James W. Hart

The isolation and some properties of a lectin (Haemagglutinin) from Cucurbita phloem exudate

The occurrence of high haemagglutinating (lectin) activity in phloem exudate from three cucurbit species is reported. The protein responsible for this lectin activity in Cucurbita maxima Duch. has been isolated by cation exchange chromatography on Sepharose and identified by gel electrophoresis. The lectin showed agglutinating activity at concentrations as low as 0.1 μg/ml. No sugar, including those transported in the phloem of these species, interacted with agglutination. The lectin could not be extracted from cucurbit seed, but appeared in 5-day old seedlings. The possible role of a lectin in the sieve element is discussed.

Chemical and morphological studies on sites of sesquiterpene accumulation in Pogostemon cablin (patchouli).

Abstract Techniques are described for the micro-analysis of the free volatile sesquiterpenes in plant tissue by direct-loading gas liquid chromatography. Evidence is presented for the accumulation of sesquiterpenes, not only in the external glandular trichomes, but also in specialized internal accumulatory cells. The morphology and development of these external and internal structures are described as seen under light and electron microscopy. Also, while the apical dome of the stem neither shows any specialized accumulatory structures nor contains any detectable sesquiterpenes, the second pair of primordial leaves has glandular trichomes and a sesquiterpene concentration approximately twelve times higher than that in any other part of the plant.

The positive hook: the role of gravity in the formation and opening of the apical hook.

Photographic observations on germinating seedlings of Lepidium sativum L., Cucumis sativus L., and Helianthus annuus L. showed that the hypocotyl hook is not present in the seed but forms during the early stages of growth. Evidence that gravity plays a major role in inducing curvature of the hypocotyl, and in maintaining the hook once it has been formed, was obtained from clinostat experiments, from the use of morphactin to remove geotropic sensitivity and from inversion of seedlings to change the direction of the geostimulus. In L. sativum and H. annuus gravity perception seemed to be the only mechanism responsible for hook formation. In C. sativus hook formation was additionally aided by the mode of emergence of the cotyledons from the seed coat but gravity played an indirect role in regulating such emergence. Further evidence that hook formation is linked to a georesponse was derived from a comparison of hypocotyl development in wild-type Arabidopsis thaliana seedlings with that of an ageotropic mutant, hook formation being found to occur only in the wild type. Hook formation and maintenance is discussed in terms of contrasting geosensitivity between the apical and basal ends of the hypocotyl and it is suggested that light-induced hook opening is a reversal to a condition of uniformly negative georesponse throughout the hypocotyl.

Heterogeneity in phloem protein complements from different species : Consequences of hypotheses concerned with P-protein function.

Protein subunits present in phloem exudate from 17 cultivars, 5 species and 3 genera of the Cucurbitaceae have been fractionated by SDS-gel electrophoresis. The degree of difference in the phloem protein patterns appears to reflect the taxonomic relationships of the plants: there were major differences among genera, significant differences and similarities among species, and relatively few differences among cultivars of a single species. A representative of another family (Acer pseudoplatanus; Aceraceae) provided a completely different electrophoretic pattern. Each plant displayed a consistent protein pattern, irrespective of the organ from which exudate was obtained or the age and physiological status of the plant. Neither complete etiolation nor transition to the flowering phase effected any change in phloem proteins. The differences in phloem proteins between plants and the unusual properties of major subunits from different protein complements, render improbable many of the present ideas concerning the origin and function of P-protein.

Calcium-binding protein in sieve tube exudate.

A calcium-binding macromolecule, with an estimated molecular weight greater than 100,000, was detected in phloem exudate from Cucurbita maxima and related species. The macromolecule was a component of sieve tube sap, rather than a contaminant leached from cell walls or cut parenchyma cells during exudate collection. The protein nature of this macromolecule was deduced from its size, lability, susceptibility to proteolytic digestion, and by the dependence of calcium-binding activity on thiol-protecting agents. This protein is distinct from the major proteins of exudate and does not appear to be related to calmodulin.

A comparative analysis of phloem exudate proteins from Cucumis melo, Cucumis sativus and Cucurbita maxima by polyacrylamide gel electrophoresis and isoelectric focusing.

SummaryProteins in sieve tube exudate from Cucumis melo L., Cucumis sativus L. and Cucurbita maxima Duch. were analysed by gel electrophoresis and isoelectric focusing. Estimated molecular weights and isoelectric points for the major and minor proteins from each plant species are presented. Electrophoresis revealed striking differences between the protein complements of exudatc from the two genera investigated. Similarly, although a few exudate proteins from the two species of Cucumis possessed identical molecular weights, several major proteins were peculiar to each species. Isoelectric focusing of proteins in exudate samples from the three plants confirmed the marked differences in their protein complements. Furthermore, focusing also revealed differences between cultivars of Cucumis sativus. Both Cucumis sativus and Cucurbita maxima possessed relatively large amounts of basic proteins; these were absent in exudate from Cucumis melo. The implications of these results are discussed in relation to present concepts regarding the interrelationships and possible functional roles of P-proteins.

Phototropic responses of hypocotyls of etiolated and green seedlings

Abstract Hypocotyl curvature of cress, lettuce,and mustard and radish seedlings in continuous, unilateral radiation was much greater with green than with etiolated material. Exposure of cress seedlings to increasing periods of blue irradiation gave only little indication of a distinct first positive curvature type response with substantial curvature requiring relatively extensive irradiation (e.g. 60 min at 2 μE · m−2 · s−1). The greater photocurvature of green seedlings arose from a marked enhancement of this latter response to high irradiation dosage. Curvature in green seedlings also occured sooner and in a more basal region of the hypocotyl. When the cotyledons plus hypocotyl tips were capped with aluminium foil, blue light-induced photocurvature was totally prevented in etiolated cress seedlings, but only delayed and reduced in green material.

The heterogeneity of phloem exudate proteins from different plants: A comparative survey of ten plants using polyacrylamide gel electrophoresis.

SummaryProteins in sieve tube exudate from Ricinus communis L., Acer pseudoplatanus L., Aesculus hippocastanum L., Cucumis melo L., and two cultivars each of Cucumis sativus L., Cucurbita pepo L. and Cucurbita maxima Duchesne were fractionated and compared using polyacrylamide gel electrophoresis. Striking differences in major exudate proteins were displayed among the genera and species examined. Even cultivars within a single species, although showing general similarities, differed in some prominent proteins. Estimated molecular weights of the major exudate proteins from each plant are presented. The effects of reducing and chaotropic agents on the aggregation and subunit composition of exudate proteins from Cucumis sativus have been investigated. The problems involved in relating structure, function and biochemistry of P-protein are discussed.

Colchicine-binding protein from phloem and xylem of a higher plant.

SummaryColchicine has been demonstrated by ion-exchange and by gelfiltration assay to bind to a protein fraction derived from the higher plant Heracleum mantegazzianum. Colchicine-binding protein from a plant source was much more unstable than tubulin from animal preparations. The tissues of Heracleum vary in their content of colchicine-binding activity. No activity was obtained from non-vascular tissue. Phloem has at most, twice as much activity as xylem. The significance of these results is discussed in relation to a proposed degree of homology between P protein of phloem and microtubule protein.

Studies on the possible occurrence of actomyosin-like proteins in phloem

SummaryExtracts of large quantities of petioles and of isolated vascular tissues of Heracleum mantegazzianum have been analysed for actomyosin-like contractile proteins. Concentrated preparations, subjected to a standard isolation and purification procedure for actomyosin, failed to demonstrate either superprecipitation or viscosity changes in response to the addition of ATP and divalent cations. In addition, ATPase activities in phloem and xylem extracts have been fractionated by gel electrophoresis and characterised with regard to their substrate specificity, pH optima and ion requirements. Phloem extracts provide two phosphatases: one is non-specific in its substrate requirements; the other is a nucleoside triphosphatase but is stimulated only by monovalent cations and is also present in xylem extracts. All the enzymes are strongly inhibited by divalent cations and do not possess any of the characteristics of ATPases associated with contractile systems. The results are discussed in relation to the postulated involvement of contractile proteins in the translocation of sugars in phloem.