Variation in growth form in relation to spectral light quality (red/far-red ratio) in Plantago lanceolata L. in sun and shade populations

Plants from a sun and shade population were grown in two environments differing in the ratio of red to far-red light (R/FR ratio). A low R/FR ratio, simulating vegetation shade, promoted the formation of long, upright-growing leaves and allocation towards shoot growth, whereas a high R/FR ratio had the opposite effects. The increase in plant height under the low R/FR ratio was accompanied by a reduction in the number of leaves. Population differences in growth form resembled the differences between plants grown in different light environments: plants from the shade population had rosettes with long erect leaves, whereas plants from the sun population formed prostrate rosettes with short leaves. Plants from the shade population were more responsive to the R/FR ratio than plants from the sun population: the increases in leaf length, plant height, and leaf area ratio under a low R/FR ratio were larger in the shade population. However, differences in plasticity were small compared to the population difference in growth form itself. We argue that plants do not respond optimally to shading and that developmental constraints might have limited the evolution of an optimal response. Received: 8 December 1996 / Accepted: 31 March 1997     

The C-terminal KDEL sequence increases the expression level of a single-chain antibody designed to be targeted to both the cytosol and the secretory pathway in transgenic tobacco

The effects of subcellular localization on single-chain antibody (scFv) expression levels in transgenic tobacco was evaluated using an scFv construct of a model antibody possessing different targeting signals. For translocation into the secretory pathway a secretory signal sequence preceded the scFv gene (scFv-S). For cytosolic expression the scFv antibody gene lacked such a signal sequence (scFv-C). Also, both constructs were provided with the endoplasmic reticulum (ER) retention signal KDEL (scFv-SK and scFv-CK, respectively). The expression of the different scFv constructs in transgenic tobacco plants was controlled by a CaMV 35S promoter with double enhancer. The scFv-S and scFv-SK antibody genes reached expression levels of 0.01% and 1% of the total soluble protein, respectively. Surprisingly, scFv-CK transformants showed considerable expression of up to 0.2% whereas scFv-C transformants did not show any accumulation of the scFv antibody. The differences in protein expression levels could not be explained by the steady-state levels of the mRNAs. Transient expression assays with leaf protoplasts confirmed these expression levels observed in transgenic plants, although the expression level of the scFv-S construct was higher. Furthermore, these assays showed that both the secretory signal and the ER retention signal were recognized in the plant cells. The scFv-CK protein was located intracellularly, presumably in the cytosol. The increase in scFv protein stability in the presence of the KDEL retention signal is discussed.     

Electron microscopic structural analysis of Photosystem I,Photosystem II,and the cytochromeb6/f complex from green plants and cyanobacteria

Electron microscopy (EM) in combination with image analysis is a powerful technique to study protein structure at low- and high resolution. Since electron micrographs of biological objects are very noisy, substantial improvement of image quality can be obtained by averaging individual projections. Crystallographic and noncrystallographic averaging methods are available and have been applied to study projections of the large protein complexes embedded in photosynthetic membranes from cyanobacteria and higher plants. Results of EM on monomeric and trimeric Photosystem I complexes, on monomeric and dimeric Photosystem II complexes, and on the monomeric cytochromeb6/f complex are discussed.     

Sugar beet guard cell protoplasts demonstrate a remarkable capacity for cell division enabling applications in stomatal physiology and molecular breeding

A highly-efficient protocol for the large-scale isolation ofguard cell protoplasts from sugar beet (Beta vulgaris L.) hasbeen developed. Optimization of conditions for culturing theseprotoplasts resulted in extensive cell division and colony formation,at frequencies exceeding 50%. Plants can subsequently be regeneratedfrom these guard cell-derived colonies. This provides definitiveconfirmation that, in sugar beet leaf protoplast populations,only guard cells are the source of totipotent protoplasts. Thesefindings are the outcome of a directed, non-empirical approachto overcoming plant cell recalcitrance which was initiated byexploiting computer-assisted microscopy to couple in vitro responseto cell origin. The results reaffirm the conclusion that, inplants, extreme degrees of cytodifferentiation need not entailterminal specialization. The responsive nature of this systemcan be ascribed to the unique use of cultures essentially comprisinga single in vivo cell type. A uniform model system has thusbeen created with potential for widespread application. Theirdistinct morphological (and mechanical) features make guardcells a valuable choice for studying various fundamental aspects,not only of stomatal physiology, but also of plant cell (de)differentiation,differential gene expression etc. Furthermore, an applied valuefor such a system can also be envisaged. Results indicate thatthese cells are highly amenable to genetic manipulation techniques.The importance of these observations to our understanding ofplant cell function and behaviour is discussed. Key words: Beta, guard cells, stomatal physiology, totipotency, transformation     

Development of a pollen-mediated transformation method forNicotiana glutinosa

The development is described of a new procedure to genetically transform plant species using the male gametophyte as a natural transformation vector. Our system avoids the need for complicated regeneration procedures thus making it broadly applicable. Naked plasmid DNA encoding kanamycin resistance and GUS activity was introduced by particle gun bombardment into mature pollen grains ofNicotiana glutinosa. Bombarded pollen was used for pollinations and the resulting seeds were selected for kanamycin resistance. Two different kanamycin-resistant plants, designated VIP A and VIP B, were obtained in two independent experiments. In VIP A, TR2-driven GUS activity was observed in vascular bundles, trichomes and in a small number of pollen grains. DNA gel blot analysis indicated that the introduced DNA was integrated independently into the genome of VIP A and VIP B. It was shown that male and female gametophyte development and seed set were highly aberrant in both VIP A and VIP B and that the offspring of self- and cross-pollinations did not contain the transgenes. This might be caused by a recombination event during the integration of the naked DNA resulting in a deletion of part of the target chromosome. After meiosis such a deletion is lethal for the gametes. Our observation that the transgenes were detected in DNA isolated from sporophytic tissues but not in DNA from VIP A and VIP B pollen grains is in line with this explanation. Future experiments designed to increase the frequency of transformation and to transfer the transgenes to the offspring are discussed.     

Inter-Network Interactions: Impact of Connections between Oscillatory Neuronal Networks on Oscillation Frequency and Pattern

Oscillations in electrical activity are a characteristic feature of many brain networks and display a wide variety of temporal patterns. A network may express a single oscillation frequency, alternate between two or more distinct frequencies, or continually express multiple frequencies. In addition, oscillation amplitude may fluctuate over time. The origin of this complex repertoire of activity remains unclear. Different cortical layers often produce distinct oscillation frequencies. To investigate whether interactions between different networks could contribute to the variety of oscillation patterns, we created two model networks, one generating on its own a relatively slow frequency (20 Hz; slow network) and one generating a fast frequency (32 Hz; fast network). Taking either the slow or the fast network as source network projecting connections to the other, or target, network, we systematically investigated how type and strength of inter-network connections affected target network activity. For high inter-network connection strengths, we found that the slow network was more effective at completely imposing its rhythm on the fast network than the other way around. The strongest entrainment occurred when excitatory cells of the slow network projected to excitatory or inhibitory cells of the fast network. The fast network most strongly imposed its rhythm on the slow network when its excitatory cells projected to excitatory cells of the slow network. Interestingly, for lower inter-network connection strengths, multiple frequencies coexisted in the target network. Just as observed in rat prefrontal cortex, the target network could express multiple frequencies at the same time, alternate between two distinct oscillation frequencies, or express a single frequency with alternating episodes of high and low amplitude. Together, our results suggest that input from other oscillating networks may markedly alter a network''s frequency spectrum and may partly be responsible for the rich repertoire of temporal oscillation patterns observed in the brain.     

The effectiveness of flower strips and hedgerows on pest control,pollination services and crop yield: a quantitative synthesis

Floral plantings are promoted to foster ecological intensification of agriculture through provisioning of ecosystem services. However, a comprehensive assessment of the effectiveness of different floral plantings, their characteristics and consequences for crop yield is lacking. Here we quantified the impacts of flower strips and hedgerows on pest control (18 studies) and pollination services (17 studies) in adjacent crops in North America, Europe and New Zealand. Flower strips, but not hedgerows, enhanced pest control services in adjacent fields by 16% on average. However, effects on crop pollination and yield were more variable. Our synthesis identifies several important drivers of variability in effectiveness of plantings: pollination services declined exponentially with distance from plantings, and perennial and older flower strips with higher flowering plant diversity enhanced pollination more effectively. These findings provide promising pathways to optimise floral plantings to more effectively contribute to ecosystem service delivery and ecological intensification of agriculture in the future.     

Cholesterol accumulation caused by low density lipoprotein receptor deficiency or a cholesterol-rich diet results in ectopic bone formation during experimental osteoarthritis

Introduction

Osteoarthritis (OA) is associated with the metabolic syndrome, however the underlying mechanisms remain unclear. We investigated whether low density lipoprotein (LDL) accumulation leads to increased LDL uptake by synovial macrophages and affects synovial activation, cartilage destruction and enthesophyte/osteophyte formation during experimental OA in mice.

Methods

LDL receptor deficient (LDLr^−/−) mice and wild type (WT) controls received a cholesterol-rich or control diet for 120 days. Experimental OA was induced by intra-articular injection of collagenase twelve weeks after start of the diet. OA knee joints and synovial wash-outs were analyzed for OA-related changes. Murine bone marrow derived macrophages were stimulated with oxidized LDL (oxLDL), whereupon growth factor presence and gene expression were analyzed.

Results

A cholesterol-rich diet increased apolipoprotein B (ApoB) accumulation in synovial macrophages. Although increased LDL levels did not enhance thickening of the synovial lining, S100A8 expression within macrophages was increased in WT mice after receiving a cholesterol-rich diet, reflecting an elevated activation status. Both a cholesterol-rich diet and LDLr deficiency had no effect on cartilage damage; in contrast, ectopic bone formation was increased within joint ligaments (fold increase 6.7 and 6.1, respectively). Moreover, increased osteophyte size was found at the margins of the tibial plateau (4.4 fold increase after a cholesterol-rich diet and 5.3 fold increase in LDLr^−/− mice). Synovial wash-outs of LDLr^−/− mice and supernatants of macrophages stimulated with oxLDL led to increased transforming growth factor-beta (TGF-β) signaling compared to controls.

Conclusions

LDL accumulation within synovial lining cells leads to increased activation of synovium and osteophyte formation in experimental OA. OxLDL uptake by macrophages activates growth factors of the TGF-superfamily.