBA also induced anthocyanin accumulation in reproductive tissues of transgenic tobacco; (iv) the expression of MdMYBA was induced by UV-B irradiation and low-temperature treatment, both of which are known to be important in the promotion of anthocyanin accumulation in apple skin; (v) MdMYBA bound specifically to an anthocyanidin synthase (MdANS) promoter region in a gel-shift assay; and (vi) MdMYBA was mapped to the near region of the BC226-STS (a1) marker for the red skin color locus (R(f)). These results suggest that MdMYBA is a key regulatory gene in anthocyanin biosynthesis in apple skin.
503 citations
TL;DR: It is demonstrated that HvAACT1 is an Al-activated citrate transporter responsible for Al resistance in barley.
Abstract: Soluble ionic aluminum (Al) inhibits root growth and reduces crop production on acid soils Al-resistant cultivars of barley (Hordeum vulgare L) detoxify Al by secreting citrate from the roots, but the responsible gene has not been identified yet Here, we identified a gene (HvAACT1) responsible for the Al-activated citrate secretion by fine mapping combined with microarray analysis, using an Al-resistant cultivar, Murasakimochi, and an Al-sensitive cultivar, Morex This gene belongs to the multidrug and toxic compound extrusion (MATE) family and was constitutively expressed mainly in the roots of the Al-resistant barley cultivar Heterologous expression of HvAACT1 in Xenopus oocytes showed efflux activity for (14)C-labeled citrate, but not for malate Two-electrode voltage clamp analysis also showed transport activity of citrate in the HvAACT1-expressing oocytes in the presence of Al Overexpression of this gene in tobacco enhanced citrate secretion and Al resistance compared with the wild-type plants Transiently expressed green fluorescent protein-tagged HvAACT1 was localized at the plasma membrane of the onion epidermal cells, and immunostaining showed that HvAACT1 was localized in the epidermal cells of the barley root tips A good correlation was found between the expression of HvAACT1 and citrate secretion in 10 barley cultivars differing in Al resistance Taken together, our results demonstrate that HvAACT1 is an Al-activated citrate transporter responsible for Al resistance in barley
455 citations
TL;DR: This work down-regulated the endogenous dihydroflavonol 4-reductase (DFR) gene and overexpressed the Irisxhollandica DFR gene in addition to the viola F3'5'H gene in a rose cultivar, resulting in the accumulation of a high percentage of delphinidin and a novel bluish flower color.
Abstract: Flower color is mainly determined by anthocyanins. Rosa hybrida lacks violet to blue flower varieties due to the absence of delphinidin-based anthocyanins, usually the major constituents of violet and blue flowers, because roses do not possess flavonoid 3',5'-hydoxylase (F3'5'H), a key enzyme for delphinidin biosynthesis. Other factors such as the presence of co-pigments and the vacuolar pH also affect flower color. We analyzed the flavonoid composition of hundreds of rose cultivars and measured the pH of their petal juice in order to select hosts of genetic transformation that would be suitable for the exclusive accumulation of delphinidin and the resulting color change toward blue. Expression of the viola F3'5'H gene in some of the selected cultivars resulted in the accumulation of a high percentage of delphinidin (up to 95%) and a novel bluish flower color. For more exclusive and dominant accumulation of delphinidin irrespective of the hosts, we down-regulated the endogenous dihydroflavonol 4-reductase (DFR) gene and overexpressed the Irisxhollandica DFR gene in addition to the viola F3'5'H gene in a rose cultivar. The resultant roses exclusively accumulated delphinidin in the petals, and the flowers had blue hues not achieved by hybridization breeding. Moreover, the ability for exclusive accumulation of delphinidin was inherited by the next generations.
408 citations
TL;DR: Co-expression network analysis is a powerful approach for data-driven hypothesis construction and gene prioritization, and provides novel insights into the system-level understanding of plant cellular processes.
Abstract: Gene co-expression, in many cases, implies the presence of a functional linkage between genes. Co-expression analysis has uncovered gene regulatory mechanisms in model organisms such as Escherichia coli and yeast. Recently, accumulation of Arabidopsis microarray data has facilitated a genome-wide inspection of gene co-expression profiles in this model plant. An approach using network analysis has provided an intuitive way to represent complex co-expression patterns between many genes. Co-expression network analysis has enabled us to extract modules, or groups of tightly co-expressed genes, associated with biological processes. Furthermore, integrated analysis of gene expression and metabolite accumulation has allowed us to hypothesize the functions of genes associated with specific metabolic processes. Co-expression network analysis is a powerful approach for data-driven hypothesis construction and gene prioritization, and provides novel insights into the system-level understanding of plant cellular processes.
345 citations
TL;DR: It is demonstrated that the phytohormone ABA is a potential signal for cold-induced pollen sterility (CIPS) and the cold tolerance phenotype of R31 is correlated with lower endogenous ABA levels and a different regulation of ABA metabolism.
Abstract: Cold temperatures cause pollen sterility and large reductions in grain yield in temperate rice growing regions of the world. Induction of pollen sterility by cold involves a disruption of sugar transport in anthers, caused by the cold-induced repression of the apoplastic sugar transport pathway in the tapetum. Here we demonstrate that the phytohormone ABA is a potential signal for cold-induced pollen sterility (CIPS). Cold treatment of the cold-sensitive cultivar Doongara resulted in increased anther ABA levels. Exogenous ABA treatment at the young microspore stage induced pollen sterility and affected cell wall invertase and monosaccharide transporter gene expression in a way similar to cold treatment. In the cold-tolerant cultivar R31, ABA levels were significantly lower under normal circumstances and remained low after cold treatment. The differences in endogenous ABA levels in Doongara and R31 correlated with differences in expression of the ABA biosynthetic genes encoding zeaxanthin epoxidase (OSZEP1) and 9-cis-epoxycarotenoid dioxygenase (OSNCED2, OSNCED3) in anthers. The expression of three ABA-8-hydroxylase genes (ABA8OX1, 2 and 3) in R31 anthers was higher under control conditions and was regulated differently by cold compared with Doongara. Our results indicate that the cold tolerance phenotype of R31 is correlated with lower endogenous ABA levels and a different regulation of ABA metabolism.
255 citations
TL;DR: It is demonstrated that the PTGS suppressor (2b) of a severe strain of cucumber mosaic virus (CMV) can bind to in vitro synthesized siRNAs and even to long dsRNAs to a lesser extent, however, the 2b suppressor weakly bound to a miRNA (miR171) duplex in contrast to another small RNA-binding suppressor, p19 of tombusvirus that can effectively bind miRNAs.
Abstract: Double-stranded (ds) RNAs and imperfect hairpin RNAs of endogenous genes trigger post-transcriptional gene silencing (PTGS) and are cleaved by a Dicer-like nuclease into small interfering RNAs (siRNAs) and microRNs (miRNAs), respectively. Such small RNAs (siRNAs and miRNAs) then guide an RNA-induced silencing complex (RISC) for sequence-specific RNA degradation. While PTGS serves as an antiviral defense in plants, many plant viruses encode suppressors as a counter defense. Here we demonstrate that the PTGS suppressor (2b) of a severe strain (CM95R) of cucumber mosaic virus (CMV) can bind to in vitro synthesized siRNAs and even to long dsRNAs to a lesser extent. However, the 2b suppressor weakly bound to a miRNA (miR171) duplex in contrast to another small RNA-binding suppressor, p19 of tombusvirus that can effectively bind miRNAs. Because the 2b suppressor of an attenuated strain of CMV (CM95), which differs in a single amino acid from the 2b of CM95R, could barely bind siRNAs, we hypothesized that the weak suppressor activity of the attenuated strain resulted from a loss of the siRNA-binding property of 2b via a single amino acid change. Here we consider that 2b interferes with the PTGS pathway by directly binding siRNAs (or long dsRNA).
223 citations
TL;DR: Analyses of TBL1 expression in hormone signaling mutants with aberrant branching suggest that T BL1 acts downstream of auxin and the MAX-related hormone to coordinate bud outgrowth, which is consistent with Arabidopsis TBL 1 providing functionality similar to monocot TB1.
Abstract: Axillary bud outgrowth is controlled by developmental and environmental signals through the integrated action of hormones and other factors that probably regulate the cell cycle in the buds. While hormonal regulators have been studied extensively, less is known about downstream mechanisms regulating bud outgrowth. The TCP domain protein TEOSINTE BRANCHED1 (TB1) is a putative transcriptional regulator that represses bud outgrowth in grasses. Phylogenetic analyses have indicated that three hypothetical Arabidopsis (Arabidopsis thaliana) proteins, TCP1, TCP12 and TCP18 [TEOSINTE BRANCHED1-LIKE 1 (TBL1)], are closely related to monocot TB1 proteins. A reverse genetics approach was used to identify TBL1 and TCP12 mutants to assess the function of the hypothetical proteins. No obvious phenotype was observed in tcp12 mutants. tbl1 null mutants exhibited a non-pleiotropic hyperbranching phenotype that was due to enhanced outgrowth of primary and secondary buds. The role of TBL1 as a repressor of bud outgrowth was supported by TBL1 mRNA accumulation: abundance was high in unelongated buds, and decreased to low levels in buds that were elongating. Analyses of TBL1 expression in hormone signaling mutants with aberrant branching suggest that TBL1 acts downstream of auxin and the MAX-related hormone to coordinate bud outgrowth. The data are consistent with Arabidopsis TBL1 providing functionality similar to monocot TB1, and highlight the conservation of mechanisms regulating branching across large evolutionary distances.
214 citations
TL;DR: It is demonstrated that overexpression of miR165 recapitulates the phenotypes caused by loss-of-function mutations of HD-ZIP III genes, such as loss of SAM, altered organ polarity and defects in development of vascular tissues and interfascicular fibers.
Abstract: The class III homeodomain leucine-zipper (HD-ZIP III) genes are thought to be targets of microRNAs (miRNAs) 165 and 166, but it is not known whether all the developmental processes affected by mutations of the HD-ZIP III genes could be recapitulated by an alteration in the expression of miR165 and miR166. Previous work showed that overexpression of miR166 by activation tagging results in down-regulation of the ATHB-9/PHV, ATHB-14/PHB and ATHB-15 genes, and concomitantly causes an enlargement of shoot apical meristems (SAMs) and an enhancement in vascular development. Here we demonstrated that overexpression of miR165 causes a drastic reduction in the transcript levels of all five HD-ZIP III genes in Arabidopsis. The miR165 overexpressors display prominent phenotypes reminiscent of loss-of-function mutants of rev phb phv and rev/ifl1, including loss of SAM, alteration of organ polarity, abnormal formation of carpels, inhibition of vascular development and aberrant differentiation of interfascicular fibers. Global gene expression analysis revealed a link between miR165 overexpression and altered expression of genes involved in auxin signaling and vascular development. Our results demonstrate that overexpression of miR165 recapitulates the phenotypes caused by loss-of-function mutations of HD-ZIP III genes, such as loss of SAM, altered organ polarity and defects in development of vascular tissues and interfascicular fibers.
211 citations
TL;DR: Overexpression of the CK-inducible type-A response regulator OsRR6 abolished shoot regeneration, suggesting that Os RR6 acts as a negative regulator of CK signaling, and increased content of trans-zeatin-type CKs inOsRR6-ox lines indicates that homeostatic control of CK levels is regulated by OsRR 6 signaling.
Abstract: Genome-wide analyses of rice (Oryza sativa L.) cytokinin (CK)-responsive genes using the Affymetrix GeneChip(R) rice genome array were conducted to define the spectrum of genes subject to regulation by CK in monocotyledonous plants. Application of trans-zeatin modulated the expression of a wide variety of genes including those involved in hormone signaling and metabolism, transcriptional regulation, macronutrient transport and protein synthesis. To understand further the function of CK in rice plants, we examined the effects of in planta manipulation of a putative CK signaling factor on morphology, CK metabolism and expression of CK-responsive genes. Overexpression of the CK-inducible type-A response regulator OsRR6 abolished shoot regeneration, suggesting that OsRR6 acts as a negative regulator of CK signaling. Transgenic lines overexpressing OsRR6 (OsRR6-ox) had dwarf phenotypes with poorly developed root systems and panicles. Increased content of trans-zeatin-type CKs in OsRR6-ox lines indicates that homeostatic control of CK levels is regulated by OsRR6 signaling. Expression of genes encoding CK oxidase/dehydrogenase decreased in OsRR6-ox plants, possibly accounting for elevated CK levels in transgenic lines. Expression of a number of stress response genes was also altered in OsRR6-ox plants.
202 citations
TL;DR: Results indicate that AOX can dissipate the excess reducing equivalents, which are transported from the chloroplasts, and serve in efficient photosynthesis.
Abstract: Alternative oxidase (AOX), the unique terminal oxidase in plant mitochondria, catalyzes the energy-wasteful cyanide (CN)-resistant respiration. Although it has been suggested that AOX might prevent chloroplast over-reduction through the efficient dissipation of excess reducing equivalents, direct evidence for this in the physiological context has been lacking. In this study, we examined the mitochondrial respiratory properties, especially AOX, connected to the accumulation of reducing equivalents in the chloroplasts and the activities of enzymes needed to transport the reducing equivalents. We used Arabidopsis thaliana mutants defective in cyclic electron flow around PSI, in which the reducing equivalents accumulate in the chloroplast stroma due to an unbalanced ATP/NADPH production ratio. These mutants showed higher activities of the enzymes needed to transport the reducing equivalents even in low-light growth conditions. The amounts of AOX protein and CN-resistant respiration in the mutants were also higher than those in the wild type. After high-light treatment, AOX, even in the wild type, was preferentially up-regulated concomitant with the accumulation of reducing equivalents in the chloroplasts and an increase in the activities of enzymes needed to transport reducing equivalents. These results indicate that AOX can dissipate the excess reducing equivalents, which are transported from the chloroplasts, and serve in efficient photosynthesis.
199 citations
TL;DR: This study reports physiological and molecular responses of wild watermelon, which exhibits extraordinarily high drought resistance, and suggests that this xerophyte switches survival strategies from drought avoidance to drought tolerance during the progression of drought stress, by regulating its root proteome in a temporally programmed manner.
Abstract: Water availability is a critical determinant for the growth and ecological distribution of terrestrial plants. Although some xerophytes are unique regarding their highly developed root architecture and the successful adaptation to arid environments, virtually nothing is known about the molecular mechanisms underlying this adaptation. Here, we report physiological and molecular responses of wild watermelon (Citrullus lanatus sp.), which exhibits extraordinarily high drought resistance. At the early stage of drought stress, root development of wild watermelon was significantly enhanced compared with that of the irrigated plants, indicating the activation of a drought avoidance mechanism for absorbing water from deep soil layers. Consistent with this observation, comparative proteome analysis revealed that many proteins induced in the early stage of drought stress are involved in root morphogenesis and carbon/nitrogen metabolism, which may contribute to the drought avoidance via the enhancement of root growth. On the other hand, lignin synthesis-related proteins and molecular chaperones, which may function in the enhancement of physical desiccation tolerance and maintenance of protein integrity, respectively, were induced mostly at the later stage of drought stress. Our findings suggest that this xerophyte switches survival strategies from drought avoidance to drought tolerance during the progression of drought stress, by regulating its root proteome in a temporally programmed manner. This study provides new insights into the complex molecular networks within plant roots involved in the adaptation to adverse environments.
TL;DR: This work showed that the clock-associated genes PSEUDO-RESPONSE REGULATOR (PRR) PRR9, PRR7 and PRR5 are involved in activation of CO expression during the daytime, and suggested that PRR genes act coordinately in a manner parallel with and antagonistic to CCA/LHY, upstream of the canonical CO-FLOWERING LOCUS T (FT) photoperiodic flowering pathway.
Abstract: Photoperiodism allows organisms to measure daylength, or external photoperiod, and to anticipate coming seasons. Daylength measurement requires the integration of light signal and temporal information by the circadian clock. In the long-day plant Arabidopsis thaliana, CONSTANS (CO) plays a crucial role in integrating the circadian rhythm and environmental light signals into the photoperiodic flowering pathway. Nevertheless, the molecular mechanism by which the circadian clock modulates the cyclic expression profile of CO is poorly understood. Here, we first showed that the clock-associated genes PSEUDO-RESPONSE REGULATOR (PRR) PRR9, PRR7 and PRR5 are involved in activation of CO expression during the daytime. Then, extensive genetic studies using CIRCADIAN CLOCK-ASSOCIATED1 (CCA1)/LATE ELONGATED HYPOCOTYL (LHY) double mutants (cca1/lhy) and prr7/prr5 were conducted. The results suggested that PRR genes act coordinately in a manner parallel with and antagonistic to CCA/LHY, upstream of the canonical CO-FLOWERING LOCUS T (FT) photoperiodic flowering pathway. Finally, we provided evidence to propose a model, in which CCA1/LHY repress CO through GIGANTEA (GI), while PRR9, PRR7 and PRR5 activate CO predominantly by repressing CYCLING DOF FACTOR1 (CDF1) encoding a DNA-binding transcriptional repressor.
TL;DR: Analysis of monosaccharide composition in conjunction with digestion with pectolytic enzymes conclusively demonstrated that the principal pectic domain of both layers was rhamnogalacturonan I, and that in the outer layer this was unbranched.
Abstract: A comprehensive analysis was carried out of the composition of seed coat mucilage from Arabidopsis thaliana using the Columbia-0 accession. Pectinaceous mucilage is released from myxospermous seeds upon imbibition, and in Arabidopsis consists of a water-soluble, outer layer and an adherent, inner layer. Analysis of monosaccharide composition in conjunction with digestion with pectolytic enzymes conclusively demonstrated that the principal pectic domain of both layers was rhamnogalacturonan I, and that in the outer layer this was unbranched. The macromolecular characteristics of the water-soluble mucilage indicated that the rhamnogalacturonan molecules in the outer layer were in a slightly expanded random-coil conformation. The inner, adherent layer remained attached to the seed, even after extraction with acid and alkali, suggesting that its integrity was maintained by covalent bonds. Confocal microscopy and monosaccharide composition analyses showed that the inner layer can be separated into two domains. The internal domain contained cellulose microfibrils, which could form a matrix with RGI and bind it to the seed. In effect, in the mum5-1 mutant where most of the inner and outer mucilage layers were water soluble, cellulose remained attached to the seed coat. Immunolabeling with anti-pectin antibodies indicated the presence of galactan and arabinan in the inner layer, with the latter only present in the non-cellulose-containing external domain. In addition, JIM5 and JIM7 antibodies labeled different domains of the inner layer, suggesting the presence of stretches of homogalacturonan with different levels of methyl esterification.
TL;DR: It is proposed that XET activity is essential for G-fiber shrinking by repairing xyloglucan cross-links between G- and S(2)-layers and thus maintaining their contact.
Abstract: Tension wood is a specialized tissue of deciduous trees that functions in bending woody stems to optimize their position in space. Tension wood fibers that develop on one side of the stem have an increased potency to shrink compared with fibers on the opposite side, thus creating a bending moment. It is believed that the gelatinous (G) cell wall layer containing almost pure cellulose of tension wood fibers is pivotal to their shrinking. By analyzing saccharide composition and linkage in isolated G-layers of poplar, we found that they contain some matrix components in addition to cellulose, of which xyloglucan is the most abundant. Xyloglucan, xyloglucan endo-transglycosylase (XET) activity and xyloglucan endo-transglycosylase/hydrolase (XTH) gene products were detected in developing G-layers by labeling using CCRC-M1 monoclonal antibody, in situ incorporation of XXXG-SR and the polyclonal antibody to poplar PttXET16-34, respectively, indicating that xyloglucan is incorporated into the G-layer during its development. Moreover, several XTH transcripts were altered and were generally up-regulated in developing tension wood compared with normal wood. In mature G-fibers, XTH gene products were detected in the G-layers while the XET activity was evident in the adjacent S 2 wall layer. We propose that XET activity is essential for G-fiber shrinking by repairing xyloglucan cross-links between G- and S 2 -layers and thus maintaining their contact. Surprisingly, XTH gene products and XET activity persisted in mature G-fibers for several years, suggesting that the enzyme functions after cell death repairing the cross-links as they are being broken during the shrinking process.
TL;DR: It is demonstrated that PARVUS, which is a member of family GT8, is required for the biosynthesis of the tetrasaccharide primer sequence, located at the reducing end of glucuronoxylan (GX) biosynthesis.
Abstract: Xylan, cellulose and lignin are the three major components of secondary walls in wood, and elucidation of the biosynthetic pathway of xylan is of importance for potential modification of secondary wall composition to produce wood with improved properties. So far, three Arabidopsis glycosyltransferases, FRAGILE FIBER8, IRREGULAR XYLEM8 and IRREGULAR XYLEM9, have been implicated in glucuronoxylan (GX) biosynthesis. In this study, we demonstrate that PARVUS, which is a member of family GT8, is required for the biosynthesis of the tetrasaccharide primer sequence, beta-D-Xyl-(1 --> 3)-alpha-l-Rha-(1 --> 2)-alpha-D-GalA-(1 --> 4)-D-Xyl, located at the reducing end of GX. The PARVUS gene is expressed during secondary wall biosynthesis in fibers and vessels, and its encoded protein is predominantly localized in the endoplasmic reticulum. Mutation of the PARVUS gene leads to a drastic reduction in secondary wall thickening and GX content. Structural analysis of GX using (1)H-nuclear magnetic resonance (NMR) spectroscopy revealed that the parvus mutation causes a loss of the tetrasaccharide primer sequence at the reducing end of GX and an absence of glucuronic acid side chains in GX. Activity assay showed that the xylan xylosyltransferase and glucuronyltransferase activities were not affected in the parvus mutant. Together, these findings implicate a possible role for PARVUS in the initiation of biosynthesis of the GX tetrasaccharide primer sequence and provide novel insights into the mechanisms of GX biosynthesis.
TL;DR: It is demonstrated that AtSZF1 and AtSzF2 negatively regulate the expression of salt-responsive genes and play important roles in modulating the tolerance of Arabidopsis plants to salt stress.
Abstract: The molecular mechanism governing the response of plants to salinity stress, one of the most significant limiting factors for agriculture worldwide, has just started to be revealed. Here, we report AtSZF1 and AtSZF2, two closely related CCCH-type zinc finger proteins, involved in salt stress responses in Arabidopsis. The expression of AtSZF1 and AtSZF2 is quickly and transiently induced by NaCl treatment. Mutants disrupted in the expression of AtSZF1 or AtSZF2 exhibit increased expression of a group of salt stress-responsive genes in response to high salt. Significantly, the atszf1-1/atszf2-1 double mutant displays more sensitive responses to salt stress than the atszf1-1 or atszf2-1 single mutants and wild-type plants. On the other hand, transgenic plants overexpressing AtSZF1 show reduced induction of salt stress-responsive genes and are more tolerant to salt stress. We also showed that AtSZF1 is localized in the nucleus. Taken together, these results demonstrated that AtSZF1 and AtSZF2 negatively regulate the expression of salt-responsive genes and play important roles in modulating the tolerance of Arabidopsis plants to salt stress.
TL;DR: Analysis of promoter::GUS (beta-glucuronidase) transgenic plants representing six different GASA loci reveals that the promoters are activated in a variety of stage- and tissue-specific patterns during development, indicating that the GASS genes are involved in diverse processes.
Abstract: Members of the plant-specific gibberellic acid-stimulated Arabidopsis (GASA) gene family play roles in hormone response, defense and development. We have identified six new Arabidopsis GASA genes, bringing the total number of family members to 14. Here we show that these genes all encode small polypeptides that share the common structural features of an N-terminal putative signal sequence, a highly divergent intermediate region and a conserved 60 amino acid C-terminal domain containing 12 conserved cysteine residues. Analysis of promoter::GUS (β-glucuronidase) transgenic plants representing six different GASA loci reveals that the promoters are activated in a variety of stage- and tissue-specific patterns during development, indicating that the GASA genes are involved in diverse processes. Characterization of GASA4 shows that the promoter is active in the shoot apex region, developing flowers and developing embryos. Phenotypic analyses of GASA4 loss-of-function and gain-of-function lines indicate that GASA4 regulates floral meristem identity and also positively affects both seed size and total seed yield.
TL;DR: It is shown that microsomes isolated from the stems of wild-type Arabidopsis exhibit XylT and GlcAT activities in the presence of exogenous 1,4-linked beta-d-xylooligomers.
Abstract: Xylan is the second most abundant polysaccharide in dicot wood, and thus elucidation of the xylan biosynthetic pathway is required to understand the mechanisms controlling wood formation. Genetic and chemical studies in Arabidopsis have implicated three genes, FRAGILE FIBER8 (FRA8), IRREGULAR XYLEM8 (IRX8) and IRREGULAR XYLEM9 (IRX9), in the biosynthesis of glucuronoxylan (GX), but the biochemical functions of the encoded proteins are not known. In this study, we determined the effect of the fra8, irx8 and irx9 mutations on the activities of xylan xylosyltransferase (XylT) and glucuronyltransferase (GlcAT). We show that microsomes isolated from the stems of wild-type Arabidopsis exhibit XylT and GlcAT activities in the presence of exogenous 1,4-linked β-D-xylooligomers. Xylooligomers ranging in size from two to six can be used as acceptors by XylT to form xylooligosaccharides with up to 12 xylosyl residues. We provide evidence that the irx9 mutation results in a substantial reduction in XylT activity but has no discernible effect on GlcAT activity. In contrast, neither XylT nor GlcAT activity is affected by fra8 and irx8 mutations. Our results provide biochemical evidence that the irx9 mutation results in a deficiency in xylan XylT activity, thus leading to a defect in the elongation of the xylan backbone.
TL;DR: Examination of local and systemic gene expression revealed that GP17-2 and its CF modulate the expression of genes involved in both the SA and JA/ET signaling pathways, hypothesizing the involvement of multiple defense mechanisms leading to an ISR of Arabidopsis by GP 17-2.
Abstract: Arabidopsis thaliana grown in soil amended with barley grain inocula of Penicillium simplicissimum GP17-2 or receiving root treatment with its culture filtrate (CF) exhibited clear resistance to Pseudomonas syringae pv. tomato DC3000 (Pst). To assess the contribution of different defense pathways, Arabidopsis genotypes implicated in salicylic acid (SA) signaling expressing the NahG transgene or carrying disruption in NPR1 (npr1), jasmonic acid (JA) signaling (jar1) and ethylene (ET) signaling (ein2) were tested. All genotypes screened were protected by GP17-2 or its CF. However, the level of protection was significantly lower in NahG and npr1 plants than it was in similarly treated wild-type plants, indicating that the SA signaling pathway makes a minor contribution to the GP17-2-mediated resistance and is insufficient for a full response. Examination of local and systemic gene expression revealed that GP17-2 and its CF modulate the expression of genes involved in both the SA and JA/ET signaling pathways. Subsequent challenge of GP17-2-colonized plants with Pst was accompanied by direct activation of SA-inducible PR-2 and PR-5 genes as well as potentiated expression of the JA-inducible Vsp gene. In contrast, CF-treated plants infected with Pst exhibited elevated expression of most defense-related genes (PR-1, PR-2, PR-5, PDF1.2 and Hel) studied. Moreover, an initial elevation of SA responses was followed by late induction of JA responses during Pst infection of induced systemic resistance (ISR)-expressing plants. In conclusion, we hypothesize the involvement of multiple defense mechanisms leading to an ISR of Arabidopsis by GP17-2.
TL;DR: Genes from roots of B-tolerant cultivars of wheat and barley with high similarities to previously reported B efflux transporters from Arabidopsis and rice were cloned and show tolerance in proportion to the level of expression of the transporter gene.
Abstract: Tolerance to boron (B) toxicity in cereals is known to be associated with reduced tissue accumulation of B. Genes from roots of B-tolerant cultivars of wheat and barley with high similarities to previously reported B efflux transporters from Arabidopsis and rice were cloned. Expression of these genes was strongly correlated with the ability of tolerant genotypes to lower the concentration of B in roots. The gene from barley located to chromosome 4. Backcross lines containing a B tolerance locus on chromosome 4 showed tolerance in proportion to the level of expression of the transporter gene, whereas those lacking the locus were sensitive to B and had very low levels of gene expression. The results are consistent with a widespread mechanism of tolerance to high B based on efflux of B from root cells.
TL;DR: A regulator of transcription in berberine biosynthesis is identified using functional genomics with a transient RNA interference (RNAi) and overexpression of the candidate gene, which is named CjWRKY1.
Abstract: Selected cultured Coptis japonica cells produce a large amount of the benzylisoquinoline alkaloid berberine. Previous studies have suggested that berberine productivity is controlled at the transcript level of biosynthetic genes. We have identified a regulator of transcription in berberine biosynthesis using functional genomics with a transient RNA interference (RNAi) and overexpression of the candidate gene. The 24 primary candidate clones were selected from 1,014 expressed sequence tags (ESTs) that were obtained from a C. japonica cell line producing high levels of berberine. Further characterization of the expression profiles of these ESTs suggested that five ESTs would be good candidates as regulators of berberine production. A newly developed transient RNAi system with C. japonica protoplasts indicated that double-stranded RNA of an EST clone significantly reduced the level of transcripts of 3'-hydroxy N-methylcoclaurine 4'-O-methyltransferase. Sequence analysis showed that this EST encoded a group-II WRKY, and we named it CjWRKY1. When the effects of double-stranded RNA of the CjWRKY1 gene were examined in detail, a marked reduction in the transcripts of all genes involved in berberine biosynthesis was detected, whereas little effect was found in the transcript levels of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and chorismate mutase (CM) that are associated with primary metabolism. Ectopic expression of CjWRKY1 cDNA in C. japonica protoplasts clearly increased the level of transcripts of all berberine biosynthetic genes examined compared with control treatment, whereas the levels of GAPDH and CM were not affected. The functional role of CjWRKY1 as a specific and comprehensive regulator of berberine biosynthesis is discussed.
TL;DR: Experimental evidence is provided that this model monocot Oryza sativa also has a set of highly conserved clock-associated genes, such as those designated as OsCCA1, OsPRR-series including OsTOC1/OsPRR 1, OsZTLs, OsPCL1 as well as OsGI, which will provide insight into the general roles of plant circadian clocks.
Abstract: In higher plants, circadian rhythms are highly relevant to a wide range of biological processes. To such circadian rhythms, the clock (oscillator) is central, and recent intensive studies on the model higher plant Arabidopsis thaliana have begun to shed light on the molecular mechanisms underlying the functions of the central clock. Such representative clock-associated genes of A. thaliana are the homologous CCA1 and LHY genes, and five PRR genes that belong to a small family of pseudo-response regulators including TOC1. Others are GI, ZTL, ELF3, ELF4, LUX/PCL1, etc. In this context, a simple question arose as to whether or not the molecular picture of the model Arabidopsis clock is conserved in other higher plants. Here we made an effort to answer the question with special reference to Oryza sativa, providing experimental evidence that this model monocot also has a set of highly conserved clock-associated genes, such as those designated as OsCCA1, OsPRR-series including OsTOC1/OsPRR1, OsZTLs, OsPCL1 as well as OsGI. These results will provide us with insight into the general roles of plant circadian clocks, such as those for the photoperiodic control of flowering time that has a strong impact on the reproduction and yield in many higher plants.
TL;DR: It is shown that atwbc11 mutants displayed organ fusions and stunted growth, and became vulnerable to chlorophyll leaching and toluidine blue staining, and results support that AtWBC11 is involved in cuticle development.
Abstract: Cuticle, including wax and cutin, is the barrier covering plant aerial organs and protecting the inner tissues. The Arabidopsis thaliana ATP-binding cassette (ABC) transporter CER5 (AtWBC12) has been identified as a wax exporter. In agreement with the latest report of another wax exporter, AtWBC11, here we show that atwbc11 mutants displayed organ fusions and stunted growth, and became vulnerable to chlorophyll leaching and toluidine blue staining. Chemical analysis showed that wax and cutin monomers were both reduced in the atwbc11 mutant. AtWBC11 was widely expressed in aerial organs. Interestingly, we found that the expression was light dependent, and the phytohormone ABA up-regulated AtWBC11 expression. We also found that while the AtWBC11 promoter had a broad pattern of activity, the expression was converted to epidermis specific when the reporter gene was fused to AtWBC11 cDNA. Furthermore, RNA blot analysis supported epidermis-specific expression of AtWBC11. Our results support that AtWBC11 is involved in cuticle development.
TL;DR: Investigation of the CLE peptide function in Arabidopsis and rice revealed 47 putative CLE genes in the rice genome and multiple CLE domains in some CLE genes, indicating diverse CLE function in these plants.
Abstract: Using 26 chemically synthetic CLAVATA3/ESR (CLE) peptides, which correspond to the predicted products of the 31 Arabidopsis CLE genes, we investigated the CLE peptide function in Arabidopsis and rice. Treatment with some CLE peptides inhibited root elongation in rice as well as in Arabidopsis. It also reduced the size of the shoot apical meristem in Arabidopsis but not in rice. Database searches revealed 47 putative CLE genes in the rice genome and multiple CLE domains in some CLE genes, indicating diverse CLE function in these plants.
TL;DR: It is shown that Arabidopsis WRKY62, a member of WRKY group III transcription factors, was induced by methyl jasmonate and SA treatment and acts downstream of cytosolic NPR1 and negatively regulates JA-responsive gene expression, suggesting thatWRKY62 may be involved in the SA-mediated suppression of JA signaling.
Abstract: Cytosolic NPR1 has been shown to be essential for the salicylic acid (SA)-mediated suppression of jasmonic acid (JA)-responsive gene expression. However, factors downstream of NPR1 in the cross-talk between SA and JA signaling are unclear. Here we show that Arabidopsis WRKY62, a member of WRKY group III transcription factors, was induced by methyl jasmonate (MeJA) and SA treatment. The presence of basal SA is required for the MeJA-induced WRKY62 expression, and both chemicals exhibit a synergistic effect on WRKY62 induction. In addition, upon treatment with an extremely low concentration of SA, cytosolic NPR1 controls the MeJA-induced expression of WRKY62. TGA transcription factors, which up-regulate SA-induced expression of WRKY62, are dispensable for the induction of WRKY62 in JA signaling. Genetic dissection of both wrky62 mutants and WRKY62-overexpressing plants indicated that WRKY62 down-regulates JA-responsive LOX2 and VSP2 expression. Our results demonstrate that WRKY62 acts downstream of cytosolic NPR1 and negatively regulates JA-responsive gene expression, suggesting that WRKY62 may be involved in the SA-mediated suppression of JA signaling.
TL;DR: G-6-PDH plays a pivotal role in NR-dependent NO production, and in establishing tolerance of red kidney bean roots to salt stress, in investigation of tolerance to oxidative stress induced by 100 mM NaCl in red kidneybean roots.
Abstract: The pivotal role of glucose-6-phosphate dehydrogenase (G-6-PDH)-mediated nitric oxide (NO) production in the tolerance to oxidative stress induced by 100 mM NaCl in red kidney bean (Phaseolus vulgaris) roots was investigated. The results show that the G-6-PDH activity was enhanced rapidly in the presence of NaCl and reached a maximum at 100 mM. Western blot analysis indicated that the increase of G-6-PDH activity in the red kidney bean roots under 100 mM NaCl was mainly due to the increased content of the G-6-PDH protein. NO production and nitrate reductase (NR) activity were also induced by 100 mM NaCl. The NO production was reduced by NaN(3) (an NR inhibitor), but not affected by N(omega)-nitro-L-arginine (L-NNA) (an NOS inhibitor). Application of 2.5 mM Na(3)PO(4), an inhibitor of G-6-PDH, blocked the increase of G-6-PDH and NR activity, as well as NO production in red kidney bean roots under 100 mM NaCl. The activities of antioxidant enzymes in red kidney bean roots increased in the presence of 100 mM NaCl or sodium nitroprusside (SNP), an NO donor. The increased activities of all antioxidant enzymes tested at 100 mM NaCl were completely inhibited by 2.5 mM Na(3)PO(4). Based on these results, we conclude that G-6-PDH plays a pivotal role in NR-dependent NO production, and in establishing tolerance of red kidney bean roots to salt stress.
TL;DR: Investigating the intracellular localization and intermolecular interaction of these molecules using imaging techniques, including bimolecular fluorescence complementation and fluorescence resonance energy transfer techniques, found that DCL1, HYL1 and SE formed bodies which localized in the nuclei.
Abstract: There has been much recent research on the contribution of microRNA (miRNA) in plant organogenesis and hormone action. In plants, it has been reported that Dicer-like 1 (DCL1), HYPONASTIC LEAVES1 (HYL1) and SERRATE (SE) are involved in the production of miRNAs. The means by which miRNAs are processed and transported is not well understood in detail, however. In this study, we investigated the intracellular localization and intermolecular interaction of these molecules using imaging techniques, including bimolecular fluorescence complementation and fluorescence resonance energy transfer techniques, making use of various enhanced fluorescent proteins. We found that DCL1, HYL1 and SE formed bodies which localized in the nuclei. We were also able to locate the miRNA primary transcript using an MS2-tagged method on these bodies. It appears very likely that the observed DCL1-HYL1-SE nuclear body is involved in miRNA production. Co-expression of SmD3 or SmB proteins revealed the localization of DCL1-HYL1-SE complexes in the SmD3/SmB nuclear bodies.
TL;DR: Novel evidence is provided indicating that the His-Asp phosphorelay is connected to diverse regulatory levels of cytokinin-responsive phenomena through ARR1 direct-target genes.
Abstract: Plant cells respond to cytokinins by changing their gene expression patterns. The histidyl–aspartyl (His–Asp) phosphorelay mediates the signal from cytokinin receptors to type-B response regulators including ARR1, which transactivate cytokinin primary response genes. However, the overall architecture of the signal cascade leading to cytokininresponsive phenomena is still unclear, mainly because it is not known how the His–Asp phosphorelay is connected to downstream phenomena. To reveal events immediately downstream from the phosphorelay-mediated transcriptional activation, we searched for direct-target genes of ARR1 by exploiting ARR1DDK–GR, a chimeric transcription factor that transactivates ARR1 direct-target genes in transgenic plants by glucocorticoid induction. We identified 23 directtarget genes, most of which were found to be cytokinin primary response genes. The arr1-1 mutation clearly affected the primary response in at least 17 genes, meaning that they respond primarily to cytokinins through the function of ARR1. The 17 genes encode proteins with diverse functions, including type-A response regulators, cytokinin metabolic enzymes and putative disease resistance response proteins. These results provide novel evidence indicating that the His–Asp phosphorelay is connected to diverse regulatory levels of cytokinin-responsive phenomena through ARR1 direct-target genes.
TL;DR: It is shown that salt stress-induced wild-type Arabidopsis seedling roots display right-handed skewed growth and depolymerization of the cortical microtubules and seedling survival may be mediated by calcium influx in salt stress.
Abstract: Although the results of some studies indicate that salt stress affects the organization of microtubules, it remains an open question whether microtubules play an active role in the plant's ability to withstand salt stress. In the present study, we showed that salt stress-induced wild-type Arabidopsis seedling roots display right-handed skewed growth and depolymerization of the cortical microtubules. The results of a long-term observational study showed that cortical microtubules depolymerized then reorganized themselves under salt stress. Stabilization of microtubules with paclitaxel resulted in more seedling death under salt stress, while disruption of microtubules with oryzalin or propyzamide rescued seedlings from death. Seedlings in which the cortical microtubules were reorganized did not succumb to salt stress. These results suggest that both depolymerization and reorganization of the cortical microtubules are important for the plant's ability to withstand salt stress. Depolymerizing microtubules by drugs rescues seedlings from death under salt stress. This rescue effect was abolished by removing calcium from the medium or treatment with a calcium channel inhibitor. Depolymerization of the microtubules is followed by an increase in the free cytoplasmic calcium concentration. The addition of calcium to the growth medium increased the number of seedlings in which recovery of the cortical microtubules occurred, whereas the removal of calcium decreased the number of seedlings in which recovery occurred. Therefore, depolymerization of the cortical microtubules raises intracellular calcium concentrations, while reorganization of the cortical microtubules and seedling survival may be mediated by calcium influx in salt stress.
TL;DR: It is concluded that LAZY1 is specifically involved in shoot gravitropism and that LAzY1-dependent and -independent signaling pathways occur in coleoptiles, and it is further concluded that only the LAZy1- dependent gravity signaling involves asymmetric distribution of auxin between the two lateral halves and is required for circumnutation.
Abstract: We identified the gene responsible for three allelic lazy1 mutations of Japonica rice (Oryza sativa L.) by map-based cloning, complementation and RNA interference. Sequence analysis and database searches indicated that the wild-type gene (LAZY1) encodes a novel and unique protein (LAZY1) and that rice has no homologous gene. Two lazy1 mutants were LAZY1 null. Confirming and advancing the previously reported results on lazy1 mutants, we found the following. (i) Gravitropism is impaired, but only partially, in lazy1 coleoptiles. (ii) Circumnutation, observed in dark-grown coleoptiles, is totally absent from lazy1 coleoptiles. (iii) Primary roots of lazy1 mutants show normal gravitropism and circumnutation. (iv) LAZY1 is expressed in a tissue-specific manner in gravity-sensitive shoot tissues (i.e. coleoptiles, leaf sheath pulvini and lamina joints) and is little expressed in roots. (v) The gravitropic response of lazy1 coleoptiles is kinetically separable from that absent from lazy1 coleoptiles. (vi) Gravity-induced lateral translocation of auxin, found in wild-type coleoptiles, does not occur in lazy1 coleoptiles. Based on the genetic and physiological evidence obtained, it is concluded that LAZY1 is specifically involved in shoot gravitropism and that LAZY1-dependent and -independent signaling pathways occur in coleoptiles. It is further concluded that, in coleoptiles, only the LAZY1-dependent gravity signaling involves asymmetric distribution of auxin between the two lateral halves and is required for circumnutation.