Id Gene regulation and function in the prosensory domains of the chicken inner ear: a link between Bmp Signaling and Atoh1

Bone morphogenetic proteins (Bmps) regulate the expression of the proneural gene Atoh1 and the generation of hair cells in the developing inner ear. The present work explored the role of Inhibitor of Differentiation genes (Id1-3) in this process. The results show that Id genes are expressed in the prosensory domains of the otic vesicle, along with Bmp4 and Bmp7. Those domains exhibit high levels of the phosphorylated form of Bmp-responding R-Smads (P-Smad1,5,8), and of Bmp-dependent Smad transcriptional activity as shown by the BRE-tk-EGFP reporter. Increased Bmp signaling induces the expression of Id1-3 along with the inhibition of Atoh1. Conversely, the Bmp antagonist Noggin or the Bmp-receptor inhibitor Dorsomorphin elicit opposite effects, indicating that Bmp signaling is necessary for Id expression and Atoh1 regulation in the otocyst. The forced expression of Id3 is sufficient to reduce Atoh1 expression and to prevent the expression of hair cell differentiation markers. Together, these results suggest that Ids are part of the machinery that mediates the regulation of hair cell differentiation exerted by Bmps. In agreement with that, during hair cell differentiation Bmp4 expression, P-Smad1,5,8 levels and Id expression are downregulated from hair cells. However, Ids are also downregulated from the supporting cells which contrarily to hair cells exhibit high levels of Bmp4 expression, P-Smad1,5,8, and BRE-tk-EGFP activity, suggesting that in these cells Ids escape from Bmp/Smad signaling. The differential regulation of Ids in time and space may underlie the multiple functions of Bmp signaling during sensory organ development.

VAMP-2 and cellubrevin are expressed in pancreatic beta-cells and are essential for Ca(2+)-but not for GTP gamma S-induced insulin secretion.

VAMP proteins are important components of the machinery controlling docking and/or fusion of secretory vesicles with their target membrane. We investigated the expression of VAMP proteins in pancreatic beta-cells and their implication in the exocytosis of insulin. cDNA cloning revealed that VAMP-2 and cellubrevin, but not VAMP-1, are expressed in rat pancreatic islets and that their sequence is identical to that isolated from rat brain. Pancreatic beta-cells contain secretory granules that store and secrete insulin as well as synaptic-like microvesicles carrying gamma-aminobutyric acid. After subcellular fractionation on continuous sucrose gradients, VAMP-2 and cellubrevin were found to be associated with both types of secretory vesicle. The association of VAMP-2 with insulin-containing granules was confirmed by confocal microscopy of primary cultures of rat pancreatic beta-cells. Pretreatment of streptolysin-O permeabilized insulin-secreting cells with tetanus and botulinum B neurotoxins selectively cleaved VAMP-2 and cellubrevin and abolished Ca(2+)-induced insulin release (IC50 approximately 15 nM). By contrast, the pretreatment with tetanus and botulinum B neurotoxins did not prevent GTP gamma S-stimulated insulin secretion. Taken together, our results show that pancreatic beta-cells express VAMP-2 and cellubrevin and that one or both of these proteins selectively control Ca(2+)-mediated insulin secretion.

Context-dependent dual role of SKI8 homologs in mRNA synthesis and turnover.

Eukaryotic mRNA transcription and turnover is controlled by an enzymatic machinery that includes RNA polymerase II and the 3' to 5' exosome. The activity of these protein complexes is modulated by additional factors, such as the nuclear RNA polymerase II-associated factor 1 (Paf1c) and the cytoplasmic Superkiller (SKI) complex, respectively. Their components are conserved across uni- as well as multi-cellular organisms, including yeast, Arabidopsis, and humans. Among them, SKI8 displays multiple facets on top of its cytoplasmic role in the SKI complex. For instance, nuclear yeast ScSKI8 has an additional function in meiotic recombination, whereas nuclear human hSKI8 (unlike ScSKI8) associates with Paf1c. The Arabidopsis SKI8 homolog VERNALIZATION INDEPENDENT 3 (VIP3) has been found in Paf1c as well; however, whether it also has a role in the SKI complex remains obscure so far. We found that transgenic VIP3-GFP, which complements a novel vip3 mutant allele, localizes to both nucleus and cytoplasm. Consistently, biochemical analyses suggest that VIP3-GFP associates with the SKI complex. A role of VIP3 in the turnover of nuclear encoded mRNAs is supported by random-primed RNA sequencing of wild-type and vip3 seedlings, which indicates mRNA stabilization in vip3. Another SKI subunit homolog mutant, ski2, displays a dwarf phenotype similar to vip3. However, unlike vip3, it displays neither early flowering nor flower development phenotypes, suggesting that the latter reflect VIP3's role in Paf1c. Surprisingly then, transgenic ScSKI8 rescued all aspects of the vip3 phenotype, suggesting that the dual role of SKI8 depends on species-specific cellular context.

Shelf life and sensory assessment of tilapia quenelle during frozen storage

The objective of this work was to evaluate the shelf life and sensory attributes of tilapia quenelle. Treatments consisted of two types of packages - polyethylene zipper (retort pouch) (QA) and polyethylene waxed paper box (QB) - stored at -18ºC for 120 days. Tilapia quenelle was stable for all parameters established by Brazilian legislation. Average values of the evaluated attributes in different packages, during storage, showed no significant difference, except for the "refrigeration" flavor. However, during the storage period, there were significant differences for sensory attributes, as "moist appearance", fish and product aroma, and off flavors of "mud" and "refrigeration". Preserving product quality as for its sensory attributes, during storage, shows that tilapia quenelle is a convenience product and contributes to the increase of fish consumption.

Market Area Selection of aSawmill Machinery Manufacturer

Tämän diplomityön tavoitteena on kartoittaa suomalaisen sahakonevalmistaja Veisto Oy:n kannalta lähitulevaisuuden merkittävimmät markkina-alueet, joiden sahateollisuuteen tehdään lähivuosina eniten korkean teknologian investointeja. Markkina-alueiden valinnassa sovelletaan sekä numeerisiin tilastoihin että asiantuntijahaastatteluihin pohjautuvia ranking-menetelmiä. Työn ensimmäinen osa käsittelee kansainvälisten teollisten markkinoiden ominaispiirteitä ja niiden analysointia. Pääpaino on kuitenkin screening-menetelmillä, markkina-alueiden vertailumenetelmilläja päätöksenteon työkaluilla. Työn toisessa osassa keskitytään markkina-alueiden screeningiin, analysointiin ja maiden eri ominaisuuksien vertailuun. Päätöksentekomatriiseja hyödyntäen valitaan Veisto Oy:lle kolme tällä hetkellä houkuttelevinta markkina-aluetta, joita ovat Venäjä, USA:n kaakkoisosan Southern Yellow Pine -alue sekä Etelä-Amerikan suurimmat sahaajamaat (Brasilia, Argentiina ja Chile) yhtenä alueena. Valituilla alueilla on Veiston kannalta omat haasteensa: USA:ssa vahvat kotimaiset kilpailijat ja uusien referenssien saaminen, Venäjällä investointien epävarmuus ja markkina-alueen laajuuden tuoma monimuotoisuus sekä Etelä-Amerikassa vahvat ruotsalaiset kilpailijat sekä etenkin Brasilian osalta tuntuvat suojatullit.

Review. Precision viticulture. Research topics, challenges and opportunities in site-specific vineyard management

Precision Viticulture (PV) is a concept that is beginning to have an impact on the wine-growing sector. Its practical implementation is dependant on various technological developments: crop sensors and yield monitors, local and remote sensors, Global Positioning Systems (GPS), VRA (Variable-Rate Application) equipment and machinery, Geographic Information Systems (GIS) and systems for data analysis and interpretation. This paper reviews a number of research lines related to PV. These areas of research have focused on four very specific fields: 1) quantification and evaluation of within-field variability, 2) delineation of zones of differential treatment at parcel level, based on the analysis and interpretation of this variability, 3) development of Variable-Rate Technologies (VRT) and, finally, 4) evaluation of the opportunities for site-specific vineyard management. Research in these fields should allow winegrowers and enologists to know and understand why yield variability exists within the same parcel, what the causes of this variability are, how the yield and its quality are interrelated and, if spatial variability exists, whether site-specific vineyard management is justifiable on a technical and economic basis.

EndoG links Bnip3-induced mitochondrial damage and caspase-independent DNA fragmentation in ischemic cardiomyocytes

Mitochondrial dysfunction, caspase activation and caspase-dependent DNA fragmentation are involved in cell damage in many tissues. However, differentiated cardiomyocytes repress the expression of the canonical apoptotic pathway and their death during ischemia is caspase-independent. The atypical BH3-only protein Bnip3 is involved in the process leading to caspase-independent DNA fragmentation in cardiomyocytes. However, the pathway by which DNA degradation ensues following Bnip3 activation is not resolved. To identify the mechanism involved, we analyzed the interdependence of Bnip3, Nix and EndoG in mitochondrial damage and DNA fragmentation during experimental ischemia in neonatal rat ventricular cardiomyocytes. Our results show that the expression of EndoG and Bnip3 increases in the heart throughout development, while the caspase-dependent machinery is silenced. TUNEL-positive DNA damage, which depends on caspase activity in other cells, is caspase-independent in ischemic cardiomyocytes and ischemia-induced DNA high and low molecular weight fragmentation is blocked by repressing EndoG expression. Ischemia-induced EndoG translocation and DNA degradation are prevented by silencing the expression of Bnip3, but not Nix, or by overexpressing Bcl-xL. These data establish a link between Bnip3 and EndoG-dependent, TUNEL-positive, DNA fragmentation in ischemic cardiomyocytes in the absence of caspases, defining an alternative cell death pathway in postmitotic cells.

Design and implementation of FPGA-based LQ control of active magnetic bearings

The need for high performance, high precision, and energy saving in rotating machinery demands an alternative solution to traditional bearings. Because of the contactless operation principle, the rotating machines employing active magnetic bearings (AMBs) provide many advantages over the traditional ones. The advantages such as contamination-free operation, low maintenance costs, high rotational speeds, low parasitic losses, programmable stiffness and damping, and vibration insulation come at expense of high cost, and complex technical solution. All these properties make the use of AMBs appropriate primarily for specific and highly demanding applications. High performance and high precision control requires model-based control methods and accurate models of the flexible rotor. In turn, complex models lead to high-order controllers and feature considerable computational burden. Fortunately, in the last few years the advancements in signal processing devices provide new perspective on the real-time control of AMBs. The design and the real-time digital implementation of the high-order LQ controllers, which focus on fast execution times, are the subjects of this work. In particular, the control design and implementation in the field programmable gate array (FPGA) circuits are investigated. The optimal design is guided by the physical constraints of the system for selecting the optimal weighting matrices. The plant model is complemented by augmenting appropriate disturbance models. The compensation of the force-field nonlinearities is proposed for decreasing the uncertainty of the actuator. A disturbance-observer-based unbalance compensation for canceling the magnetic force vibrations or vibrations in the measured positions is presented. The theoretical studies are verified by the practical experiments utilizing a custom-built laboratory test rig. The test rig uses a prototyping control platform developed in the scope of this work. To sum up, the work makes a step in the direction of an embedded single-chip FPGA-based controller of AMBs.

Evolution and cellular function of monothiol glutaredoxins: involvement in iron-sulphur cluster assembly

A number of bacterial species, mostly proteobacteria, possess monothiol glutaredoxins homologous to the Saccharomyces cerevisiae mitochondrial protein Grx5, which is involved in iron–sulphur cluster synthesis. Phylogenetic profiling is used to predict that bacterial monothiol glutaredoxins also participate in the iron–sulphur cluster (ISC) assembly machinery, because their phylogenetic profiles are similar to the profiles of the bacterial homologues of yeast ISC proteins. High evolutionary cooccurrence is observed between the Grx5 homologues and the homologues of the Yah1 ferredoxin, the scaffold proteins Isa1 and Isa2, the frataxin protein Yfh1 and the Nfu1 protein. This suggests that a specific functional interaction exists between these ISC machinery proteins. Physical interaction analyses using low-definition protein docking predict the formation of strong and specific complexes between Grx5 and several components of the yeast ISC machinery. Two-hybrid analysis has confirmed the in vivo interaction between Grx5 and Isa1. Sequence comparison techniques and cladistics indicate that the other two monothiol glutaredoxins of S. cerevisiae, Grx3 and Grx4, have evolved from the fusion of a thioredoxin gene with a monothiol glutaredoxin gene early in the eukaryotic lineage, leading to differential functional specialization. While bacteria do not contain these chimaeric glutaredoxins, in many eukaryotic species Grx5 and Grx3/4-type monothiol glutaredoxins coexist in the cell.

Characterization and occurence of eairly softening disorder in 'golden' papaya fruits

The occurrence of green skin and soft pulp in 'Golden' papaya fruit during certain seasons has been reported by farmers in the northern of the state of Espirito Santo, Brazil. The objective of this study was to characterize and determine the occurrence of this disorder, which was referred as "early softening disorder". Fruits were harvested weekly for 11 months (from September to July). The fruits were stored at 10°C, and then fruit flesh firmness and skin color were analyzed. The results of the firmness test were submitted to regression analysis assuming a linear trendline. The slope of the curve was called the 'softening index' (SI). Fruits with early softening are characterized by a loss of firmness in less than 10 days, even when stored under refrigeration. Although softened, the skin of the fruit remains partially green. Fruits with the disorder occurred more frequently from mid-summer to mid-autumn (February to May). It is not possible to distinguish early softening disorder fruits from those without the disorder by skin color and flesh firmness analysis at the time of the harvest.

Monitoring the viability of citrus rootstocks seeds stored under refrigeration

The citrus nursery tree is produced through the bud grafting process, in which rootstock is usually grown from seed germination. The objective of this research was to evaluate, in two dissimilar environmental conditions, the viability and polyembryony expression of five citrus rootstocks seeds stored in different periods under refrigeration. The rootstock varieties evaluated were: Rangpur lime (Citrus limonia Osb. cv. Limeira), Trifoliate orange (Poncirus trifoliata Raf. cv. Limeira), Citrumelo (P. trifoliata x C. paradisi Macf. cv. Swingle), Sunki mandarin (C. sunki Hort. ex Tanaka) and Volkamer lemon (C. volkameriana Ten. & Pasq. cv. Catania 2). The experimental design was the randomized blocks in a 11 x 5 x 2 factorial scheme, evaluating from time zero to the tenth month of storage, the five varieties of rootstock in two environments: germination and growth B.O.D type chamber (Biological Oxygen Demand - Eletrolab Brand Model FC 122) at 25 °C; and greenhouse seedbed with partial temperature control (22 °C to 36 °C) and humidity control (75-85%). The plot had 24 seeds in four replicates, using trays with substrate in greenhouse and Petri dishes with filter paper in B.O.D. chamber. The seed germination rate and polyembryony expression were evaluated monthly. It was concluded that Trifoliate and Citrumelo Swingle seeds can be stored for up to seven months, while Volkamer lemon, Rangpur lime and Sunki seeds can be stored for up to ten months. The polyembryony expression rate was slightly higher when measured in greenhouse than in B.O.D. chamber and remained stable in both environments until the seventh month, from which dropped sharply. Citrumelo Swingle seeds expressed the highest polyembryony rate (18.8%), followed by Rangpur lime and Volkamer lemon (average value of 13.7%), Sunki (9.4%) and Trifoliate (3.2%). Despite some differences among varieties, the viability of rootstock stored seeds can be monitored either in the greenhouse or in B.O.D. germination chamber, the latter being the faster and more suitable method.

Moderately oxidized oils and dietary zinc and alpha-tocopheryl acetate supplementation: effects on the oxidative stability of rabbit plasma, liver, and meat

The aim of this study was to assess the alterations in plasma, liver, and meat oxidative stability and R-tocopherol content when moderately oxidized sunflower oils were added to feeds and when feeds were supplemented with R-tocopheryl acetate (100 mg/kg) and Zn (200 mg/kg). The effects of cooking the meat and its subsequent refrigeration were also studied. When the content of primary oxidation compounds of the oil was high, rabbit plasma, liver, and meat R-tocopherol content was reduced and meat susceptibility to oxidation increased. The addition of oil with a high content of secondary oxidation compounds (oil heated at 140 'C, 31 h) to feed also led to an increase in meat susceptibi- lity to oxidation, although it presented an R-tocopherol content similar to that of nonheated oil. Feed supplementation with R-tocopheryl acetate increased tissue R-tocopherol content and improved the oxidative stability of liver and meat. However, in the latter, it was less effective when oil heated at 55 'C was added.

Potential Acquisition Candidates for a Supplier of Wood Processing Machinery

Diplomityön tavoitteena oli sopivimman yritysostokohteen valitseminen useiden kilpailijoiden joukosta puunkäsittelykoneiden toimittajalle. Ensin esiteltiin Suomen metsäteollisuus sekä sen osaamistarpeista noussut metsäklusteri pääosin kohdeyrityksen näkökulmasta. Seuraavaksi annettiin kuva yrityksen tuotteista, kilpailijoista ja asiakkaista. Yritysostoprosessi kuvattiin sekä esille tuotiin yleiset motiivit ja kriittiset menestystekijät. Lisäksi kuvattiin kilpailijoiden ja liiketoimintaympäristön analysointi yrityksen menestyksen edellytyksenä. Puuntyöstökoneiden markkinat segmentoitiin ja analysoitiin vuodesta 1990 aina tähän päivään asti, jotta löydettäisiin kehityskelpoiset osa-alueet eli alueet, joissa yrityksen markkinaosuutta voitaisiin kasvattaa. Kandidaattien ominaisuuksia verrattiin yritysoston motiiveihin. Yritysten tuotteet sekä maantieteellinen sijainti pisteytettiin, jotta sopivimmat yritykset nousisivat esille. Kolme yritystä valittiin syvällisempään tarkasteluun. Yritysten tuotteita, taloudellista asemaa ja globaalia verkostoa vertailtiin keskenään muiden tekijöiden, kuten maailmantalouden ohessa. Taloudellisesti vakaa ja teknisesti monipuolinen yritys kohtasi yritysoston motiivit parhaiten. Kohteen positiivisia puolia olivat sijainti, tuotteet ja palvelut. Lisäksi, yritys sopii ostajan strategiaan sekä auttaa kohtaamaan asiakkaiden nykyiset ja tulevat tarpeet.

Development of support and drive systems for defiberization drum

Diplomityön tarkoituksena oli kuidutusrumpulaitteiston käytön- ja kannatuksen kehittä-minen. Työ rajattiin laajuutensa vuoksi koskemaan tuotesarjan viittä pienintä kokoa. Työn alkuosassa käsitellään kuidutuksen teoriaa ja siihen soveltuvia laitteistoja. Käytön suunnittelun kannalta olennaista käynnistystehon tarvetta on tarkasteltu lähtökohdaisesti fysiikan avulla. Perustietoja teorialle on haettu aiemmista tutkimuksista sekä kirjallisuu-desta. Tarkastelun tuloksena teoriaa on kehitty ja se on saatu vastaamaan todellisuutta aiempaa paremmin. Kannatuksen ja käytön toteuttamisvaihtoja etsittäessä on käytetty systemaattisen koneen-suunnittelun keinoja. Saatuja ideoita on arvioitu teknillis-taloudellisin perustein ja näistä on valittu parhaat vaihtoehdot jatkokehitykseen. Jatkokehitysvaiheessa ratkaisuvaihto-ehtoja on tarkasteltu komponenttitasolla ja näistä on tehty yksityiskohtaiset kustannus-laskelmat. Työn tuloksena on esitetty kannatuksen ja käytön toteutusvaihtoehto, jonka avulla voidaan saavuttaa merkittäviä kustannussäästöjä. Korkea, 30 prosentin kustannussäästö-tavoite saavutettiin.

Combined loss of the BH3-only proteins Bim and Bmf restores B-cell development and function in TACI-Ig transgenic mice.

Terminal differentiation of B cells depends on two interconnected survival pathways, elicited by the B-cell receptor (BCR) and the BAFF receptor (BAFF-R), respectively. Loss of either signaling pathway arrests B-cell development. Although BCR-dependent survival depends mainly on the activation of the v-AKT murine thymoma viral oncogene homolog 1 (AKT)/PI3-kinase network, BAFF/BAFF-R-mediated survival engages non-canonical NF-κB signaling as well as MAPK/extracellular-signal regulated kinase and AKT/PI3-kinase modules to allow proper B-cell development. Plasma cell survival, however, is independent of BAFF-R and regulated by APRIL that signals NF-κB activation via alternative receptors, that is, transmembrane activator and CAML interactor (TACI) or B-cell maturation (BCMA). All these complex signaling events are believed to secure survival by increased expression of anti-apoptotic B-cell lymphoma 2 (Bcl2) family proteins in developing and mature B cells. Curiously, how lack of BAFF- or APRIL-mediated signaling triggers B-cell apoptosis remains largely unexplored. Here, we show that two pro-apoptotic members of the 'Bcl2 homology domain 3-only' subgroup of the Bcl2 family, Bcl2 interacting mediator of cell death (Bim) and Bcl2 modifying factor (Bmf), mediate apoptosis in the context of TACI-Ig overexpression that effectively neutralizes BAFF as well as APRIL. Surprisingly, although Bcl2 overexpression triggers B-cell hyperplasia exceeding the one observed in Bim(-/-)Bmf(-/-) mice, Bcl2 transgenic B cells remain susceptible to the effects of TACI-Ig expression in vivo, leading to ameliorated pathology in Vav-Bcl2 transgenic mice. Together, our findings shed new light on the molecular machinery restricting B-cell survival during development, normal homeostasis and under pathological conditions. Our data further suggest that Bcl2 antagonists might improve the potency of BAFF/APRIL-depletion strategies in B-cell-driven pathologies.