Regulation of growth by the trehalose pathway: Relationship to temperature and sucrose

Carbon signaling can override carbon supply in the regulation of growth. At least some of this regulation is imparted by the sugar signal trehalose 6-phosphate (T6P) through the protein kinase, SnRK1. This signaling pathway regulates biosynthetic processes involved in growth under optimal growing conditions. Recently, using a seedling system we showed that under sub-optimal conditions, such as cold, carbon signaling by T6P/ SnRK1 enables recovery of growth following relief of the stress. The T6P/ SnRK1 mechanism thus could be selected as a means of improving low temperature tolerance. High-throughput automated Fv/Fm measurements provide a potential means to screen for T6P/ SnRK1, and here we confirm through measurements of Fv/Fm in rosettes that T6P promotes low temperature tolerance and recovery during cold to warm transfer. Further, to better understand the coordination between sugars, trehalose pathway, and temperature-dependent growth, we examine the interrelationship between sugars, trehalose phosphate synthase (TPS), and trehalose phosphate phosphatase (TPP) gene expression and T6P content in seedlings. Sucrose, particularly when fed exogenously, correlated well with TPS1 and TPPB gene expression, suggesting that these enzymes are involved in maintaining carbon flux through the pathway in relation to sucrose supply. However, when sucrose accumulated to higher levels under low temperature and low N, TPS1 and TPPB expression were less directly related to sucrose; other factors may also contribute to regulation of TPS1 and TPPB expression under these conditions. TPPA expression was not related to sucrose content and all genes were not well correlated with endogenous glucose. Our work has implications for understanding acclimation to sink-limited growth conditions such as low temperature and for screening cold-tolerant genotypes with altered T6P/ SnRK1 signaling.     

Receptor Polymorphism and Genomic Structure Interact to Shape Bitter Taste Perception

The ability to taste bitterness evolved to safeguard most animals, including humans, against potentially toxic substances, thereby leading to food rejection. Nonetheless, bitter perception is subject to individual variations due to the presence of genetic functional polymorphisms in bitter taste receptor (TAS2R) genes, such as the long-known association between genetic polymorphisms in TAS2R38 and bitter taste perception of phenylthiocarbamide. Yet, due to overlaps in specificities across receptors, such associations with a single TAS2R locus are uncommon. Therefore, to investigate more complex associations, we examined taste responses to six structurally diverse compounds (absinthin, amarogentin, cascarillin, grosheimin, quassin, and quinine) in a sample of the Caucasian population. By sequencing all bitter receptor loci, inferring long-range haplotypes, mapping their effects on phenotype variation, and characterizing functionally causal allelic variants, we deciphered at the molecular level how a subjects’ genotype for the whole-family of TAS2R genes shapes variation in bitter taste perception. Within each haplotype block implicated in phenotypic variation, we provided evidence for at least one locus harboring functional polymorphic alleles, e.g. one locus for sensitivity to amarogentin, one of the most bitter natural compounds known, and two loci for sensitivity to grosheimin, one of the bitter compounds of artichoke. Our analyses revealed also, besides simple associations, complex associations of bitterness sensitivity across TAS2R loci. Indeed, even if several putative loci harbored both high- and low-sensitivity alleles, phenotypic variation depended on linkage between these alleles. When sensitive alleles for bitter compounds were maintained in the same linkage phase, genetically driven perceptual differences were obvious, e.g. for grosheimin. On the contrary, when sensitive alleles were in opposite phase, only weak genotype-phenotype associations were seen, e.g. for absinthin, the bitter principle of the beverage absinth. These findings illustrate the extent to which genetic influences on taste are complex, yet arise from both receptor activation patterns and linkage structure among receptor genes.     

Exceptional seed longevity and robust growth: ancient Sacred Lotus from China

A 1,288 ± 271-yr-old (1,350 ± 220 yr BP, radiocarbon age) seed of Sacred Lotus (Nelumbo nucifera Gaertn.) from an ancient lake bed at Pulantien, Liaoning Province, China, has been germinated and subsequently radiocarbon dated. This is the oldest demonstrably viable and directly dated seed ever reported, the preserved relict of one of the early crops of lotus cultivated by Buddhists at Pulantien after introduction of the religion into the region prior to 372 A.D. A small portion of the dry pericarp of a second lotus fruit from the same locale has been dated as being 332 ± 135-yr-old (270 ± 60 yr BP, radiocarbon age) by accelerator mass spectroscopy at the Lawrence Livermore National Laboratory. This polycentenarian seed not only germinated but is still growing (since March, 1994). Of six old lotus fruits tested, two-thirds germinated, almost all in fewer than 4 d, as rapidly as fruits harvested from the progeny of Pulantien Sacred Lotus plants (under cultivation by the National Park Service in Washington, DC), and more rapidly than fresh fruits of Yellow Lotus [N. lutea (Willd.) Pers.]. Growth of the old lotus is robust: rhizome formation and leaf emergence at rhizome nodes are more rapid than those of the Pulantien progeny, although the leaf width is smaller. Activity of the protein-repair enzyme L-isoaspartyl methyltransferase in the old lotus seed is persistent during germination and is as robust as that in the progeny, and the degree of aspartyl racemization in proteins of the two groups of plants is minimal and essentially identical. The six dated ancient Sacred Lotus fruits range in age from 95 to 1,288 yr (with a mean age of 595 ± 380 yr), evidently reflecting their production, deposition, and preservation at varying times during the intervening millennium.     

Physiological responses of Caloglossa leprieurii (Ceramiales, Rhodophyta) to salinity stress

Marine, estuarine and freshwater isolates of Caloglossa leprieurii (Montagne) J. Agardh exhibit a high salinity tolerance, reflected by broad cell viability and growth rate. Osmotic adjustment is shown to rely to a large extent on ion-transport systems, with K⁺ and Cl- accumulated in osmotically- significant quantities and active Na⁺ extrusion. The ion concentrations contribute a large proportion (67–94%) to internal osmotic pressure. The concentration of the organic osmolyte mannitol in all populations was strongly salinity dependent. Mannitol made a lower contribution to the internal osmotic pressure, when compared to ion concentrations, but nonetheless represented an important proportion of the internal osmolality. The response of the three isolates is discussed in relation to the salinity of their respective environments.     

Manually and automatically produced pellet cultures of human primary chondrocytes: A comparative analysis

Cartilage defects are often associated with restriction of the locomotor system. New methods are required to investigate cartilage tissue and for the repair of cartilage tissue. 3D cultures are promising due to better simulation of in vivo conditions. The aim of this study was to provide a model system for studying cartilage tissue. We solved this problem by automated production of pellet cultures of human primary chondrocytes in media with and without antibiotics using the Biomek® Cell Workstation and consequent automated bioscreening with a high‐throughput screening system, and compared with the regular manual processes. The Biomek® Cell Workstation allows the cultivation of different cell types (suspensions cells and adherent cells) and 3D cell cultures (pellet cultures, alginate beads and spheroid cultures). The proliferation was analyzed by DNA quantification and compared with the EZ4U proliferation assay as a new tool for pellet cultures. The toxicity was evaluated by the detection of ubiquitous adenylate kinases. The proliferation increased from day 14 until day 35 and was associated with a decrease in the cytotoxicity. The comparative analysis showed similar results for manual and automated processes. We concluded that the manual methods can be replaced by automated processes (pellet manufacturing and screening), which would allow large‐scale procedures to support studies on cartilage regeneration.     

Speicherung und mobilisierung von Kohlenhydraten und Lipiden in der Blattwespe Gilpinia hercyniae

Von Larven (drei Populationen, L₃, L₄-, L₄, L₅- und alte L₅-Larven), adulten Weibchen vor (I_v) und nach (I_n) Eiablage, Kokons (K) und Exuvien (E) wurden die Konzentrationen von löslichen Kohlenhydraten, Alanin, Prolin, Glykogen und Lipiden sowie die Frisch- und Trockengewichte bestimmt. Aus den Ergebnissen werden Rückschlüsse auf den Energiestoffwechsel der Entwicklungsstadien Embryo, Larve und Imago gezogen.

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Summary The concentrations of soluble carbohydrates (Fig. 2), alanine and proline (Fig. 3), glycogen and lipids (Fig. 4) and the weights (Fig. 1, Tab. I) of larvae (three populations, L₃, L₄-, L₄, L₅- and old L₅-instar larvae), adult females before (I_v) and after (I_n) oviposition, cocoons (K) and exuviae (E) (Tab. II) were determined. Embryonic, larval and adult stages of development have characteristic types of energy metabolism. The lipid catabolism of the embryonic stage is substituted during larval development by anabolic pathways based on resorbed sugars absorbed from spruce needles. During the first larval stages these soluble carbohydrates are directed to a glycogen pool in relatively large amounts. This pool is only slightly enlarged whereas lipids are synthesized in relatively large amounts between the 14th and 28th day after oviposition. Metamophosis is correlated with a decrease of glycogen. Lipids are used as an energy source by imagines and in eggs. Energy is distributed in the body by movement of trehalose in the larval stage and by a proline-alanine-shuttle in the adult stage.

     

Endokrinologische Untersuchungen an dem Wirt-Parasit-System: Pieris brassicae-Apanteles glomeratus

Die von Apanteles glomeratus L. parasitierten Raupen von Pieris brassicae zeigen in Abhängigkeit des Parasitierungstermins eine deutliche Veränderung des Juvenilhormon (JH)-Titer-Verlaufs während des letzten Larvenstadiums. Dabei tritt ein steiler Anstieg des JH-Gehaltes der Wirtshämolymphe im Zusammenhang mit der Häutung der Parasitenlarven vom 1. zum 2. Larvenstadium auf. Aufgrund von Ligations-experimenten konnte nachgewiesen werden, daß die Parasitenlarven selbst für den erhöhten JH-Titer ihrer Wirtsraupen verantwortlich sind, indem sie während ihrer Häutungsphase anscheinend JH in die Wirtshämolymphe abgeben.Eine durch die Parasitierung gesteigerte Syntheseaktivität läßt sich aus den Befunden histologischer Schnitte der Corpora allata frühparasitierter Raupen nicht feststellen. Dagegen weisen die Prothoraxdrüsen parasitierter Raupen zur Mitte des letzten Stadiums eine deutlich kleinere Querschnittsfläche auf als unparasitierte Tiere. Eine dadurch im Zusammenhang mit dem erhöhten JH-Titer bestehende Beziehung zur Häutungsunfähigkeit parasitierter Pieris-Raupen am Ende des letzten Larvenstadiums wird diskutiert.

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Summary The effects of parasitism by Apanteles glomeratus on the hemolymph juvenile hormone (JH) titers of Pieris brassicae during the last larval instar were determined using the Galleria bioassay.Depending on the time of parasitization, a significant increase of the JH titer could be observed when moulting of the parasites from the first to the second larval instar occurred.As neck-ligatured, parasitized Pieris larvae showed a similar increase of the JH titer at this time, it is concluded that the parasite larvae themselves are responsible for the elevation of the titer by delivering JH during their ecdysis into the host's hemolymph.This is supported by histological results from the corpora allata of parasitized and unparasitized caterpillars at the first and third day of the last larval instar, indicating no differences in its secretory activity. The prothoracid glands of parasitized host larvae, however, appear significantly smaller than those of comparable unparasitized ones in the middle of the last instar. A reduced secretory activity at this time, which is assumed from their decreased size, combined with an elevated JH titer may explain why parasitized larvae fail to moult at the end of their larval development.

     

Enhancement of BODIPY505/515 lipid fluorescence method for applications in biofuel-directed microalgae production

This paper describes a microalgal cell lipid fluorescence enhancement method using BODIPY(505/515), which can be used to screen for lipids in wild-type microalgae and to monitor lipid content within microalgae production processes to determine optimal harvesting time. The study was based on four microalgae species (Dunaliella teteriolecta, Tetraselmis suecica, Nannochloropsis oculata, and Nannochloris atomus) selected because of their inherent high lipid content. An extended analysis was carried out with N. oculata due to the depressed fluorescence observed when compared with the other experimental strains. BODIPY(505/515) lipid fluorescence was determined for two solvent pre-treatment methods (DMSO and glycerol) and four staining condition parameters (analysis time, staining temperature, dye concentration, and algal cell concentration). It was found that lipid fluorescence of thick cell-walled microalgae, such as N. oculata, is significantly enhanced by both the pre-treatment methods and staining condition parameters, thereby significantly enhancing lipid fluorescence by ca. 800 times the base autofluorescence. The lipid fluorescence enhancement method provides a quick and simple index for in vivo Flow Cytometry quantification of total lipid contents for purposes of species screening or whole culture monitoring in biofuel-directed microalgae production.     

Modelling the effects of climate change on the potential feeding activity of Thaumetopoea pityocampa (Den. & Schiff.) (Lep., Notodontidae) in France

Aim We investigated whether climate change has affected the potential feeding activity of a winter active larva, the pine processionary moth (PPM), Thaumetopoea pityocampa L., and whether it may explain its range expansion. Location The study area is France and, at a smaller scale, the Paris Basin. Methods We used a statistical model derived from Huchon and Démolin [1970 Revue Forestière Française (special issue: La lutte biologique en forêt), 220–234] to test whether their model, updated with climate change, could explain the observed range expansion. Since Battisti and colleagues have recently shown that climate could affect survival of the PPM through its effect on feeding activity, we also developed a mechanistic model based on larval feeding requirements (night air temperature above 0 °C and temperature inside the nest above 9 °C on the preceding day). We reconstructed the geographical distribution of feeding activity and we compared the resulting change with the PPM range expansion. Results The statistical model did not successfully predict the observed expansion but the mechanistic model showed considerable change in the feeding activity of the PPM. In the Paris Basin, the PPM border coincided with a zone unfavourable for feeding activity in the period 1992–96. Feeding conditions became more favourable in the period 2001–04, and the PPM succeeded in crossing this zone. Over larger temporal and spatial scales improved feeding conditions in the north‐western part of France were forecast by the mechanistic model. Main conclusions (1) The range distribution of the PPM in the Paris Basin is no longer limited by unfavourable feeding conditions. (2) The pattern of range expansion of the PPM is now governed mainly by its dispersal capabilities and host tree distribution. (3) At the country scale, this approach gives an approximate prediction of the potential distribution of the PPM, though the model may not be reliable in mountainous regions.