Showing papers in "Plant and Cell Physiology in 1997"
TL;DR: It is suggested that levels of Pro are regulated at the level of transcriptional the genes of these two enzymes during dehydration and rehydration, which appear to be the rate-limiting factors under water stress.
Abstract: Compatible osmolytes are potent osmoprotectants that play a role in counteracting the effects of osmotic stress. Proline (Pro) is one of the most common compatible osmolytes in water-stressed plants. The accumulation of Pro in dehydrated plants is caused both by activation of the biosynthesis of Pro and by inactivation of the degradation of Pro. In plants, L-Pro is synthesized from L-glutamic acid (L-G1U) via zl'-pyrroline-S-carboxylate (P5C) by two enzymes, P5C synthetase (P5CS) and P5C reductase (P5CR). L-Pro is metabolized to L-G1U via P5C by two enzymes, proline dehydrogenase (oxidase) (ProDH; EC 1.5.99.8) and P5C dehydrogenase (P5CDH; EC 1.5.1.12). Such metabolism of Pro is inhibited when Pro accumulates during dehydration and it is activated when rehydration occurs. Under dehydration conditions, when expression of the gene for P5CS is strongly induced, expression of the gene for ProDH is inhibited. By contrast, under rehydration conditions, when the expression of the gene for ProDH is strongly induced, the expression of the gene for P5CS is inhibited. Thus, P5CS, which acts during the biosynthesis of Pro, and ProDH, which acts during the metabolism of Pro, appear to be the rate-limiting factors under water stress. Therefore, it is suggested that levels of Pro are regulated at the level of transcriptional the genes of these two enzymes during dehydration and rehydration. Moreover, it has been demonstrated that Pro acts as an osmoprotectant and that overproduction of Pro results in increased tolerance to osmotic stress of transgenic tobacco plants. Genetically engineered crop plants that overproduce Pro might, thus, acquire osmotolerance, namely, the ability to tolerate environmental stresses such as drought and high salinity.
653 citations
TL;DR: Generations of superoxide anions and hydrogen peroxide in the vascular tissue were particularly apparent in the xylem and associated with the sites of distribution of CuZn-SOD as determined by an immunohistochemical method, and also with the location of lignin as determined with the phloroglucin-HCl reaction.
Abstract: The sites of generations of superoxide anions and hydrogen peroxide in cross sections of hypocotyls from spinach seedlings were located by staining with nitroblue tetrazolium (NBT) and with starch-iodide, respectively. Formazan, produced upon the reduction of NBT by superoxide, was observed mainly in the vascular tissue only in the presence of inhibitors of CuZn-superoxide dismutase (CuZn-SOD), and its formation was suppressed under anaerobic conditions. Thus, NBT was reduced to formazan specifically by the superoxide anions generated in vascular tissue. The reduction of NBT was suppressed by inhibitors of NAD(P)H oxidase, but neither by cyanide nor azide, indicating the involvement of NAD(P)H oxidase in the generation of superoxide anions in the vascular tissue. Starch-I2 complex also was formed in the vascular tissue, but not in the presence of either the CuZn-SOD inhibitor or the NAD(P)H oxidase inhibitor, indicating that the hydrogen peroxide is produced via the catalytic disproportionation with CuZn-SOD of the superoxide generated by NAD(P)H oxidase. Generations of superoxide anions and hydrogen peroxide in the vascular tissue were particularly apparent in the xylem and associated with the sites of distribution of CuZn-SOD as determined by an immunohistochemical method, and also with the location of lignin as determined by the phloroglucin-HCl reaction.
235 citations
TL;DR: Findings indicate that the secretion of citric acid is a response specific to Al stress in C. tora and constitutes a mechanism of Al resistance.
Abstract: A rapid and sensitive assay method for Al-chelating activity was established to screen Al-chelating substances secreted from roots of Al-resistant species in response to Al stress. From one Al-resistant species, Cassia tora L., an Alchelating substance was detected in the root exudates when they were exposed to SO fM Al in 0.5 mM CaCl2 solution at pH 4.5; the dominant component was identified as citric acid. The secretion of citric acid was very low during the first 4 h after initiation of Al treatment, but increased markedly thereafter. A 3-h pulse with 50 //M Al also induced significant secretion of citric acid after 6 h. The lag between Al addition and secretion of citric acid suggests that inducible processes are involved. A dose-response experiment showed that the amount of secreted citric acid increased with increasing external concentrations of Al. Eight-d treatment of P deficiency did not induce the secretion of citric acid. Exposure to 50 //M of either lanthanum (La3*) or ytterbium (Yb3+) did not induce the secretion of citric acid either. These findings indicate that the secretion of citric acid is a response specific to Al stress in C. tora and constitutes a mechanism of Al resistance.
214 citations
TL;DR: To understand the entire genetic system in cyanobacteria, the nucleotide sequence of the entire genome of the unicellular cyanobacterium Synechocystis sp.
Abstract: Cyanobacteria are photoautotrophic organisms capable of oxygen-producing photosynthesis similar to that in eukaryotic algae and plants, and because of this, they have been used as model organisms for the study of the mechanism and regulation of oxygen-producing photosynthesis. To understand the entire genetic system in cyanobacteria, the nucleotide sequence of the entire genome of the unicellular cyanobacterium Synechocystis sp. PCC6803 has been determined. The total length of the circular genome is 3,573,470 bp, with a GC content of 47.7%. A total of 3,168 potential protein coding genes were assigned. Of these, 145 (4.6%) were identical to reported genes, and 1,259 (39.6%) and 342 (10.8%) showed similarity to reported and hypothetical genes, respectively. The remaining 1,422 (45.0%) showed no apparent similarity to any genes registered in the databases. Classification of the genes by their biological function and comparison of the gene complement with those of other organisms have revealed a variety of features of the genetic information characteristic of a photoautotrophic organism. The sequence data, as well as other information on the Synechocystis genome, is presented in CyanoBase on WWW [http:/(/)www.kazusa.or.jp/cyano/].
187 citations
TL;DR: A new oxygenic photosynthetic prokaryote was isolated that contained chlorophyll (/ as a predominant pigment withchlorophyll a being a minor pigment and the existence of a new light utilization mechanism involving chlorophylla d was indicated.
Abstract: The principal pigment found in the majority of oxygenic photosynthetic organisms is known to be chlorophyll a. However, we isolated a new oxygenic photosynthetic prokaryote that contained chlorophyll (/ as a predominant pigment with chlorophyll a being a minor pigment. Chlorophyll d had previously been noted but its natural occurrence and function remained unclear. Cells of the new prokaryote had an absorption maximum at red region of 714-718 nm due to chlorophyll d absorption, but no characteristic absorption peak of chlorophyll a around 680 nm was observed. Chlorophyll d of the new organism was identified spectrophotometrically in several solvents and its chemical structure was confirmed by NMR and FABMS analysis. The cell also contained a chlorophyll c-like pigment, zeaxanthin and a-carotene but not chlorophyll b and /^-carotene. The content of chlorophyll d accounted for more than 2% of the cell dry weight, while the content of chlorophyll a was less than 0.1%. The chlorophyll a/d ratio remained between 0.03 and 0.09 under different culture conditions. The light absorption characteristics and the high content of chlorophyll d along with the small content of chlorophyll a indicated the existence of a new light utilization mechanism involving chlorophyll d.
182 citations
TL;DR: Results strongly suggest that endogenous brassinosteroids induce entry into the final stage of differentiation in Zinnia cells.
Abstract: To elucidate the involvement of brassinosteroids in the progression of tracheary element differentiation in cultured Zinnia cells, we analyzed the effects of uniconazole, an inhibitor of brassinosteroid synthesis, and brassinolide, a biologically active brassinosteroid, on the accumulation of raRNAs for various genes that were expressed in different stages of differentiation. Uniconazole specifically suppressed the accumulation of transcripts for genes that were induced in the final stage of differentiation in association with secondary wall formation and cell death. This suppression was recovered with the addition of brassinolide. These results strongly suggest that endogenous brassinosteroids induce entry into the final stage of differentiation.
181 citations
TL;DR: Two new PAM fluorometers (pulse amplitude modulated) were used in an investigation of photosynthetic performance of Prochloron resident as a symbiont in the ascidian Lissoclinum patella, growing in a coral reef of Heron Island on the Great Barrier Reef, and the pattern of fluorescence responses differed markedly from those found in cyanobacteria.
Abstract: Two new PAM fluorometers (pulse amplitude modulated) were used in an investigation of photosynthetic performance of Prochloron resident as a symbiont in the ascidian Lissoclinum patella, growing in a coral reef of Heron Island on the Great Barrier Reef. With a new DIVINGPAM in situ measurements of effective PSII quantum yield (/1F/Fm') as a function of quantum flux density (rapid light curves) were carried out in 2.5 m depth in the reef and in a seawater tank. Photosynthetic electron transport rates were measured on in hospite Prochloron both in situ and in collected material. Both light-limited and light-saturated yields were exceptionally high. Maximal yields (Fv/Fm) were ~0.83. A new TEACHING-PAM was employed for analysing dark-light induction and light-dark relaxation kinetics in collected samples with Prochloron in hospite. Considerable variability in kinetic responses was observed which was found to be at least in part due to differences in O2 concentration. It is suggested that endogenous reductants feed electrons into the intersystem transport chain, which normally is reoxidized by O2 (chlororespiration), and that in the dark, the reduction level of PSII acceptors is increased due to a decline in O2 concentration. The pattern of fluorescence responses differed markedly from those found in cyanobacteria and provides new insights into light-harvesting responses of a photosynthetic prokaryote with a membrane bound light-harvesting system, as contrasted with an extrinsic light-harvesting system.
174 citations
TL;DR: N-terminal amino acid sequence analysis of the four S-proteins revealed that they were highly homologous to each other and similar to the S-RNases of Malus, Pyrus, Scrophulariaceae, and Solanaceae.
Abstract: Stylar proteins of 13 almond (Prunus dulcis) cultivars with known 5-genotypes were surveyed by IEF and IDPAGE combined with immunoblot and N-terminal amino acid sequence analyses to identify S-RNases associated with gametophytic self-incompatibility (SI) in this plant species. RNase activities corresponding to Sa and Sb, two of the four S-alleles tested, were identified by IEF and RNase activity staining. The Sa-RNase band reacted with the antiiS>serum prepared from Japanese pear (Pyrus serotina); no reaction with the antiserum was observed with the ^-RNase band. When the 5,,-RNase band was excised from an IEF gel stained for RNase activity, subjected to SDSPAGE, and detected by immunoblotting, it appeared that this band consisted of a single protein that reacted with the anti-5rserum with M, of about 28 kDa. With 2D-PAGE and silver staining of the stylar extracts, all four S-proteins could be successfully distinguished from each other in the highly basic zone of the gel. Although Sb-, Sc-, and Sd-proteins had roughly the same Mr of about 30 kDa, the ^-protein seemed to be slightly smaller than the ^-protein and slightly larger than the 5 serum. N-terminal amino acid sequence analysis of the four 5-proteins revealed that they were highly homologous to each other and similar to the 5-RNases of Malus, Pyrus, Scrophulariaceae, and Solanaceae. Taken together, RNases in the style are strongly suggested to be associated with the gametophytic SI of alAbbreviations: CIG, cross incompatibility group; 2D-PAGE, two dimensional polyacrylarnide gel electrophoresis; IEF, isoelectric focusing; NEPHGE, nonequilibrium pH gradient electrophoresis; Mr, relative molecular weight; SC, self-compatibility; SI, self-incompatibility. Correspondence to Ryutaro Tao (FAX: 075-753-6068, Email: rtao@kais.kyoto-u.ac.jp). mond. This is the first report identifying and characterizing S-RNase in almond.
167 citations
TL;DR: The results suggest that the large subunit of Rubisco was directly degraded to the 37-kDa fragment containing the N-terminal region and the 16-k da fragmentcontaining the C-Terminal region of the LSU by active oxygen, probably the hydroxyl radical, generated in the lysates of chloroplasts.
Abstract: Lysates of chloroplasts isolated from wheat (Triticum aestivum L. cv. Aoba) leaves were incubated on ice (pH 5.7) for 0 to 60 min in light (15 mumol quanta m-2 s-1), and degradation of the large subunit (LSU) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco: EC 4.1.1.39) was analyzed by applying immunoblotting with site-specific antibodies against the N-terminal, internal, and C-terminal amino acid sequences of the LSU of wheat Rubisco. The most dominant product of the breakdown of the LSU and that which was first to appear was an apparent molecular mass of 37-kDa fragment containing the N-terminal region of the LSU. A 16-kDa fragment containing the C-terminal region of the LSU was concomitantly seen. This fragmentation of the LSU was inhibited in the presence of EDTA or 1,10-phenanthroline. The addition of active oxygen scavengers, catalase (for H2O2) and n-propyl gallate (for hydroxyl radical) to the lysates also inhibited the fragmentation. When the purified Rubisco from wheat leaves was exposed to a hydroxyl radical-generating system comprising H2O2, FeSO4 and ascorbic acid, the LSU was degraded in the same manner as observed in the chloroplast lysates. The results suggest that the large subunit of Rubisco was directly degraded to the 37-kDa fragment containing the N-terminal region and the 16-kDa fragment containing the C-terminal region of the LSU by active oxygen, probably the hydroxyl radical, generated in the lysates of chloroplasts.
158 citations
TL;DR: The results suggest NTH15 is involved in tobacco morphogenesis and abnormal leaf morphology in transgenic plants results from altered hormone levels.
Abstract: The shoot apical meristem functions to generate the lateral organs of a plant throughout the vegetative and reproductive phases Homeobox genes play key roles in controlling such developmental programs, but their modes of action have not been well defined Here we describe isolation and biological functions of a novel tobacco homeobox gene, designated NTH15 (Nicotiana tabacum homeobox 15), from a tobacco shoot apex cDNA library NTH15 encodes a polypeptide of 342 amino acids, its homeodomain is very similar to the class 1 KNOTTED-type homeodomains NTH15 mRNA is mainly localized in corpus cells in the tobacco shoot apical meristem, but not in tunica layers nor in differentiated lateral organs The NTH15 cDNA was fused to the cauliflower mosaic virus 35S promoter and used to generate transgenic tobacco plants Almost all transgenic tobacco plants showed abnormal leaf and/or flower morphology, and were categorized into three groups depending on severity of the leaf phenotype In transgenic leaves, drastic decrease of GA1 and increase of cytokinin were observed, while the levels of other phytohormones were only slightly changed Taken together, our results suggest NTH15 is involved in tobacco morphogenesis and abnormal leaf morphology in transgenic plants results from altered hormone levels
147 citations
TL;DR: The results suggest that gravity perception may require a greater plastid mass in hypocotyls compared to roots, and that 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.
Abstract: 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.
TL;DR: Cl cloning and expression analysis of a sucrose transporter cDNA from a monocot (the rice plant, Oryza sativa L.) and when OsSUT1 cDNA was introduced into yeast and expressed, the cells rapidly accumulated sucrose demonstrating that OsS UT1 does, in fact, encode a Sucrose transporter.
Abstract: We describe the cloning and expression analysis of a sucrose transporter cDNA from a monocot (the rice plant, Oryza sativa L.). The cDNA clone (OsSUT1) encoded an open reading frame of 1,611 bp (537 amino acids) and showed 76.8 to 79.7% similarity at the amino acid level to other sucrose transporters of dicot species. The predicted membrane topology of OsSUT1 protein is made up of 12 transmembrane helices which is consistent with most of the mono- and disaccharide transporters previously identified. When OsSUT1 cDNA was introduced into yeast and expressed, the cells rapidly accumulated sucrose demonstrating that OsSUT1 does, in fact, encode a sucrose transporter. From genomic Southern hybridization OsSUT1 appeared to be a single copy gene. OsSUT1 was expressed in source organs such as leaf blade, leaf sheath and germinating seed whereas little or no expression was observed in some sink organs such as the panicles before heading and the roots. Transcript was observed at high levels in panicles after heading, particularly in the portion containing endosperm and embryo. In addition, expression of OsSUT1 was high in etiolated seedlings and decreased during light-induced greening.
TL;DR: The results suggest that a strong positive feedback regulation is involved in ethylene biosynthesis at the gene transcriptional level in tomato fruit, even at the stage with a burst of ethylene production.
Abstract: We have examined whether or not a positive feedback regulation of gene expression for 1-aminocyclopropane-1-carboxylate (ACC) synthase and ACC oxidase also operates in ripening tomato (Lycopersicon esculentum) fruit during the burst of ethylene production. Two cDNA fragments for ACC synthase and one for ACC oxidase were cloned with high homology to already known genes involved in ethylene biosynthesis in ripening tomato fruit. Accumulation of mRNAs which hybridize to these cDNA probes were induced in mature green fruit within two days by treatment with propylene. In the fruit ripened from the turning stage, red color development, ethylene production, ACC content, and activities of ACC synthase and ACC oxidase increased as maturity progressed. The abundance of two ACC synthase and one ACC oxidase mRNAs in the fruit increased from the turning to pink stage and were followed by a slight decline towards the red stage. These increases in mRNAs abundance with ripening were prevented to a large extent by treatment with the ethylene action inhibitor, 1-methylcyclopropene (MCP). This was most pronounced in the fruit treated with MCP at the turning stage, in which the accumulation of ACC synthase and ACC oxidase transcripts was almost completely eliminated in the first two d, precisely the same stage at which the control fruit had the greatest level of each mRNA accumulation. The inhibition of transcript accumulation recovered to the control level within two to four d. MCP also decreased ethylene biosynthetic activity, although this decrease did not reflect the reduction in the mRNAs accumulation. These results suggest that a strong positive feedback regulation is involved in ethylene biosynthesis at the gene transcriptional level in tomato fruit, even at the stage with a burst of ethylene production.
TL;DR: Differences in expression of sodB and sodCp might explain why both plastid-located superoxide dismutase enzymes are needed, particularly under stress conditions.
Abstract: Chloroplasts of Nicotiana tabacum have two superoxide dismutases: a Fe- and a CuZn-containing enzyme, encoded by the nuclear genes sodB and sodCp, respectively. As a first step in studying the physiological function of these two enzymes, we compared the expression of sodB and sodCp in different plant organs, in response to hormonal treatments, and upon treatment with paraquat and Norflurazon. The sodCp transcript and active enzyme were detected only in young leaves of mature plants. The sodB transcript was more abundant in young compared to old leaves, but the enzymatic activity was higher in mature and senescent leaves. sodCp and sodB exhibited a different expression pattern upon treatment with abscisic acid, indole-3-acetic acid, kinetin, gibberellin, and 1-aminocyclopropane-1-carboxylate. Paraquat treatment caused a decrease in abundance of both transcripts, although the dose dependency of this decrease differed. Norflurazon-induced photooxidation resulted in a 10-fold increase of sodCp mRNA whereas the sodB transcript level was 25% higher than the control. These differences in expression might explain why both plastid-located superoxide dismutase enzymes are needed, particularly under stress conditions.
TL;DR: Changes in CuZn-SOD activity and content in isolated wheat chloroplasts under the light and the involvement of protease(s) and/or active oxygen species in this process were studied.
Abstract: Changes in CuZn-SOD activity and content in isolated wheat chloroplasts under the light, and the involvement of protease(s) and/or active oxygen species in this process were studied. Both SOD activity and content decayed with exposure time to photooxidative stress. Ascorbate, a H2O2 scavenger, prevented photooxidation-associated inactivation of SOD, while benzoate, a .OH scavenger, prevented SOD degradation. Wheat chloroplasts incubated in the dark did not hydrolyze exogenous or endogenous SOD, either H2O2-pretreated or not. Protease inhibitors did not prevent SOD degradation under photooxidative treatment, suggesting that plastid protease(s) did not participate in this process. Purified chloroplast CuZn-SOD was exposed to H2O2 and O2- or .OH-generating systems. O2- had no effect on either SOD activity or stability (estimated by native PAGE). H2O2 up to 700 microM inhibited SOD in a dose-dependent manner and induced charge/mass changes as seen by native PAGE. .OH also reduced SOD activity by inducing its fragmentation. High levels of active oxygen, as can be generated under strong stress conditions, could directly inactivate and degrade chloroplastic SOD.
TL;DR: Results suggest that PV72 and PV82 are potential sorting receptors for 2S albumin to protein-storage vacuoles.
Abstract: Precursor-accumulating (PAC) vesicles were previously shown to mediate the transport of the precursor of a major storage protein (pro2S albumin) to protein-storage vacuoles in developing pumpkin cotyledons. In this study, we characterized two homologous proteins from PAC vesicles, a 72 kDa protein (PV72) and an 82 kDa protein (PV82). PV72 and PV82 showed an ability to bind to peptides derived from both an internal propeptide and a C-terminal peptide of pro2S albumin. PV72 was predicted to be a type I integral membrane protein with epidermal growth factor (EGF)-like motifs. These results suggest that PV72 and PV82 are potential sorting receptors for 2S albumin to protein-storage vacuoles.
TL;DR: These phenomena indicate that apoptosis-like cell death can occur under salt stress.
Abstract: Salt stress-induced cell death was investigated in barley roots. Cleavage of nuclear DNA was observed 1 h after salt stress. Oligonucleosomal fragments of DNA were detected electrophoretically 8 h after salt stress. These phenomena indicate that apoptosis-like cell death can occur under salt stress.
TL;DR: The endo-1,3-beta-glucanase mRNA level was very low in the elongating cells but increased gradually at the onset of secondary wall synthesis, accompanying the massive deposition of cellulose.
Abstract: mRNAs for cell wall-related enzymes in developing cotton fiber cells were measured by reverse transcription-PCR analysis. Both endo-1,4-beta-glucanase and expansin mRNA levels were high during cell elongation but decreased when cell elongation ceased, and xyloglucan decreased. The endo-1,3-beta-glucanase mRNA level was very low in the elongating cells but increased gradually at the onset of secondary wall synthesis, accompanying the massive deposition of cellulose. Endoxyloglucan transferase and sucrose synthase mRNA levels were constant during all stages of growth.
TL;DR: The results suggest that the peroxidation of Iipids in the plasma membrane that is caused by both Al and Fe leads to the accumulation of Al andFe and the loss of viability.
Abstract: Aluminum (Al) and ferrous iron [Fe(II)] are separately non-toxic to cultured tobacco cells in nutrient solution. However, Al and Fe(II) together cause the peroxidation of membrane iipids, the accumulation of Al and Fe, and the loss of viability [Ono et al. (1995) Plant Cell Physiol. 36: 115]. We investigated the cause-and-effect relationships of these various responses. In cells exposed to Fe(TT) or Fe(III)-EDTA, both the peroxidation of Iipids and the loss of viability were similarly enhanced by A1C13 in a dose-dependent manner. During exposure to A1C13, the accumulation of Al and the loss of viability became apparent rapidly and simultaneously at 8 h, whereas both the peroxidation of Iipids and the accumulation of Fe occurred at later times. However, lipophilic antioxidants protected cells efficiently not only from the peroxidation of Iipids but also from the loss of viability and the accumulation of Al and Fe. These results suggest that the peroxidation of Iipids in the plasma membrane that is caused by both Al and Fe leads to the accumulation of Al and Fe and the loss of viability.
TL;DR: The data demonstrate that cytokinin or auxin overproduction decreases the content of the other hormone, apparently by decreasing its rate of synthesis and/or transport, rather than by increasing rates of turnover or conjugation.
Abstract: Cytokinins and auxins are important regulators of plant growth and development, but there is incomplete and conflicting evidence that auxins affect cytokinin metabolism and vice versa. We have investigated these interactions in Nicotiana tabacum L. by separate in planta manipulation of levels of the hormones followed by analysis of the induced changes in the metabolism of the other hormone. Cytokinin-overproducing plants (expressing the Agrobacterium tumefaciens ipt gene) had lower than wild-type levels of free IAA, and reduced rates of IAA synthesis and turnover, but there were no differences in the profiles of metabolites they produced from fed IAA. Similarly, auxin-overproducing plants (expressing the A. tumefaciens iaaM and iaaH genes), had lower levels of the major cytokinins than wild-type plants and lower cytokinin oxidase activity, but there were no differences in the profiles of metabolites they produced from fed cytokinins. The data demonstrate that cytokinin or auxin overproduction decreases the content of the other hormone, apparently by decreasing its rate of synthesis and/or transport, rather than by increasing rates of turnover or conjugation. Implications for the importance of cytokinin : auxin ratios in plant development are considered. et al. 1988). Further evidence for the hormones' mutual effects on one another has come from studies of crown gall tissues in which mutations in Agrobacterium genes involved in cytokinin or auxin synthesis, led to changes in concentrations of both auxins and cytokinins (Akiyoshi et al. 1983, Rudelsheim et al. 1987, Ishikawa et al. 1988, McGaw et al. 1988). There are also reports of direct auxinactivation of the cytokinin-catabolising enzyme, cytokinin oxidase (Hare and Van Staden 1994). The data so far amassed on the reciprocal effects of cytokinins on auxins, and auxins on cytokinins, however, is incomplete and conflicting. Our understanding of these hormonal interactions and their role in regulating plant development would therefore be greatly enhanced by separate in planta manipulation of the levels of the hormones, followed by high-resolution, kinetic analysis of the induced changes in the metabolism of the other hormone. In this investigation we have used fully-regenerated, fertile, seedsown, cytokinin and auxin overproducing, transgenic Nicotiana tabacum L. plants to study effects of cytokinin on auxin metabolism and vice versa. The data obtained has given a number of insights into the interactions, or "cross-communication", between the two hormones.
TL;DR: It is shown that the polymerized, filamentous state of actin is necessary for plants to block fungal penetration, which strongly suggest that actin microfilaments may play important roles in the expression of nonhost resistance of higher plants.
Abstract: We investigated the role of actin microfilaments in nonhost resistance of higher plants. Here we present several lines of evidence to indicate that microfilaments are indeed involved in blocking fungal penetration of nonhost plants. Erysiphe pisi, a pathogen of pea, normally fails to penetrate into nonhost plants such as barley, wheat, cucumber and tobacco. When tissues of these nonhost plants were treated with cytochalasins, specific inhibitors of actin polymerization, this fungus became able to penetrate and formed haustoria in epidermal cells of these plants. Moreover, treatment of these plants with various kinds and concentrations of cytochalasins allowed several other nonpathogenic fungi, E.graminis hordei, E.graminis tritici, Sphaerotheca fuliginea, Colletotrichum graminicola, Mycosphaella pinodes, C. lagenarium, Altemaria kikuchiana and Corynespora melonis, to also penetrate the cells of these plants. The degree of microfilament depolymerization varied depending on the kinds and concentrations of cytochalasins applied and we show that this is significantly correlated with the penetration efficiency of C. graminicola. This indicates that the polymerized, filamentous state of actin is necessary for plants to block fungal penetration. These results strongly suggest that actin microfilaments may play important roles in the expression of nonhost resistance of higher plants.
TL;DR: Deduced amino acid sequences encoded by the cDNAs related to the MIP gene family from Nicotiana excelsior were characterized and suggested that they are water channel proteins localized in the plasma membrane.
Abstract: Deduced amino acid sequences encoded by the cDNAs related to the MIP gene family from Nicotiana excelsior were characterized. Phylogenetic characterization of the products of corresponding genes named NeMip1, NeMip2, and NeMip3 strongly suggested that they are water channel proteins localized in the plasma membrane. Organ specificity of the gene expression was examined in leaves, roots, and reproductive organs. NeMip1 was expressed in roots and reproductive organs; however, it was hardly detectable in leaves. Two other genes, NeMip2 and NeMip3, were expressed in all of organs examined. mRNA accumulation from the genes was investigated in leaves under salt- and drought-stresses. The results demonstrated that mRNA accumulation from all three genes increased under salt- and drought-stresses within one day. However, they showed different accumulation patterns. In addition to their up-regulation under salt- and drought-stresses, daily changes in NeMip2 and NeMip3 mRNA accumulation was observed under unstressed conditions in leaves.
TL;DR: Strict structural requirements for the induction of ion fluxes were similar to those of specific binding to the putative plasma membrane receptor as well as a series of signaling events specifically induced by the oligosaccharides, suggesting the involvement of transient changes in cytoplasmic ion concentration in oligOSaccharide signaling for defense responses.
Abstract: /V-Acetylchitooligosaccharides, fragments of a main backbone polymer of fungal ceil wall, elicit defense responses including phytoalexin production in suspensioncultured rice cells. The purified oligosaccharide triggers rapid, transient membrane depolarization. Ion fluxes induced by the oligosaccharides were analyzed by using ionselective electrodes. Treatment of the cells with the oligosaccharides induced transient efflux of K + and influx of H + immediately after the elicitation. To monitor the pH values of the cytoplasm and the vacuoles noninvasively under a physiological condition, in vivo 31 P-nuclear magnetic resonance spectroscopy was applied to the cells to which oxygenated growth medium was perfused continuously. The cytoplasmic pH showed significant transient decrease, correspondingly. Only the N-acetylchitooligosaccharides with a degree of polymerization higher than 5 were active, whereas deacetylated chitosan oligomers caused no effect. Less than 1 nM of iV-acetylchitoheptaose was sufficient to induce rapid flux of ions. Such strict structural requirements for the induction of ion fluxes were similar to those of specific binding to the putative plasma membrane receptor as well as a series of signaling events specifically induced by the oligosaccharides, suggesting the involvement of transient changes in cytoplasmic ion concentration in oligosaccharide signaling for defense responses.
TL;DR: Immunocytochemical analysis demonstrated that BiP is localized in a non-stochastic fashion in the endoplasmic reticulum membrane complex of developing endosperm cells, which support a model which proposes thatBiP associates with the newly synthesized prolamine polypeptide to facilitate its folding and assembly into a protein inclusion body, and is then recycled.
Abstract: The ER luminal binding protein, BiP, has been linked to prolamine protein body formation in rice. To obtain further information on the possible role of this chaperone in protein body formation we have cloned and sequenced a BiP cDNA homolog from rice endosperm. The rice sequence is very similar to the maize BiP exhibiting 92% nucleotide identity and 96% deduced amino acid sequence identity in the coding region. Substantial amino acid sequence homology exists between rice BiP and BiP homologs from several other plant and animal species including long stretches of conservation through the amino-terminal ATPase domain. Considerable variation, however, is observed within the putative carboxy-terminal peptide-binding domain between the plant and nonplant BiP sequences. A single hand of approximately 2.4 kb was visible when RNA gel blots of total RNA purified from seed tissue were probed with radiolabeled rice BiP cDNA. This band increased in intensity during seed development up to 10 days after flowering, and then decreased gradually until seed maturity. Protein gel blots indicated that BiP polypeptide accumulation parallels that of the prolamine polypeptides throughout seed development. Immunocytochemical analysis demonstrated that BiP is localized in a non-stochastic fashion in the endoplasmic reticulum membrane complex of developing endosperm cells. It is abundant on the periphery of the protein inclusion body but not in the central portion of the protein body or in the cisternal ER membranes connecting the protein bodies. These data support a model which proposes that BiP associates with the newly synthesized prolamine polypeptide to facilitate its folding and assembly into a protein inclusion body, and is then recycled.
TL;DR: Results clearly revealed that both elastic and viscous properties of avocado mesocarp tissues were substantially altered during ripening, and that the solubility changes in acidic polysaccharides and decreases in the average molecular weight of cell wall xyloglucan components were associated with significant changes in fruit texture.
Abstract: Changes in avocado (Persea americana) fruit texture during ripening were evaluated by stress-relaxation analysis. A conical probe was imposed into the mesocarp tissue to a depth of 0.6 mm and the initial stress and the stress relaxation over 60 s were determined. The initial stress, an elastic parameter, was substantially reduced within one day when ripening was initiated by transferring the fruit from 15 to 25°C. The minimum and maximum relaxation time, parameters which reflect viscosity, were also reduced within one day. Mesocarp cell walls were fractionated into water-soluble (WS), hot EDTA-soluble (EDTA), alkaline soluble (hemicellulose) and the residual (cellulose) fractions. The amount of cellulose did not change during ripening. Rhamnose, arabinose and uronic acids in the WS fraction increased during the initial day of ripening; those same components decreased in the EDTA fraction. A molecular weight downshift in the WS acidic polysaccharides was detected within one day, while only slight changes were observed in the molecular weights of the EDTA fraction. The quantities of individual sugar components of major hemicellulose fraction were unchanged, but there was a prominent molecular weight downshift in the xyloglucan components within one day. These results clearly revealed that both elastic and viscous properties of avocado mesocarp tissues were substantially altered during ripening, and that the solubility changes in acidic polysaccharides and decreases in the average molecular weight of cell wall xyloglucan components were associated with significant changes in fruit texture.
TL;DR: It is suggested that the NADPH-binding subunit of NDH complex is easily dissociated from the thylakoid membranes during the process of the washing the thyakoids by centrifugation, suggesting that this is mediated mainly by FQR.
Abstract: activity was suppressed by inhibitors of the respiratory NADH dehydrogenase (NDH) complex in mitochondria, capsaicin and amobarbital, suggesting that the activity was mediated by chloroplastic NDH complex. Antimycin A, an inhibitor of ferredoxin-quinone reductase (FQR), and the protonophore nigericin also inhibited the increase in Chi fluorescence by NADPH. By contrast, JV-ethylmaleimide (NEM), an inhibitor of ferredoxin-NADP+ reductase (FNR), did not suppress the fluorescence increase, showing that FNR is not involved in this reaction. When the osmotically ruptured chloroplasts were washed by centrifugation, a further addition of ferredoxin as well as NADPH was required for an increase in fluorescence. This ferredoxin-dependent activity also was suppressed by antimycin A, but only partly inhibited by capsaicin or amobarbital, suggesting that this is mediated mainly by FQR. These findings suggest that the NADPH-binding subunit of NDH complex is easily dissociated from the thylakoid membranes during the process of the washing the thylakoids by centrifugation.
TL;DR: The level of a protein analogous to VM23 (VM23P) was especially high during the active cell-expansion stage in young fruit, and VM23P might, therefore, play an important role in the rapid expansion of cells as a vacuolar water channel.
Abstract: Vacuolar H(+)-ATPase (V-ATPase) was purified from pear fruit and antibodies were raised against the subunits of 55 and 33 kDa. Antibodies against mung bean H(+)-pyrophosphatase (V-PPase) and radish VM23, which is a tonoplast intrinsic protein (TIP) and a water channel, cross-reacted with the vacuolar membrane proteins of pear fruit. To clarify the roles of these proteins in development of pear fruit, we determined their levels relative to the total amount of protein by immunoblot analysis. The levels of subunits of the V-ATPase increased with fruit development. By contrast, the level of V-PPase was particularly high at the cell-division stage and remained almost the same at other stages. The changes in the activities of V-ATPase and V-PPase corresponded to those in their protein levels. The ratio of V-PPase activity to V-ATPase activity indicated that V-PPase is a major H(+)-pump of the vacuolar membranes of young fruit and that the contribution of V-ATPase increases with fruit development, finally, V-ATPase becomes the major H(+)-pump during the later stages of fruit development. The level of a protein analogous to VM23 (VM23P) was especially high during the active cell-expansion stage in young fruit, and VM23P might, therefore, play an important role in the rapid expansion of cells as a vacuolar water channel. Our results show that the levels of V-ATPase, V-PPase and VM23P change differently and reflect the roles of the respective protein in the development of pear fruit.
TL;DR: To study the antifungal mechanism based on three dimensional structure of PR-5d, purified PR- 5d was crystallized and revealed that the crystals belong to space group C2, with cell dimensions a = 80.2 degrees, and diffract at least 1.8 A resolution.
Abstract: Cultured tobacco cells accumulate several pathogenesis-related proteins. A neutral PR-5 protein, PR-5d, was purified to homogeneity from such cells. PR-5d has highly hydrophobic characteristics, but hydropathy analysis of its primary structure did not show a hydrophobic domain. In a series of bioassays, purified PR-5d showed inhibitory activity against several phytopathogenic and non-phytopathogenic fungi as do other members of the PR-5 protein family. To study the antifungal mechanism based on three dimensional structure of PR-5d, purified PR-5d was crystallized. The preliminary X-ray analysis of the crystal revealed that the crystals belong to space group C2, with cell dimensions a = 80.2 A, b = 63.8 A, c = 45.7 A, and beta = 107.2 degrees, and diffract at least 1.8 A resolution.