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A study of ecdysteroids control molting and metamorphosis in insects via a heterodimeric complex of two nuclear receptors, the ecdysone receptor (EcR) and ultraspiracle protein (Usp), is presented. Fluorescence resonance energy transfer (FRET) is used to study the natural pseudopalindromic element in complex with DNA -binding domains of Usp and EcR (UspDBD and EcRDBD).

The use of biostimulants (amino acid containing protein hydrolysate) in forestry field has received much less attention so far than in agriculture. Promising evidences were reported in literature with useful application for nursery activities, stimulating early growth and reducing transplanting stress. This study investigates the potential benefits induced by an amino acid-based animal-derived protein hydrolysate biostimulant (Siapton® by Isagro) in containerized Eucalyptus globulus seedlings following transplanting. Foliar and soil drench applications were applied using two different concentrations each (2.5 ml.l-1 and 5.0 ml.l-1 for foliar, 10 ml.l-1 and 20 ml.l-1 for soil). Measures on seedling height, apical shot formation, total aboveground (dry weight of leaves and stem) and belowground biomass (dry weight of roots) 120 days after transplanting were made to quantify the effect on growth. The contrasts analysis on results revealed a positive effect of the biostimulant on many of the measured parameters, especially with foliar application using the lowest concentration (2.5 ml.l-1). The foliar application was overall suggested as more efficient than soil drench also allowing lower inputs (i.e. biostimulant quantity). The main consequence of the treatment was an increased biomass allocation in the stem (above ground biomass) due to a stimulated leaves production which might suggest an increased photosynthetic activity and growth. Conversely no influence was detectable on total height of seedlings neither on the collar diameter. The biostimulant treatment on containerized Eucalyptus globulus positively influenced some features of seedlings' growth after transplanting and the use of biostimulant with foliar application during the hardening phase in the nursery, appears to be a promising technique to potentially improve seedling growth after transplanting. An interesting impact from application of biostimulant on biomass accumulation following transplanting was here demonstrated. Anyway, further research to verify the results on different tree species as well as under open field conditions is envisioned. [ABSTRACT FROM AUTHOR]

The present study examines the three-dimensional hygric and mechanical behavior of oak wood. The moisture equilibrium state, characterized by the sorption isotherms, was obtained from measurements taken during adsorption and desorption cycles. Sorption behavior was analyzed with the Dent theory and compared considering the sorption direction (adsorption/desorption cycle). Sorption parameters were provided for possible numerical applications in hygric material models. The corresponding swelling and shrinkage behavior was examined and characterized by the moisture expansion parameters for all anatomical directions. Orthotropic mechanical material behavior was characterized by determining the elastic engineering (Young's moduli, shear moduli, and Poisson's ratios) and the bending, compressive and compressive shear strength material parameters. Influence of moisture content (MC) on the mechanical material properties was studied using Young's moduli, Poisson's ratios, and the investigated strength parameters. A significant difference between the sorption behavior in adsorption and desorption, known as the hysteresis effect, could be proved. Furthermore, swelling and shrinkage behavior did not show any dependency on the adsorption/desorption cycle. The results confirm the significant influence of MC on the Young's moduli and the strength properties, however, did not validate an influence on the Poisson's ratios. [ABSTRACT FROM PUBLISHER]

Hardwood is increasingly trying to get established in the building industry in order to realise a larger material output and to take advantage of the superior mechanical properties of hardwood in comparison to softwood for structural applications. For calculations of stress and strain in boards and glued-laminated timbers it is essential to know the specific values of wood in the main directions: longitudinal, radial and tangential. Therefore, the specific values of Young's modulus, shear modulus, Poisson's ratio, the specific values of strength and also these of mass and heat flux (diffusion coefficient, thermal conductivity) were determined on European beech. [ABSTRACT FROM AUTHOR]

The influence of moisture content (MC) on the tension-compression (Te-Co) asymmetry of beech wood has been examined. The elastic and strength parameters, including Te and Co Young's moduli, Poisson's ratios, and ultimate and yield stress values, were determined and compared in terms of different MCs for all orthotropic directions. The results reveal a distinctive Te-Co strength asymmetry with a moisture dependency that is visualized clearly by the Te to Co yield stress ratio. The Te-Co asymmetry is further shown by the inequality of the elastic properties, known as the 'bimodular behavior'. The latter is proven for the Young's moduli values in the radial and tangential directions and for individual Poisson's ratios. Although the bimodularity of the Young's moduli is significant at low MC levels, there is no evidence of moisture dependency on the Te-Co asymmetry of the Poisson's ratios. [ABSTRACT FROM AUTHOR]

The heterodimer consisting of ecdysteroid receptor (EcR) and ultraspiracle (USP), both of which are members of the nuclear receptor superfamily, is considered to be the functional ecdysteroid receptor. Here we analyzed the subcellular distribution of EcR and USP fused to fluorescent proteins. The experiments were carried out in mammalian COS-7, CHO-K1 and HeLa cells to facilitate investigation of the subcellular trafficking of EcR and USP in the absence of endogenous expression of these two receptors. The distribution of USP tagged with a yellow fluorescent protein (YFP-USP) was almost exclusively nuclear in all cell types analyzed. The nuclear localization remained constant for at least 1 day after the first visible signs of expression. In contrast, the intracellular distribution of EcR tagged with a yellow fluorescent protein (YFP-EcR) varied and was dependent on time and cell type, although YFP-EcR alone was also able to partially translocate into the nuclear compartment. Coexpression of YFP-EcR with USP tagged with a cyan fluorescent protein (CFP-USP) resulted in exclusively nuclear localization of both proteins in all cell types analyzed. The USP-induced nuclear localization of YFP-EcR was stable for at least 20 hours. These experiments suggest that USP has a profound effect on the subcellular distribution of EcR. [ABSTRACT FROM AUTHOR]

Juvenile hormone (JH) and ecdysone are the key hormones controlling insect growth and development. The juvenile hormone binding protein (JHBP) is the first member in the array of proteins participating in JH signal transmission. In the present report a whole jhbp gene sequence (9790 bp) is described. The jhbp gene contains four introns (A-D). All the introns have common flanking sequences: GT at the 5' and AG at the 3' end. The first intron is in phase 1, the second in phase 2, and the third and fourth in phase 1. An analysis of these sequences suggests that U2-class spliceosomes are involved in intron excision from pre-mRNA. Several horizontally transmitted elements from other genes were found in the introns. All jhbp exons are positioned in local AT-reach regions of the gene. A search for core promoter regulatory elements revealed that the TATA box starts 29 bp preceding the start of transcription; the sequence TCAGTA representing a putative initiator sequence (Inr) starts at position +14. Eight characteristic sequences for binding Broad-Complex gene products, which coordinate the ecdysone temporal response, are present in the non-coding sequence of the jhbp gene. An analysis of exon locations and intron phases indicates that jhbp gene organization is related to the retinol binding protein gene, a member of the lipocalin family. [ABSTRACT FROM AUTHOR]

Studies the polarity of the ecdysone receptor complex's interaction with the palindromic response element from the hsp27 gene promoter. Heterodimer of two members of the nuclear hormone receptors superfamily; Function of natural 20E response elements as functionally asymmetric elements in a manner similar to the action of direct repeats in vertebrates; Theoretical significance of the study.

Drosophila melanogaster germ cell-expressed protein (GCE) belongs to the family of bHLH-PAS transcription factors that are the regulators of gene expression networks that determine many physiological and developmental processes. GCE is a homolog of D. melanogaster methoprene tolerant protein (MET), a key mediator of anti-metamorphic signaling in insects and the putative juvenile hormone receptor. Recently, it has been shown that the functions of MET and GCE are only partially redundant and tissue specific. The ability of bHLH-PAS proteins to fulfill their function depends on proper intracellular trafficking, determined by specific sequences, i.e. the nuclear localization signal (NLS) and the nuclear export signal (NES). Nevertheless, until now no data has been published on the GCE intracellular shuttling and localization signals. We performed confocal microscopy analysis of the subcellular distribution of GCE fused with yellow fluorescent protein (YFP) and YFP-GCE derivatives which allowed us to characterize the details of the subcellular traffic of this protein. We demonstrate that GCE possess specific pattern of localization signals, only partially consistent with presented previously for MET. The presence of a strong NLS in the C-terminal part of GCE, seems to be unique and important feature of this protein. The intracellular localization of GCE appears to be determined by the NLSs localized in PAS-B domain and C-terminal fragment of GCE, and NESs localized in PAS-A, PAS-B domains and C-terminal fragment of GCE. NLSs activity can be modified by juvenile hormone (JH) and other partners, likely 14-3-3 proteins.

The effect of pyridoxal 5′-phosphate on the binding of the ecdysteroid receptor from a nuclear extract of Drosophila rnelanogaster to DNA-cellulose was studied. The binding of hormone-receptor complexes to DNA- cellulose was completely blocked after a 30-min incubation with 3 mM pyridoxal 5′-phosphate at 0-4°C. The effect was specific for pyridoxal Y-phosphate since related compounds (pyridoxal, pyridoxamine 5′-phosphate and pyridoxamine) were not effective or gave only 17% inhibition (pyridoxal). Under standard conditions, none of the compounds tested exerted a significant effect on the stability of [3H](20R,22R)-2β,3β,14α,20,22-pentahydroxy-5β-cholest-7-en-6-one ([3H]ponasterone A)-receptor complexes. The loss of DNA-binding activity caused by pyridoxal 5′-phosphate is accompanied by changes in the molecular properties of [3H]ponasterone-A- receptor complexes. A shift of [3H]ponasterone-A binding was observed from the 8.0– 8.5 S to the 4.5–5.0 S region, when [3H]ponasterone-A-receptor complexes were exposed to pyridoxal 5′-phosphate during sucrose- gradient centrifugation. The inhibition of DNA-cellulose binding by pyridoxal 5′-phosphate can be reversed. Probably, pyridoxal 5′- phosphate forms a Schiff base with a critical lysine group of the eedysteroid receptor, presumably at its DNA- binding site. The hormone-receptor complexes obtained after removal of pyridoxal 5′-phosphate had the same affinity for DNA-cellulose as ‘native’ complexes. DNA-cellulose-bound [3H]ponasterone-A complexes were efficiently eluted from DNA-celhilose with pyfidoxal 5′-phosphate in 0.1 M KCI resulting in a 104-fold purification of the ecdysteroid receptor. The results reflect possible structural similarities between ecdysteroid and vertebrate steroid receptors. [ABSTRACT FROM AUTHOR]

A juvenile-hormone-binding protein (JHBP) has been isolated from Galleria mellonella hemolymph by gel filtration, phosphocellulose chromatography, and by chromatofocusing. The isolated protein is homogenous as judged by column chromatography and gel electrophoresis in the presence and absence of denaturing agent. It has a relative molecular mass of 32 000, Stokes radius 2.4 nm, sedimentation coefficient of 2.3 S, molar absorption coefficient at 280 nm ε = 2.34 x 104 M-1 cm-1, and is composed of a single polypeptide chain. Chromatofocusing analysis (pI 8.6) and isoelectric focusing (pI 8.1) indicate that the JHBP is an alkaline protein. Its amino acid composition and fluorescence absorption spectra indicate that the protein does not contain tryptophan residues. The protein exhibits one class of binding sites for juvenile hormone (JH), 0.8 per molecule, with the following dissociation constants: JH I, 8.5 × 10-8 M; JH II, 7.2 x 10-8 M; JH III, 47 x 10-8 M. The JHBP binds (10R, 11S)-JH II enantiomer with 2.3-times higher affinity then (10S, l lR)-JH II enantiomer. The pH optimum of binding is 7.0. [ABSTRACT FROM AUTHOR]

Juvenile hormone-binding protein (JHBP) isolated from Galleria mellonella haemolymph has been crystallized using the hanging-drop method in two polymorphic forms. The best diffracting crystals (2.7 Å) are trigonal, space group P3[sub 1]21 (or P3[sub 2]21), with unit-cell parameters a = 110.4, c = 93.9 Å. X-ray diffraction data have been collected for the native crystals using synchrotron radiation and cryogenic conditions (100 K). [ABSTRACT FROM AUTHOR]