SciCombinator

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Concept: Plant stem

173

Plants continuously extend their root and shoot systems through the action of meristems at their growing tips. By regulating which meristems are active, plants adjust their body plans to suit local environmental conditions. The transport network of the phytohormone auxin has been proposed to mediate this systemic growth coordination, due to its self-organising, environmentally sensitive properties. In particular, a positive feedback mechanism termed auxin transport canalization, which establishes auxin flow from active shoot meristems (auxin sources) to the roots (auxin sinks), has been proposed to mediate competition between shoot meristems and to balance shoot and root growth. Here we provide strong support for this hypothesis by demonstrating that a second hormone, strigolactone, regulates growth redistribution in the shoot by rapidly modulating auxin transport. A computational model in which strigolactone action is represented as an increase in the rate of removal of the auxin export protein, PIN1, from the plasma membrane can reproduce both the auxin transport and shoot branching phenotypes observed in various mutant combinations and strigolactone treatments, including the counterintuitive ability of strigolactones either to promote or inhibit shoot branching, depending on the auxin transport status of the plant. Consistent with this predicted mode of action, strigolactone signalling was found to trigger PIN1 depletion from the plasma membrane of xylem parenchyma cells in the stem. This effect could be detected within 10 minutes of strigolactone treatment and was independent of protein synthesis but dependent on clathrin-mediated membrane trafficking. Together these results support the hypothesis that growth across the plant shoot system is balanced by competition between shoot apices for a common auxin transport path to the root and that strigolactones regulate shoot branching by modulating this competition.

Concepts: Protein, Plant, Feedback, Root, Plant morphology, Plant stem, Meristem, Shoot

170

BACKGROUND: Plant grafting techniques have deepened our understanding of the signals facilitating communication between the root and shoot, as well as between shoot and reproductive organs. Transmissible signalling molecules can include hormones, peptides, proteins and metabolites: some of which travel long distances to communicate stress, nutrient status, disease and developmental events. While hypocotyl micrografting techniques have been successfully established for Arabidopsis to explore root to shoot communications, inflorescence grafting in Arabidopsis has not been exploited to the same extent. Two different strategies (horizontal and wedge-style inflorescence grafting) have been developed to explore long distance signalling between the shoot and reproductive organs. We developed a robust wedge-cleft grafting method, with success rates greater than 87%, by developing better tissue contact between the stems from the inflorescence scion and rootstock. We describe how to perform a successful inflorescence stem graft that allows for reproducible translocation experiments into the physiological, developmental and molecular aspects of long distance signalling events that promote reproduction. RESULTS: Wedge grafts of the Arabidopsis inflorescence stem were supported with silicone tubing and further sealed with parafilm to maintain the vascular flow of nutrients to the shoot and reproductive tissues. Nearly all (87%) grafted plants formed a strong union between the scion and rootstock. The success of grafting was scored using an inflorescence growth assay based upon the growth of primary stem. Repeated pruning produced new cauline tissues, healthy flowers and reproductive siliques, which indicates a healthy flow of nutrients from the rootstock. Removal of the silicone tubing showed a tightly fused wedge graft junction with callus proliferation. Histological staining of sections through the graft junction demonstrated the differentiation of newly formed vascular connections, parenchyma tissue and lignin accumulation, supporting the presumed success of the graft union between two sections of the primary inflorescence stem. CONCLUSIONS: We describe a simple and reliable method for grafting sections of an Arabidopsis inflorescence stem. This step-by-step protocol facilitates laboratories without grafting experience to further explore the molecular and chemical signalling which coordinates communications between the shoot and reproductive tissues.

Concepts: Reproductive system, Grafting, Plant stem, Plant reproduction, Rose, Rootstock, Tree shaping, Tomato grafting

168

Circumnutation, the helical movement of growing organ tips, is ubiquitous in land plants. The mechanisms underlying circumnutation have been debated since Darwin’s time. Experiments in space and mutant analyses have revealed that internal oscillatory (tropism-independent) movement and gravitropic response are involved in circumnutation. Female flower buds of tape grass (Vallisneria asiatica var. biwaensis) circumnutate on the water surface. Our observations and experiments with an artificial model indicated that gravitropism is barely involved in circumnutation. Instead, we show that helical intercalary growth at the base of peduncle plays the primary role in all movements in Vallisneria. This growth pattern produces torsional bud rotation, and gravity and buoyancy forces have a physical effect on the direction of peduncle elongation, resulting in bud circumnutation on the water surface. In contrast to other water-pollinated hydrophilous plants, circumnutation in Vallisneria enables female flowers to actively collect male flowers from a larger surface area of water.

Concepts: Plant, Bud, Buoyancy, Root, Flower, Plant stem, Plant sexuality, Surface area

168

BACKGROUND AND AIMS: Dendrobium longicornu, commonly known as the ‘Long-horned Dendrobium’, is an endangered and medicinally important epiphytic orchid. Over-exploitation and habitat destruction seriously threaten this orchid in Northeast India. Our objective was to develop an efficient protocol for the mass propagation of D. longicornu using axillary bud segments. METHODOLOGY AND PRINCIPAL RESULTS: Axillary buds cultured in Murashige and Skoog semi-solid medium supplemented with α-naphthalene acetic acid (NAA), 2,4-dichlorophenoxy acetic acid (2,4-D) and 6-benzylaminopurine (BAP) readily developed into plantlets. These formed either directly from shoot buds or from intermediary protocorm-like bodies (PLBs). The maximum explant response (86.6 %) was obtained in medium supplemented with NAA at 30 µM, while the maximum number of shoots (4.42) and maximum bud-forming capacity (3.51) were observed in medium containing 15 µM BAP and 5 µM NAA in combination. Protocorm-like bodies were obtained when the medium contained 2,4-D. The maximum number of explants forming PLBs (41.48 %) was obtained in medium containing 15 µM BAP and 15 µM 2,4-D. Well-developed plantlets obtained after 20-25 weeks of culture were acclimatized and eventually transferred to the greenhouse. Over 60 % of these survived to form plants ∼3-4 cm tall after 90 days in glasshouse conditions using a substrate of crushed brick and charcoal, shredded bark and moss. CONCLUSIONS: The method described can readily be used for the rapid and large-scale regeneration of D. longicornu. Its commercial adoption would reduce the collection of this medicinally important and increasingly rare orchid from the wild.

Concepts: Bud, Culture, Acetic acid, Plant stem, Vinegar, Shoot, Orchidaceae

160

Many plants respond to competition signals generated by neighbors by evoking the shade avoidance syndrome, including increased main stem elongation and reduced branching. Vegetation-induced reduction in the Red light:Far Red light (R:FR) provides a competition signal sensed by phytochromes. Plants deficient in phytochrome B (phyB) exhibit a constitutive shade avoidance syndrome including reduced branching. As auxin in the polar auxin transport stream (PATS) inhibits axillary bud outgrowth, its role in regulating the phyB branching phenotype was tested. Removing the main shoot PATS auxin source by decapitation or chemically inhibiting the PATS strongly stimulated branching in Arabidopsis thaliana deficient in phyB, but had a modest effect in WT. While indole-3-acetic acid (IAA) levels were elevated in young phyB seedlings, there was less IAA in mature stems compared to WT. A split plate assay of bud outgrowth kinetics indicated that low auxin levels inhibited phyB buds more than WT. Since the auxin response could be due either to the bud’s ability to export auxin into the main shoot PATS, or to auxin signaling status, both parameters were assessed. Main shoots of phyB had less absolute auxin transport capacity compared to WT, but equal or greater capacity when based on the relative amounts of native IAA in the stems. Thus, auxin transport capacity was unlikely to restrict branching. Both shoots of young phyB seedlings and mature stem segments showed elevated expression of auxin responsive genes and expression was further increased by auxin treatment, suggesting that phyB suppresses auxin signaling to promote branching.

Concepts: Bud, Arabidopsis thaliana, Arabidopsis, Plant physiology, Plant stem, Shoot, Phytochrome, The Stems

151

The outgrowth of axillary buds into branches is regulated systemically via plant hormones and the demand of growing shoot tips for sugars. The plant hormone auxin is thought to act via two mechanisms. One mechanism involves auxin regulation of systemic signals, cytokinins and strigolactones, which can move into axillary buds. The other involves suppression of auxin transport/canalization from axillary buds into the main stem and is enhanced by a low sink for auxin in the stem. In this theory, the relative ability of buds and stem to transport auxin controls bud outgrowth. Here we evaluate whether auxin transport is required or regulated during bud outgrowth in pea (Pisum sativum). The profound, systemic and long-term effects of the auxin transport inhibitor N-1-naphthylphthalamic acid had very little inhibitory effect on bud outgrowth in strigolactone deficient mutants. Strigolactones can also inhibit bud outgrowth in N-1-naphthylphthalamic acid-treated shoots that have greatly diminished auxin transport. Moreover strigolactones can inhibit bud outgrowth despite a much diminished auxin supply in in vitro or decapitated plants. These findings demonstrate that auxin sink strength in the stem is not important for bud outgrowth in pea. Consistent with alternative mechanisms of auxin-regulation of systemic signals, enhanced auxin biosynthesis in Arabidopsis thaliana can suppress branching in yuc1D plants compared to wild-type plants, but has no effect on bud outgrowth in a strigolactone-deficient mutant background.

Concepts: Bud, Annual plant, Plant physiology, Plant hormone, Plant stem, Shoot, Auxin, Polar auxin transport

151

Panax Notoginseng flower saponins (PNFS) are the main active component of Panax notoginseng (Burk) F. H. Chen flower bud (PNF) and possess significant anti-inflammatory efficacy. This study aims to explore the mechanisms underlying PNFS' antiflammatory action in RAW264.7 macrophages.

Concepts: Gene expression, Bud, Plant stem, Ginseng, Panax, Panax pseudoginseng, Notoginseng

149

The development of outgrowths from plant shoots depends on formation of epidermal sites of cell polarity convergence with high intracellular auxin at their centre. A parsimonious model for generation of convergence sites is that cell polarity for the auxin transporter PIN1 orients up auxin gradients, as this spontaneously generates convergent alignments. Here we test predictions of this and other models for the patterns of auxin biosynthesis and import. Live imaging of outgrowths from kanadi1 kanadi2 Arabidopsis mutant leaves shows that they arise by formation of PIN1 convergence sites within a proximodistal polarity field. PIN1 polarities are oriented away from regions of high auxin biosynthesis enzyme expression, and towards regions of high auxin importer expression. Both expression patterns are required for normal outgrowth emergence, and may form part of a common module underlying shoot outgrowths. These findings are more consistent with models that spontaneously generate tandem rather than convergent alignments.

Concepts: Gene, Cell, Adenosine triphosphate, Bud, Plant morphology, Plant stem, Shoot, Convergence

136

The production of tuberous roots is usually reduced by vigorous vegetative growth because of the competition for resource between the vegetative parts and reproductive organs. In this study, we conducted root pruning to examine the vigorous vegetative growth by regulating root growth, subsequently limiting vegetative growth and improving tuber yield. Compared with the control, stem, tuber, and root biomasses were all improved, whereas both flower and leaf biomasses were increased. Tuber biomass was improved by 23.48% to 50.32%, with the largest tuber biomass obtained at root cutting radius 4/5 R. With delayed root cutting time, tuber and root biomasses increased first and then decreased. The largest tuber biomass was obtained at 65 seedling stage. With a delay in root cutting time, the trend line of aboveground, underground, and total biomasses changed gradually. However, whereas underground and total biomasses showed a gradually increasing, aboveground biomass showed a decreasing. The values of stem-leaf and shoot-root ratios under different root cutting were higher than those of the control. With a delay in root cutting time, stem-leaf ratio showed an initial increase and then decreased with largest value being obtained at 80 seedling stage, whereas the largest shoot-root ratio was obtained at 115 seedling stage.

Concepts: Root, Plant morphology, Plant stem, Asexual reproduction, Tuber, Plant reproduction, Rhizome, Vegetative reproduction

81

Cannabis sativa L. is an important herbaceous species originating from Central Asia, which has been used in folk medicine and as a source of textile fiber since the dawn of times. This fast-growing plant has recently seen a resurgence of interest because of its multi-purpose applications: it is indeed a treasure trove of phytochemicals and a rich source of both cellulosic and woody fibers. Equally highly interested in this plant are the pharmaceutical and construction sectors, since its metabolites show potent bioactivities on human health and its outer and inner stem tissues can be used to make bioplastics and concrete-like material, respectively. In this review, the rich spectrum of hemp phytochemicals is discussed by putting a special emphasis on molecules of industrial interest, including cannabinoids, terpenes and phenolic compounds, and their biosynthetic routes. Cannabinoids represent the most studied group of compounds, mainly due to their wide range of pharmaceutical effects in humans, including psychotropic activities. The therapeutic and commercial interests of some terpenes and phenolic compounds, and in particular stilbenoids and lignans, are also highlighted in view of the most recent literature data. Biotechnological avenues to enhance the production and bioactivity of hemp secondary metabolites are proposed by discussing the power of plant genetic engineering and tissue culture. In particular two systems are reviewed, i.e., cell suspension and hairy root cultures. Additionally, an entire section is devoted to hemp trichomes, in the light of their importance as phytochemical factories. Ultimately, prospects on the benefits linked to the use of the -omics technologies, such as metabolomics and transcriptomics to speed up the identification and the large-scale production of lead agents from bioengineered Cannabis cell culture, are presented.

Concepts: Medicine, Biotechnology, Cell culture, Cannabis, Plant stem, Cannabinoid, Cannabis sativa