Monoclonal antibodies to the light-harvesting chlorophyll a/b protein complex of photosystem II.

A collection of 17 monoclonal antibodies elicited against the light-harvesting chlorophyll a/b protein complex which serves photosystem II (LHC-II) of Pisum sativum shows six classes of binding specificity. Antibodies of two of the classes recognize a single polypeptide (the 28- or the 26- kD polypeptides), thereby suggesting that the two proteins are not derived from a common precursor. Other classes of antibodies cross-react with several polypeptides of LHC-II or with polypeptides of both LHC-II and the light-harvesting chlorophyll a/b polypeptides of photosystem I (LHC-I), indicating that there are structural similarities among the polypeptides of LHC-II and LHC-I. The evidence for protein processing by which the 26-, 25.5-, and 24.5-kD polypeptides are derived from a common precursor polypeptide is discussed. Binding studies using antibodies specific for individual LHC-II polypeptides were used to quantify the number of antigenic polypeptides in the thylakoid membrane. 27 copies of the 26-kD polypeptide and two copies of the 28-kD polypeptide were found per 400 chlorophylls. In the chlorina f2 mutant of barley, and in intermittent light-treated barley seedlings, the amount of the 26-kD polypeptide in the thylakoid membranes was greatly reduced, while the amount of 28-kD polypeptide was apparently not affected. We propose that stable insertion and assembly of the 28-kD polypeptide, unlike the 26-kD polypeptide, is not regulated by the presence of chlorophyll b.

Quantitative trait loci analysis of powdery mildew disease resistance in the Arabidopsis thaliana accession kashmir-1.

Powdery mildew diseases are economically important diseases, caused by obligate biotrophic fungi of the Erysiphales. To understand the complex inheritance of resistance to the powdery mildew disease in the model plant Arabidopsis thaliana, quantitative trait loci analysis was performed using a set of recombinant inbred lines derived from a cross between the resistant accession Kashmir-1 and the susceptible accession Columbia glabrous1. We identified and mapped three independent powdery mildew quantitative disease resistance loci, which act additively to confer disease resistance. The locus with the strongest effect on resistance was mapped to a 500-kbp interval on chromosome III.

The health belief model and adolescents with insulin-dependent diabetes mellitus.

We tested the predictive utility of the health belief model (HBM) for adherence with a complex, ongoing medical regimen in the context of a chronically ill youthful population (56 adolescent outpatients with insulin-dependent diabetes mellitus; mean age = 14 years). A three-construct model of health beliefs was tested: Threat (perceived susceptibility combined with severity), Benefits-Costs, and Cues to seek treatment. Multiple indicators of compliance were used, and metabolic control was measured by glycosylated hemoglobin. The Benefits-Costs and Cues constructs were related to compliance in the theoretically expected positive direction. Threat interacted with Benefits-Costs in the prediction of compliance and with Cues in the prediction of metabolic control. The greatest compliance was achieved with low perceived Threat and high perceived Benefits-Costs. Poor metabolic control was associated with high Threat and Cues. As age increased, adherence to the exercise, injection, and frequency components of the regimen decreased.

The spatial demands of graphs.

This study is about the spatial demands made by graphs and about the ability of young children to cope with these demands. We wanted to answer two questions: whether children can coordinate spatial information from two axes, and whether the fact that they have to extrapolate non-perpendicular lines in graphs causes them particular difficulty. Sixteen six- and 16 nine-year-olds were asked to extrapolate imaginary lines in a graph-like task in order to find the position on one axis of a graph corresponding to a given position on the other axis. In addition to these axis-to-axis extrapolations, they were given separate line-to-axis and axis-to-line problems requiring the extrapolation of only one line. The older children made significantly small errors than the younger ones, but on the whole both age groups made accurate extrapolations in all three types of problem. Most importantly the errors in the axis-to-axis condition were not significantly different from the mean summed errors in the other two conditions (line-to-axis and axis-to-line). This shows that children do not experience difficulty with the axis-to-axis condition over and above the difficulty of extrapolating two single lines. We found no evidence that the non-perpendicularity of the lines which the children had to extrapolate caused them any particular difficulty. We concluded that young children have a more adequate understanding of rectangular coordinate systems than has been evident from their performance in other tasks and that young children can in principle cope with the spatial demands made by graphs surprisingly well.

Purification of an infection-related, extracellular peroxidase from barley.

Increases in two extracellular peroxidases were observed following inoculation of barley (Hordeum vulgare L.) with the powdery mildew pathogen (Erysiphe graminis DC.: Fr. f. sp. hordei Em. Marchal). The more prominent isozyme, P8.5, was purified from intercellular wash fluids by acetone precipitation, ion-exchange chromatography, isoelectric focusing, and gel filtration. Purified P8.5 is a heme-containing, glycoprotein with a M(r) of 35,000. It has eight cysteine residues. A highly specific, high-titer antiserum to deglycosylated P8.5 was produced.

Use of Arabidopsis recombinant inbred lines reveals a monogenic and a novel digenic resistance mechanism to Xanthomonas campestris pv campestris.

Infiltration of the Arabidopsis thaliana accession Landsberg erecta (Ler) with Xanthomonas campestris pv campestris isolate 2D520 results in extensive necrosis and limited chlorosis within 5-6 days post-inoculation (d.p.i.), which can lead to systemic necrosis within 23 d.p.i. in contrast, the accession Columbia (Col) remains asymptomatic after infiltration. Although both accessions support bacterial growth, 5-28-fold more bacteria are present in Ler than in Col leaf tissue. Inheritance studies indicate that three independent, dominant or partially dominant, nuclear genes condition resistance to X. c. campestris 2D520. The major gene, termed RXC2, conditions monogenic resistance to X. c.; campestris and was mapped to a 5.5 cM interval of chromosome V. Segregation data indicate that the locus RXC3 in conjunction with RXC4 confers digenic resistance to X. c. campestris. The combined action of RXC3 and RXC4 is correlated with a suppression of in planta bacterial levels and a suppression of symptoms relative to Ler. The RXC3 + RXC4-mediated resistance is novel in that although the Col allele of RXC4 contributes positively to resistance, it is the Ler and not the Col allele of RXC3 that contributes positively to resistance. RXC3 was mapped to the bottom arm of chromosome V in a 2.7 cM interval within the major recognition gene complex MRC-J, a cluster of genes involved in disease resistance. RXC4 was mapped to a 12 cM interval on chromosome II that also contains RXC1, a gene conferring tolerance to X. c. campestris.

An Arabidopsis thaliana mutant defective in chloroplast dicarboxylate transport.

Reactions of the photorespiratory pathway of C(3) plants are found in three subcellular organelles. Transport processes are, therefore, particularly important for maintaining the uninterrupted flow of carbon through this pathway. We describe here the isolation and characterization of a photorespiratory mutant of Arabidopsis thaliana defective in chloroplast dicarboxylate transport. Genetic analysis indicates the defect is due to a simple, recessive, nuclear mutation. Glutamine and inorganic phosphate transport are unaffected by the mutation. Thus, in contrast to previous reports for pea and spinach, glutamine uptake by Arabidopsis chloroplasts is mediated by a transporter distinct from the dicarboxylate transporter. Both the inviability and the disruption of amino-group metabolism of the mutant under photorespiratory conditions suggest that the primary function of the dicarboxylate transporter in vivo is the transfer of 2-oxoglutarate and glutamate across the chloroplast envelope in conjunction with photorespiratory nitrogen metabolism. The role commonly ascribed to this transporter, conducting malate-aspartate exchanges for the indirect export of reducing equivalents from the chloroplast, appears to be a minor one.

Young children's use of spatial coordinates.

4-6-year-old children were given problems in which they had to decide which 1 of an array of points was in line with 2 coordinate markers. The simplest problems had 4 points to choose between and markers perpendicular to the horizontal and vertical axes. Children of all ages were able to extrapolate lines from both coordinates to solve these problems. The older children were also given more complex problems. In some of these, 1 marker was at 45 degrees to an axis, the other perpendicular: in others the array was increased to 16 points and presented sometimes in a regular, sometimes in an irregular pattern. There were developmental improvements in performance, and the complex problems were more difficult than the simpler ones. However, 5- and 6-year-olds did extremely well even on the complex problems. The results establish that young children's grasp of Euclidean spatial relationships is more adequate than has often been suggested.

Genetic characterization of five powdery mildew disease resistance loci in Arabidopsis thaliana.

This paper reports on six Arabidopsis accessions that show resistance to a wild isolate of the powdery mildew pathogen, Erysiphe cichoracearum. Resistance at 7 days post-inoculation in these accessions was characterized by limited fungal growth and sporadic development of chlorotic or necrotic lesions at inoculation sites. Three accessions, Wa-1, Kas-1 and SI-0, were highly resistant, while the other accessions permitted some fungal growth and conidiation. Papilla formation was a frequent host response; however, cell death appeared to be neither a rapid nor a common response to infection. To determine the genetic basis of resistance, segregation analyses of progeny from crosses between each of the resistant accessions and Columbia (gl1), which is susceptible to the powdery mildew pathogen, were performed. For all accessions except Sl-0, resistance was conferred by a single locus. Sl-0 was unique in that two unlinked loci controlled the disease reaction phenotype. In accessions Wa-1, Kas-1, Stw-0 and Su-0, powdery mildew resistance was encoded by a semi-dominant allele. However, susceptibility was dominant to resistance in accessions Te-0 and Sl-0. Mapping studies revealed that powdery mildew resistances in Kas-1, Wa-1, Te-0, Su-0 and Stw-0 were controlled by five independent loci. This study suggests that the Arabidopsis powdery mildew disease will be a suitable model system in which to investigate powdery mildew diseases.

Two levels of inferential behavior in young children.

5- and 6-year-old children made inferences about the spatial locations of animals and people in a series of 3 experiments. The tasks employed manipulable models to represent the spatial relations involved and were made as simple as possible. 2 levels of inferential behavior were found. The first constituted the ability to draw an inference consistent with information given, but with minimal understanding of the way in which inferences can assist in decisions between alternative outcomes. At the second level, children succeeded in discriminating inferences which were logically necessary from those which were merely consistent with the premises. Most 5-year-olds were at the first level, most 6-year-olds at the second level. 2 criteria for the identification of young children's behavior as inferential were established, and the results of the present study were discussed in terms of recent related work with both younger and older children.

Phytoalexin Accumulation in Arabidopsis thaliana during the Hypersensitive Reaction to Pseudomonas syringae pv syringae.

Inoculation of leaves of Arabidopsis thaliana (L.) Heynh. with the wheat pathogen, Pseudomonas syringae pv syringae, resulted in the expression of the hypersensitive reaction and in phytoalexin accumulation. No phytoalexin accumulation was detected after infiltration of leaves with a mutant of P. s. syringae deficient in the ability to elicit a hypersensitive reaction; with the crucifer pathogen, Xanthomonas campestris pv campestris; or with 10 millimolar potassium phosphate buffer (pH 6.9). Phytoalexin accumulation was correlated with the restricted in vivo growth of P. s. syringae. A phytoalexin was purified by a combination of reverse phase flash chromatography, thin layer chromatography, followed by reverse phase high performance liquid chromatography. The Arabidopsis phytoalexin was identified as 3-thiazol-2'-yl-indole on the basis of ultraviolet, infrared, mass spectral, (1)H-nuclear magnetic resonance, and (13)C-nuclear magnetic resonance data.

Use of monoclonal antibodies in the purification and characterization of 1-aminocyclopropane-1-carboxylate synthase, an enzyme in ethylene biosynthesis.

1-Aminocyclopropane-1-carboxylate (ACC) synthase (EC 4.4.1.14), extracted from tomato pericarp tissue, was purified 6500-fold by conventional and high-performance liquid chromatography. Two-dimensional gel electrophoresis of this preparation indicated that ACC synthase activity was associated with a protein band at 50 kDa, a value consistent with size determinations by gel filtration. Monoclonal antibodies against ACC synthase were obtained from murine hybridoma cell lines. These antibodies recognized the native enzyme, as shown with an immunoprecipitation assay. A monoclonal IgG immunoaffinity gel was used to isolate, from a relatively crude enzyme preparation, a single protein, which migrated at 50 kDa in a NaDodSO(4)/polyacrylamide gel. In vivo labeling of wounded tomato pericarp tissue with [(35)S]methionine followed by immunoaffinity purification of ACC synthase yielded a radioactive protein of 50 kDa. We conclude that the 50-kDa protein represents ACC synthase in extracts of wounded tomato pericarp tissue.

Coordinated plant defense responses in Arabidopsis revealed by microarray analysis.

Disease resistance is associated with a plant defense response that involves an integrated set of signal transduction pathways. Changes in the expression patterns of 2,375 selected genes were examined simultaneously by cDNA microarray analysis in Arabidopsis thaliana after inoculation with an incompatible fungal pathogen Alternaria brassicicola or treatment with the defense-related signaling molecules salicylic acid (SA), methyl jasmonate (MJ), or ethylene. Substantial changes (up- and down-regulation) in the steady-state abundance of 705 mRNAs were observed in response to one or more of the treatments, including known and putative defense-related genes and 106 genes with no previously described function or homology. In leaf tissue inoculated with A. brassicicola, the abundance of 168 mRNAs was increased more than 2.5-fold, whereas that of 39 mRNAs was reduced. Similarly, the abundance of 192, 221, and 55 mRNAs was highly (>2.5-fold) increased after treatment with SA, MJ, and ethylene, respectively. Data analysis revealed a surprising level of coordinated defense responses, including 169 mRNAs regulated by multiple treatments/defense pathways. The largest number of genes coinduced (one of four induced genes) and corepressed was found after treatments with SA and MJ. In addition, 50% of the genes induced by ethylene treatment were also induced by MJ treatment. These results indicated the existence of a substantial network of regulatory interactions and coordination occurring during plant defense among the different defense signaling pathways, notably between the salicylate and jasmonate pathways that were previously thought to act in an antagonistic fashion.

Map positions of 47 Arabidopsis sequences with sequence similarity to disease resistance genes.

Map positions have been determined for 42 non-redundant Arabidopsis expressed sequence tags (ESTs) showing similarity to disease resistance genes (R-ESTs), and for three Pto-like sequences that were amplified with degenerate primers. Employing a PCR-based strategy, yeast artificial chromosome (YAC) clones containing the EST sequences were identified. Since many YACs have been mapped, the locations of the R-ESTs could be inferred from the map positions of the YACs. R-EST clones that exhibited ambiguous map positions were mapped as either cleavable amplifiable polymorphic sequence (CAPS) or restriction fragment length polymorphism (RFLP) markers using F8 (Ler x Col-0) recombinant inbred (RI) lines. In all cases but two, the R-ESTs and Pto-like sequences mapped to single, unique locations. One R-EST and one Pto-like sequence each mapped to two locations. Thus, a total of 47 loci were identified in this study. Several R-ESTs occur in clusters suggesting that they may have arisen via gene duplication events. Interestingly, several R-ESTs map to regions containing genetically defined disease resistance genes. Thus, this collection of mapped R-ESTs may expedite the isolation of disease resistance genes. As the cDNA sequencing projects have identified an estimated 63% of Arabidopsis genes, a very large number of R-ESTs (approximately 95), and by inference disease resistance genes of the leucine-rich repeat-class probably occur in the Arabidopsis genome.