Rhythmic growth explained by coincidence between internal and external cues.

Most organisms use circadian oscillators to coordinate physiological and developmental processes such as growth with predictable daily environmental changes like sunrise and sunset. The importance of such coordination is highlighted by studies showing that circadian dysfunction causes reduced fitness in bacteria and plants, as well as sleep and psychological disorders in humans. Plant cell growth requires energy and water-factors that oscillate owing to diurnal environmental changes. Indeed, two important factors controlling stem growth are the internal circadian oscillator and external light levels. However, most circadian studies have been performed in constant conditions, precluding mechanistic study of interactions between the clock and diurnal variation in the environment. Studies of stem elongation in diurnal conditions have revealed complex growth patterns, but no mechanism has been described. Here we show that the growth phase of Arabidopsis seedlings in diurnal light conditions is shifted 8-12 h relative to plants in continuous light, and we describe a mechanism underlying this environmental response. We find that the clock regulates transcript levels of two basic helix-loop-helix genes, phytochrome-interacting factor 4 (PIF4) and PIF5, whereas light regulates their protein abundance. These genes function as positive growth regulators; the coincidence of high transcript levels (by the clock) and protein accumulation (in the dark) allows them to promote plant growth at the end of the night. Thus, these two genes integrate clock and light signalling, and their coordinated regulation explains the observed diurnal growth rhythms. This interaction may serve as a paradigm for understanding how endogenous and environmental signals cooperate to control other processes.

Auxin-mediated plant architectural changes in response to shade and high temperature.

The remarkable plasticity of their architecture allows plants to adjust growth to the environment and to overcome adverse conditions. Two examples of environmental stresses that drastically affect shoot development are imminent shade and high temperature. Plants in crowded environments and plants in elevated ambient temperature display very similar phenotypic adaptations of elongated hypocotyls in seedlings and elevated and elongated leaves at later developmental stages. The comparable growth responses to shade and high temperature are partly regulated through shared signaling pathways, of which the phytohormone auxin and the phytochrome interacting factors (PIFs) are important components. During both shade- and temperature-induced elongation growth auxin biosynthesis and signaling are upregulated in a PIF-dependent manner. In this review we will discuss recent progress in our understanding of how auxin mediates architectural adaptations to shade and high temperature.

Phosphorylation by casein kinase 2 facilitates rRNA gene transcription by promoting dissociation of TIF-IA from elongating RNA polymerase I.

The protein kinase casein kinase 2 (CK2) phosphorylates different components of the RNA polymerase I (Pol I) transcription machinery and exerts a positive effect on rRNA gene (rDNA) transcription. Here we show that CK2 phosphorylates the transcription initiation factor TIF-IA at serines 170 and 172 (Ser170/172), and this phosphorylation triggers the release of TIF-IA from Pol I after transcription initiation. Inhibition of Ser170/172 phosphorylation or covalent tethering of TIF-IA to the RPA43 subunit of Pol I inhibits rDNA transcription, leading to perturbation of nucleolar structure and cell cycle arrest. Fluorescence recovery after photobleaching and chromatin immunoprecipitation experiments demonstrate that dissociation of TIF-IA from Pol I is a prerequisite for proper transcription elongation. In support of phosphorylation of TIF-IA switching from the initiation into the elongation phase, dephosphorylation of Ser170/172 by FCP1 facilitates the reassociation of TIF-IA with Pol I, allowing a new round of rDNA transcription. The results reveal a mechanism by which the functional interplay between CK2 and FCP1 sustains multiple rounds of Pol I transcription.

Multi-scale systems analysis of plant development : beyond cellular signaling and tissue morphodynamics

Les plantes sont essentielles pour les sociétés humaines. Notre alimentation quotidienne, les matériaux de constructions et les sources énergétiques dérivent de la biomasse végétale. En revanche, la compréhension des multiples aspects développementaux des plantes est encore peu exploitée et représente un sujet de recherche majeur pour la science. L'émergence des technologies à haut débit pour le séquençage de génome à grande échelle ou l'imagerie de haute résolution permet à présent de produire des quantités énormes d'information. L'analyse informatique est une façon d'intégrer ces données et de réduire la complexité apparente vers une échelle d'abstraction appropriée, dont la finalité est de fournir des perspectives de recherches ciblées. Ceci représente la raison première de cette thèse. En d'autres termes, nous appliquons des méthodes descriptives et prédictives combinées à des simulations numériques afin d'apporter des solutions originales à des problèmes relatifs à la morphogénèse à l'échelle de la cellule et de l'organe. Nous nous sommes fixés parmi les objectifs principaux de cette thèse d'élucider de quelle manière l'interaction croisée des phytohormones auxine et brassinosteroïdes (BRs) détermine la croissance de la cellule dans la racine du méristème apical d'Arabidopsis thaliana, l'organisme modèle de référence pour les études moléculaires en plantes. Pour reconstruire le réseau de signalement cellulaire, nous avons extrait de la littérature les informations pertinentes concernant les relations entre les protéines impliquées dans la transduction des signaux hormonaux. Le réseau a ensuite été modélisé en utilisant un formalisme logique et qualitatif pour pallier l'absence de données quantitatives. Tout d'abord, Les résultats ont permis de confirmer que l'auxine et les BRs agissent en synergie pour contrôler la croissance de la cellule, puis, d'expliquer des observations phénotypiques paradoxales et au final, de mettre à jour une interaction clef entre deux protéines dans la maintenance du méristème de la racine. Une étude ultérieure chez la plante modèle Brachypodium dystachion (Brachypo- dium) a révélé l'ajustement du réseau d'interaction croisée entre auxine et éthylène par rapport à Arabidopsis. Chez ce dernier, interférer avec la biosynthèse de l'auxine mène à la formation d'une racine courte. Néanmoins, nous avons isolé chez Brachypodium un mutant hypomorphique dans la biosynthèse de l'auxine qui affiche une racine plus longue. Nous avons alors conduit une analyse morphométrique qui a confirmé que des cellules plus anisotropique (plus fines et longues) sont à l'origine de ce phénotype racinaire. Des analyses plus approfondies ont démontré que la différence phénotypique entre Brachypodium et Arabidopsis s'explique par une inversion de la fonction régulatrice dans la relation entre le réseau de signalisation par l'éthylène et la biosynthèse de l'auxine. L'analyse morphométrique utilisée dans l'étude précédente exploite le pipeline de traitement d'image de notre méthode d'histologie quantitative. Pendant la croissance secondaire, la symétrie bilatérale de l'hypocotyle est remplacée par une symétrie radiale et une organisation concentrique des tissus constitutifs. Ces tissus sont initialement composés d'une douzaine de cellules mais peuvent aisément atteindre des dizaines de milliers dans les derniers stades du développement. Cette échelle dépasse largement le seuil d'investigation par les moyens dits 'traditionnels' comme l'imagerie directe de tissus en profondeur. L'étude de ce système pendant cette phase de développement ne peut se faire qu'en réalisant des coupes fines de l'organe, ce qui empêche une compréhension des phénomènes cellulaires dynamiques sous-jacents. Nous y avons remédié en proposant une stratégie originale nommée, histologie quantitative. De fait, nous avons extrait l'information contenue dans des images de très haute résolution de sections transverses d'hypocotyles en utilisant un pipeline d'analyse et de segmentation d'image à grande échelle. Nous l'avons ensuite combiné avec un algorithme de reconnaissance automatique des cellules. Cet outil nous a permis de réaliser une description quantitative de la progression de la croissance secondaire révélant des schémas développementales non-apparents avec une inspection visuelle classique. La formation de pôle de phloèmes en structure répétée et espacée entre eux d'une longueur constante illustre les bénéfices de notre approche. Par ailleurs, l'exploitation approfondie de ces résultats a montré un changement de croissance anisotropique des cellules du cambium et du phloème qui semble en phase avec l'expansion du xylème. Combinant des outils génétiques et de la modélisation biomécanique, nous avons démontré que seule la croissance plus rapide des tissus internes peut produire une réorientation de l'axe de croissance anisotropique des tissus périphériques. Cette prédiction a été confirmée par le calcul du ratio des taux de croissance du xylème et du phloème au cours de développement secondaire ; des ratios élevés sont effectivement observés et concomitant à l'établissement progressif et tangentiel du cambium. Ces résultats suggèrent un mécanisme d'auto-organisation établi par un gradient de division méristématique qui génèrent une distribution de contraintes mécaniques. Ceci réoriente la croissance anisotropique des tissus périphériques pour supporter la croissance secondaire. - Plants are essential for human society, because our daily food, construction materials and sustainable energy are derived from plant biomass. Yet, despite this importance, the multiple developmental aspects of plants are still poorly understood and represent a major challenge for science. With the emergence of high throughput devices for genome sequencing and high-resolution imaging, data has never been so easy to collect, generating huge amounts of information. Computational analysis is one way to integrate those data and to decrease the apparent complexity towards an appropriate scale of abstraction with the aim to eventually provide new answers and direct further research perspectives. This is the motivation behind this thesis work, i.e. the application of descriptive and predictive analytics combined with computational modeling to answer problems that revolve around morphogenesis at the subcellular and organ scale. One of the goals of this thesis is to elucidate how the auxin-brassinosteroid phytohormone interaction determines the cell growth in the root apical meristem of Arabidopsis thaliana (Arabidopsis), the plant model of reference for molecular studies. The pertinent information about signaling protein relationships was obtained through the literature to reconstruct the entire hormonal crosstalk. Due to a lack of quantitative information, we employed a qualitative modeling formalism. This work permitted to confirm the synergistic effect of the hormonal crosstalk on cell elongation, to explain some of our paradoxical mutant phenotypes and to predict a novel interaction between the BREVIS RADIX (BRX) protein and the transcription factor MONOPTEROS (MP),which turned out to be critical for the maintenance of the root meristem. On the same subcellular scale, another study in the monocot model Brachypodium dystachion (Brachypodium) revealed an alternative wiring of auxin-ethylene crosstalk as compared to Arabidopsis. In the latter, increasing interference with auxin biosynthesis results in progressively shorter roots. By contrast, a hypomorphic Brachypodium mutant isolated in this study in an enzyme of the auxin biosynthesis pathway displayed a dramatically longer seminal root. Our morphometric analysis confirmed that more anisotropic cells (thinner and longer) are principally responsible for the mutant root phenotype. Further characterization pointed towards an inverted regulatory logic in the relation between ethylene signaling and auxin biosynthesis in Brachypodium as compared to Arabidopsis, which explains the phenotypic discrepancy. Finally, the morphometric analysis of hypocotyl secondary growth that we applied in this study was performed with the image-processing pipeline of our quantitative histology method. During its secondary growth, the hypocotyl reorganizes its primary bilateral symmetry to a radial symmetry of highly specialized tissues comprising several thousand cells, starting with a few dozens. However, such a scale only permits observations in thin cross-sections, severely hampering a comprehensive analysis of the morphodynamics involved. Our quantitative histology strategy overcomes this limitation. We acquired hypocotyl cross-sections from tiled high-resolution images and extracted their information content using custom high-throughput image processing and segmentation. Coupled with an automated cell type recognition algorithm, it allows precise quantitative characterization of vascular development and reveals developmental patterns that were not evident from visual inspection, for example the steady interspace distance of the phloem poles. Further analyses indicated a change in growth anisotropy of cambial and phloem cells, which appeared in phase with the expansion of xylem. Combining genetic tools and computational modeling, we showed that the reorientation of growth anisotropy axis of peripheral tissue layers only occurs when the growth rate of central tissue is higher than the peripheral one. This was confirmed by the calculation of the ratio of the growth rate xylem to phloem throughout secondary growth. High ratios are indeed observed and concomitant with the homogenization of cambium anisotropy. These results suggest a self-organization mechanism, promoted by a gradient of division in the cambium that generates a pattern of mechanical constraints. This, in turn, reorients the growth anisotropy of peripheral tissues to sustain the secondary growth.

Inhibition of the shade avoidance response by formation of non-DNA binding bHLH heterodimers.

In shade-intolerant plants such as Arabidopsis, a reduction in the red/far-red (R/FR) ratio, indicative of competition from other plants, triggers a suite of responses known as the shade avoidance syndrome (SAS). The phytochrome photoreceptors measure the R/FR ratio and control the SAS. The phytochrome-interacting factors 4 and 5 (PIF4 and PIF5) are stabilized in the shade and are required for a full SAS, whereas the related bHLH factor HFR1 (long hypocotyl in FR light) is transcriptionally induced by shade and inhibits this response. Here we show that HFR1 interacts with PIF4 and PIF5 and limits their capacity to induce the expression of shade marker genes and to promote elongation growth. HFR1 directly inhibits these PIFs by forming non-DNA-binding heterodimers with PIF4 and PIF5. Our data indicate that PIF4 and PIF5 promote SAS by directly binding to G-boxes present in the promoter of shade marker genes, but their action is limited later in the shade when HFR1 accumulates and forms non-DNA-binding heterodimers. This negative feedback loop is important to limit the response of plants to shade.

Microclimate and development of 'Conilon' coffee intercropped with rubber trees

The objective of this work was to evaluate the influence of intercropping 'Conilon' coffee (Coffea canephora) with rubber trees on coffee tree microclimate, nutrition, growth, and yield. Rubber trees were planted in two double rows 33 m apart, with 4x2.3 m spacing between plants. Treatments consisted of the distances from the coffee plants to the rubber trees: 3, 6, 9, 12, and 15 m. Measurements of atmospheric variables (temperature, irradiance, and relative humidity), leaf nutrient concentration, internode length of plagiotropic and orthotropic branches, individual leaf area, chlorophyll content, and yield were performed. Intercropping promotes changes in the microclimatic conditions of coffee plants close to rubber trees, with reduction of temperature and irradiance level and increase in air relative humidity. The proximity of the coffee tree to the rubber trees promotes the elongation of the plagiotropic and orthotropic branches and increases the individual leaf area; however, it does not affect leaf concentrations of N, K, Mg, Fe, Zn, and B in 'Conilon' coffee and does not have a negative impact on yield.

Prohexadione calcium on the growth and quality of eggplant seedlings

Abstract:The objective of this work was to evaluate the effects of prohexadione calcium concentrations on the growth and quality of eggplant (Solanum melongena) seedlings. The effects of prohexadione calcium concentrations of 0, 50, 100, or 150 mg L-1 on seedling growth parameters were evaluated in a greenhouse experiment. After the greenhouse experiment, the seedlings were transplanted to the field. During the field experiment, the number of days to flowering, plant height, number of fruits per plant, average fruit weight and yield were evaluated. Both experiments were carried out in a completely randomized design with four replicates. All prohexadione calcium concentrations significantly reduced shoot height and internode length, when compared to the control. The concentrations of 50, 100, and 150 mg L-1 prohexadione calcium reduced shoot height by 27, 32, and 38%, respectively. Prohexadione calcium treatments (except the one with 50 mg L-1) enhanced relative chlorophyll content of leaves in comparison to the control. There were no delays in flowering and no significant differences in number of fruits per plant among treatments with prohexadione calcium. The concentrations of 100 and 150 mg L-1 prohexadione calcium significantly reduced yield per plant and total fruit yield, whereas the concentration of 50 mg L-1 did not cause any change in yield compared to the control. The lowest prohexadione calcium concentration can be used to control excessive elongation of eggplant seedlings without yield loss.

Mangaaniseosteisen duplex-teräksen hitsattavuus

Diplomityön tavoitteena oli selvittää mangaaniseosteisen LDX2101 duplex-teräksen ja LDX2101 hitsauslisäaineiden hitsausparametrit puikkohitsaus-, MAG-täytelankahitsaus- ja plasmahitsausprosessilla. Toisena tavoitteena oli selvittää lämmöntuonnin vaikutuksia hit-sausliitosten mekaanisiin ominaisuuksiin, iskusitkeyteen (-40 °C) ja ferriitti- / austeniittipitoi-suuksien jakaantumiseen. Tutkittavat hitsit olivat päittäishitsejä ja koelevyjen aineenpaksuus oli 6 mm. Koetulosten perusteella tehtiin hitsausohjeet kyseisille hitsausprosesseille. Työn kirjallisessa osassa on selvitetty yleisesti duplex-terästen käyttökohteita, mekaanisia ja kemiallisia ominaisuuksia sekä duplex-terästen korroosio-ominaisuuksia. Lisäksi on käsitelty eri hitsausprosesseja ja duplex-terästen hitsattavuutta, jossa selvitetään mm. terästen jähmet-tymistä, erkaumia ja faaseja, lämmöntuonnin vaikutuksia ja hitsausliitosten korroosionkestä-vyyttä. Teoriaosassa on kerrottu myös hitsausliitosten tarkastuksesta ja hitsiluokista. Työn kokeellisessa osassa esitellään kokeiden suoritus sekä ainetta rikkova ja ainetta rikko-maton tarkastus. Liitoksille suoritettava koestusohjelma noudatti pääosin menetelmäkoestan-dardin SFS-EN 15614-1 ohjetta. Hitseille tehtiin ainetta rikkomaton tarkastus, joka käsittää silmämääräisen tarkastuksen, pintatarkastuksen ja röntgentarkastuksen. Rikkovaa aineen-koetusta tehtiin sisältäen kovuusmittaukset, vetokokeet, taivutuskokeet jaiskusitkeyskokeet. Lisäksi valmistettiin metallografiset hieet, joita tutkimalla selvitettiin hitsausliitoksen metal-lurgiaa ja määriteltiin ferriitti- ja austeniittipitoisuudet. Kokeiden perusteella on laadittu hit-sausohjeet kullekin hitsausprosessille. Lisäksi kokeellisessa osassa on esitelty tulokset syö-pymäkokeesta, jossa selvitettiin eri materiaalien syöpymiskestävyyttä valkolipeäliuoksessa. Tutkimustuloksien perusteella LDX2102 duplex-materiaali onhyvin hitsattavaa laajalla hit-sausparametrialueella. Mekaaniset ominaisuudet, kuten lujuus- ja iskusitkeysarvot täyttävät materiaalistandardin SFS-EN10028-7 niille asettamat vaatimukset. Taivutussitkeys ja murto-venymät jäivät kuitenkin osalla koekappaleista vaatimustasoa alhaisemmiksi. Austeniitti- ja ferriittipitoisuudet vastaavat materiaalistandardin vaatimuksia.

Kosteuden ja lämpötilan vaikutus kartonkimassojen muovattavuuteen

Työn tavoitteena oli kartoittaa yleisesti kartongin valmistukseen käytettävien massatyyppien muovattavuuspotentaali. Muovattuvuuteen eniten vaikuttava tekijänä pidettiinmassojen murtovenymää. Työssä tutkittiin kosteuden ja lämpötilan sekä vetonopeuden vaikutusta mekaanisten ja sellumassojen ominaisuuksiin laboratorio-olosuhteissa. Kirjallisuusosassa tarkastellaan paperin lasisiirtymälämpötilaa muovauksen kannalta oleellisena tekijä. Lisäksi käydään läpi tapoja, joilla murtovenymää voidaan kasvattaa. Kartongin keskikerroksen aallotuksessa ja valmistettaessa paperipohjaisia tuotteita syvävedolla, käytetään hyväksi paperin pehmenemistä lämpötilan ja kosteuden alaisena. Niiden olosuhteita tarkastellaan lyhyesti. Lisäksi luodaan katsaus lämpötilan, kosteuden ja nopeuden vaikutuksistapaperin mekaanisiin ominaisuuksiin. Työn kokeellisessa osassa massojen mekaanisia ominaisuuksia testattiin erilaisissa kosteuspitoisuuksissasekä lämpötiloissa eri vetonopeuksilla. Kokeiden perusteella pyrittiin löytämään olosuhteet kullekin massalle, joissa niillä on paras murtovenymä. Lisäksi selvitettiin kuidun kihartamisen, sellumassojen jauhatuksen sekä arkin vapaan kuivumisen vaikutusta murtovenymään. Erilaiset massat saavuttavat parhaan murtovenymän erilaisissa kosteus- ja lämpötilaolosuhteissa. Sellumassoilla paras murtovenymä saadaan huoneenlämpötilassa ja paperin kosteuspitoisuuden ollessa välillä 11...12%. CTMP-massojen paras murtovenymä saadaan vedellä kyllästetyllä paperilla kohotetussa lämpötilassa. Lämpötila riippuu CTMP-massan valmistustavasta ja raaka-aineesta. Sellumassoilla on parempi kokonaisvenymä, kuin ligniinipitoisilla massoilla. Vetonopeuden vaikutus murtovenymään riippuu massasta sekä kosteudesta. Alhaisessa kosteudessa suurempi vetonopeus antaa aina pienemmän venymän. Kosteuden noustessa riippuu massasta, miten vetonopeus vaikuttaa murtovenymään. Plastinen osa tietystä venymästä on riippumaton massasta, jauhatusasteesta ja kuivatustavasta. Ainut vaikuttava muuttuja on kosteus. Kosteuden kasvu kasvattaa plastista venymää.

Identification of genes potentially involved in solute stress response in Sphingomonas wittichii RW1 by transposon mutant recovery.

The term water stress refers to the effects of low water availability on microbial growth and physiology. Water availability has been proposed as a major constraint for the use of microorganisms in contaminated sites with the purpose of bioremediation. Sphingomonas wittichii RW1 is a bacterium capable of degrading the xenobiotic compounds dibenzofuran and dibenzo-p-dioxin, and has potential to be used for targeted bioremediation. The aim of the current work was to identify genes implicated in water stress in RW1 by means of transposon mutagenesis and mutant growth experiments. Conditions of low water potential were mimicked by adding NaCl to the growth media. Three different mutant selection or separation method were tested which, however recovered different mutants. Recovered transposon mutants with poorer growth under salt-induced water stress carried insertions in genes involved in proline and glutamate biosynthesis, and further in a gene putatively involved in aromatic compound catabolism. Transposon mutants growing poorer on medium with lowered water potential also included ones that had insertions in genes involved in more general functions such as transcriptional regulation, elongation factor, cell division protein, RNA polymerase β or an aconitase.

Networking of differentially expressed genes in human cancer cells resistant to methotrexate

BACKGROUND: The need for an integrated view of data obtained from high-throughput technologies gave rise to network analyses. These are especially useful to rationalize how external perturbations propagate through the expression of genes. To address this issue in the case of drug resistance, we constructed biological association networks of genes differentially expressed in cell lines resistant to methotrexate (MTX). METHODS: Seven cell lines representative of different types of cancer, including colon cancer (HT29 and Caco2), breast cancer (MCF-7 and MDA-MB-468), pancreatic cancer (MIA PaCa-2), erythroblastic leukemia (K562) and osteosarcoma (Saos-2), were used. The differential expression pattern between sensitive and MTX-resistant cells was determined by whole human genome microarrays and analyzed with the GeneSpring GX software package. Genes deregulated in common between the different cancer cell lines served to generate biological association networks using the Pathway Architect software. RESULTS: Dikkopf homolog-1 (DKK1) is a highly interconnected node in the network generated with genes in common between the two colon cancer cell lines, and functional validations of this target using small interfering RNAs (siRNAs) showed a chemosensitization toward MTX. Members of the UDP-glucuronosyltransferase 1A (UGT1A) family formed a network of genes differentially expressed in the two breast cancer cell lines. siRNA treatment against UGT1A also showed an increase in MTX sensitivity. Eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) was overexpressed among the pancreatic cancer, leukemia and osteosarcoma cell lines, and siRNA treatment against EEF1A1 produced a chemosensitization toward MTX. CONCLUSIONS: Biological association networks identified DKK1, UGT1As and EEF1A1 as important gene nodes in MTX-resistance. Treatments using siRNA technology against these three genes showed chemosensitization toward MTX.

Role of amphiregulin in mammary gland development

Abstract Ovarian hormones are key regulators of postnatal mammary gland development and are linked to breast carcinogenesis. In particular, estrogens induce mammary epithelial cells to proliferate at the onset of puberty, leading to the elongation of the rudimental ductal tree into the fatty stromal tissue. Elucidating the molecular events underlying estrogen mitogenic activity in the mammary gland is of value in understanding how the deregulation of this signalling pathway can lead to breast tumorigenesis. Our lab has recently shown that estrogen induces mammary proliferation via epithelial estrogen receptor alpha (ERα) by a paracrine mechanism. Based on the finding that several EGF receptor (EGFR) ligands are able to substitute for estrogens and that amphiregulin (Areg), one of these ligands, is required during mammary development, we have hypothesized that Areg is a key mediator of estrogen induced paracrine signalling during ductal morphogenesis. Our analysis of the Areg -/- mice mammary phenotype reveals that epithelial Areg is required at the onset of puberty for epithelial proliferation, terminal end bud (TEB) formation and, subsequently, ductal elongation. Hormonal stimulation experiments show that among the EGFR ligands, only Arég is specifically controlled by estrogen at the transcriptional level, via ERα, in the mammary gland. Moreover, Areg is required for the estrogen-induced mammotrophic effects of epithelial proliferation and ductal elongation. We have shown that ectopic Areg expression in ERα -/- mammary epithelial cells is sufficient to induce ductal morphogenesis. Our transplantations experiment show that when Areg -/- cells are in the presence of wt cells they contribute to all aspects of ductal development, suggesting that this growth factor acts in a paracrine fashion. Moreover, this result shows that Areg -/- epithelial cells are not intrinsically impaired in proliferation. Our transplantation experiment carried out under physiological conditions confirmed previous reports showing that stromal EGFR is needed for ductal morphogenesis. This suggests that estrogen-driven Areg signalling involves an epithelium-stroma crosstalk. Thus, these data confirmed our hypothesis of Areg being an important estrogen mediator during ductal morphogenesis. Résumé Les hormones ovariennes, régulatrices clés du développement post-natal de la glande mammaire, sont également liées à la carcinogénèse du sein. En particulier, l'oestrogène induit la division des cellules épithéliales au début de la puberté. Cette prolifération amène à l'élongation du réseau canalaire rudimentaire et permet l'invasion du compartiment stromal. L'élucidation des mécanismes moléculaires responsables de l'activité mitogénique de l'oestrogène dans la glande mammaire est précieuse pour une meilleure compréhension du développement du cancer du sein. Notre laboratoire a récemment démontré que l'cestrogène induit la prolifération des cellules épithéliales par un signal paracrine, grâce au récepteur à l'oestrogène alpha (ERα). En se basant sur le fait que plusieurs ligands du récepteur à l'EGF (EGFR) sont capables de se substituer à l'cestrogène et d'induire la prolifération épithéliale et qu'amphiregulin (Areg), un de ces ligands, est essentielle au développement de la glande mammaire, nous avons émi l'hypothèse que Areg est un médiateur essentiel du signal paracrine induit par l'oestrogène pendant la croissance du système canalaire. Nos analyses phénotypiques des glandes mammaires issues de souris transgéniques Areg -/- démontrent que cette protéine est indispensable à la prolifération des cellules épithéliales mammaires au début de la puberté et à la formation des bourgeons terminaux qui conduisent à l'élongation des canaux. Nos expériences de stimulations hormonales démontrent que, parmi l'ensemble des ligands du EGFR, seule Areg est contrôlée au niveau transcriptionnel par l'cestrogène dans la glande mammaire, ceci via le récepteur ERα. De plus, Areg est essentielle pour le effets mammotrophique induit par l'cestrogène, à savoir la prolifération épithéliál et la croissance du système canalaire. Par ailleurs, l'expression ectopique d'Areg dans des cellules epithéliales mammaires de souris transgéniques ERα -/- est suffisante pour permettre la formation du réseau canalaire. En présence de cellules normales, les cellules dépourvues du gène d'Areg contribuent à la formation des canaux. Cette expérience suggère que ce facteur de croissance agit de manière paracrine. De plus, ce résultat montre que les cellules épithéliales Areg -/- conservent leur potentiel prolifératif. Nos expériences de transplantation, réalisées dans des conditions physiologiques, ont confirmé des précédentes études qui montraient que le récepteur stromal à l'EGF est nécessaire pour la morphogénèse du système canalaire. Ceci suggère que la voie de signalisation activée par l'oestrogène et dépendante d' implique une communication entre l'épithélium et le stroma. Ainsi, ces résultats valident notre hypothèse puisqu'ils confirment Areg en tant que médiateur majeur de l'oestrogène dans la morphogénèse du système canalaire.

A novel procedure for establishing clinching parameters for high strength steel sheet

The building industry has a particular interest in using clinching as a joining method for frame constructions of light-frame housing. Normally many clinch joints are required in joining of frames.In order to maximise the strength of the complete assembly, each clinch joint must be as sound as possible. Experimental testing is the main means of optimising a particular clinch joint. This includes shear strength testing and visual observation of joint cross-sections. The manufacturers of clinching equipment normally perform such experimental trials. Finite element analysis can also be used to optimise the tool geometry and the process parameter, X, which represents the thickness of the base of the joint. However, such procedures require dedicated software, a skilled operator, and test specimens in order to verify the finite element model. In addition, when using current technology several hours' computing time may be necessary. The objective of the study was to develop a simple calculation procedure for rapidly establishing an optimum value for the parameter X for a given tool combination. It should be possible to use the procedure on a daily basis, without stringent demands on the skill of the operator or the equipment. It is also desirable that the procedure would significantly decrease thenumber of shear strength tests required for verification. The experimental workinvolved tests in order to obtain an understanding of the behaviour of the sheets during clinching. The most notable observation concerned the stage of the process in which the upper sheet was initially bent, after which the deformation mechanism changed to shearing and elongation. The amount of deformation was measured relative to the original location of the upper sheet, and characterised as the C-measure. By understanding in detail the behaviour of the upper sheet, it waspossible to estimate a bending line function for the surface of the upper sheet. A procedure was developed, which makes it possible to estimate the process parameter X for each tool combination with a fixed die. The procedure is based on equating the volume of material on the punch side with the volume of the die. Detailed information concerning the behaviour of material on the punch side is required, assuming that the volume of die does not change during the process. The procedure was applied to shear strength testing of a sample material. The sample material was continuously hot-dip zinc-coated high-strength constructional steel,with a nominal thickness of 1.0 mm. The minimum Rp0.2 proof stress was 637 N/mm2. Such material has not yet been used extensively in light-frame housing, and little has been published on clinching of the material. The performance of the material is therefore of particular interest. Companies that use clinching on a daily basis stand to gain the greatest benefit from the procedure. By understanding the behaviour of sheets in different cases, it is possible to use data at an early stage for adjusting and optimising the process. In particular, the functionality of common tools can be increased since it is possible to characterise the complete range of existing tools. The study increases and broadens the amount ofbasic information concerning the clinching process. New approaches and points of view are presented and used for generating new knowledge.

Quantitative developmental response to the length of exposure to long photoperiod in wheat and barley

The present study was designed to analyse the effect of the length of exposure to a long photoperiod imposed c. 3 weeks after sowing in spring wheat (cv. UQ189) and barley (cv. Arapiles) to (i) establish whether the response to the number of cycles of exposure is quantitative or qualitative, (ii) determine the existence of a commitment to particular stages well before the stage has been observable, and (iii) study the interrelationships between the effects on final leaf number and phyllochron when the stimulus is provided several days after seedling emergence. Both wheat and barley seemed to respond quantitatively to the number of long-day cycles they were exposed to. However, wheat showed a requirement of approximately 4 long-day cycles to be able to produce a significant response in time to heading. The barley cultivar used in the study was responsive to the minimum length of exposure. The response to extended photoperiod cycles during the stem elongation phase was due to the ‘ memory’ photoperiod effects being related, in the case of wheat, to the fact that the pre-terminal spikelet appearance phase saturated its photoperiod response well before that stage was reached. Therefore, the commitment to the terminal spikelet appearance in wheat may be reached well before this stage could be recognized. As the response in duration to heading exceeded that of the final leaf number, and the stem elongation phase responded to memory effects of photoperiod, the phyllochron of both cereals was responsive to the treatments accelerating the average phyllochron when exposed to longer periods of long days. The response in average phyllochron was due to a switch from bi-linear to linear models of leaf number v. time when the conditions were increasingly inductive, with the phyllochron of the initial (6–8) leaves being similar for all treatments (within each species), and from then on increased.

Sink limitations to yield in wheat : how could it be reduced?

Further genetic gains in wheat yield are required to match expected increases in demand. This may require the identification of physiological attributes able to produce such improvement, as well as the genetic bases controlling those traits in order to facilitate their manipulation. In the present paper, a theoretical framework of source and sink limitation to wheat yield is presented and the fine-tuning of crop development as an alternative for increasing yield potential is discussed. Following a top-down approach, most crop physiologists have agreed that the main attribute explaining past genetic gains in yield was harvest index (HI). By virtue of previous success, no further gains may be expected in HI and an alternative must be found. Using a bottom-up approach, the present paper firstly provides evidence on the generalized sink-limited condition of grain growth, determining that for further increases in yield potential, sink strength during grain filling has to be increased. The focus should be on further increasing grain number per m2, through fine-tuning pre-anthesis developmental patterns. The phase of rapid spike growth period (RSGP) is critical for grain number determination and increasing spike growth during pre-anthesis would result in an increased number of grains. This might be achieved by lengthening the duration of the phase (though without altering flowering time), as there is genotypic variation in the proportion of pre-anthesis time elapsed either before or after the onset of the stem elongation phase. Photoperiod sensitivity during RSGP could be then used as a genetic tool to further increase grain number, since slower development results in smoother floret development and more floret primordia achieve the fertile floret stage, able to produce a grain. Far less progress has been achieved on the genetic control of this attribute. None of the well-known major Ppd alleles seems to be consistently responsible for RSGP sensitivity. Alternatives for identifying the genetic factors responsible for this sensitivity (e.g. quantitative trait locus (QTL) identification in mapping populations) are being considered.