LKB1 regulates polarity remodeling and adherens junction formation in the Drosophila eye.

The serine-threonine kinase LKB1 regulates cell polarity from Caenorhabditis elegans to man. Loss of lkb1 leads to a cancer predisposition, known as Peutz-Jeghers Syndrome. Biochemical analysis indicates that LKB1 can phosphorylate and activate a family of AMPK- like kinases, however, the precise contribution of these kinases to the establishment and maintenance of cell polarity is still unclear. Recent studies propose that LKB1 acts primarily through the AMP kinase to establish and/or maintain cell polarity. To determine whether this simple model of how LKB1 regulates cell polarity has relevance to complex tissues, we examined lkb1 mutants in the Drosophila eye. We show that adherens junctions expand and apical, junctional, and basolateral domains mix in lkb1 mutants. Surprisingly, we find LKB1 does not act primarily through AMPK to regulate cell polarity in the retina. Unlike lkb1 mutants, ampk retinas do not show elongated rhabdomeres or expansion of apical and junctional markers into the basolateral domain. In addition, nutrient deprivation does not reveal a more dramatic polarity phenotype in lkb1 photoreceptors. These data suggest that AMPK is not the primary target of LKB1 during eye development. Instead, we find that a number of other AMPK-like kinase, such as SIK, NUAK, Par-1, KP78a, and KP78b show phenotypes similar to weak lkb1 loss of function in the eye. These data suggest that in complex tissues, LKB1 acts on an array of targets to regulate cell polarity.

Review on polyhydroxyalkanoate formation in the model plant Arabidopsis thaliana

In order to explore potential alternatives to the production of polyhydroxyalkanoates (PHAs) in bacteria, the enzymes of Alcaligenes eutrophus involved in the synthesis of polyhydroxybutyrate (PHB) have been expressed in the model plant Arabidopsis thaliana. Following the successful production of low amounts of high molecular weight PHB in plants expressing the acetoacetyl-CoA reductase and the PHB synthase in the cytoplasm of Arabidopsis cell, expression of the PHB pathway in the pastids was achieved by modifying the PHB enzymes with plastid targeting signals. This strategy resulted in a significant increase in the formation of PHB in Arabidopsis, with a maximum of 14% of the leaf dry weight . The increase in PHB production is most likely due to the higher flux in the plastids of acetyl-CoA, the precursor for PHB synthesis. A detailed study of metabolic fluxes in Arabidopsis plants producing high levels of PHB could help to determine the potential problems and limitations of PHB synthesis in Arabidopsis and could be useful for optimising strategies for the production of PHB in crop plants. The knowledge on PHB production could also be used for the production of PHAs other than PHB. Apart from PHB, no other PHAs have been produced in an eukaryotic system. Arabidopsis will therefore be used as a model system for the production in eukaryotes of more complex PHAs, such as poly(hydroxybutyrate-co-hydroxyvylerate) or medium-chain-lenght-PHAs.

Protein-protein interactions between adenovirus DNA polymerase and nuclear factor I mediate formation of the DNA replication preinitiation complex.

The in vitro adenovirus (Ad) DNA replication system provides an assay to study the interaction of viral and host replication proteins with the DNA template in the formation of the preinitiation complex. This initiation system requires in addition to the origin DNA sequences 1) Ad DNA polymerase (Pol), 2) Ad preterminal protein (pTP), the covalent acceptor for protein-primed DNA replication, and 3) nuclear factor I (NFI), a host cell protein identical to the CCAAT box-binding transcription factor. The interactions of these proteins were studied by coimmunoprecipitation and Ad origin DNA binding assays. The Ad Pol can bind to origin sequences only in the presence of another protein which can be either pTP or NFI. While NFI alone can bind to its origin recognition sequence, pTP does not specifically recognize DNA unless Ad Pol is present. Thus, protein-protein interactions are necessary for the targetting of either Ad Pol or pTP to the preinitiation complex. DNA footprinting demonstrated that the Ad DNA site recognized by the pTP.Pol complex was within the first 18 bases at the end of the template which constitutes the minimal origin of replication. Mutagenesis studies have defined the Ad Pol interaction site on NFI between amino acids 68-150, which overlaps the DNA binding and replication activation domain of this factor. A putative zinc finger on the Ad Pol has been mutated to a product that fails to bind the Ad origin sequences but still interacts with pTP. These results indicate that both protein-protein and protein-DNA interactions mediate specific recognition of the replication origin by Ad DNA polymerase.

Foam pore size is a critical interface parameter of suction-based wound healing devices.

BACKGROUND: Suction-based wound healing devices with open-pore foam interfaces are widely used to treat complex tissue defects. The impact of changes in physicochemical parameters of the wound interfaces has not been investigated. METHODS: Full-thickness wounds in diabetic mice were treated with occlusive dressing or a suction device with a polyurethane foam interface varying in mean pore size diameter. Wound surface deformation on day 2 was measured on fixed tissues. Histologic cross-sections were analyzed for granulation tissue thickness (hematoxylin and eosin), myofibroblast density (α-smooth muscle actin), blood vessel density (platelet endothelial cell adhesion molecule-1), and cell proliferation (Ki67) on day 7. RESULTS: Polyurethane foam-induced wound surface deformation increased with polyurethane foam pore diameter: 15 percent (small pore size), 60 percent (medium pore size), and 150 percent (large pore size). The extent of wound strain correlated with granulation tissue thickness that increased 1.7-fold in small pore size foam-treated wounds, 2.5-fold in medium pore size foam-treated wounds, and 4.9-fold in large pore size foam-treated wounds (p < 0.05) compared with wounds treated with an occlusive dressing. All polyurethane foams increased the number of myofibroblasts over occlusive dressing, with maximal presence in large pore size foam-treated wounds compared with all other groups (p < 0.05). CONCLUSIONS: The pore size of the interface material of suction devices has a significant impact on the wound healing response. Larger pores increased wound surface strain, tissue growth, and transformation of contractile cells. Modification of the pore size is a powerful approach for meeting biological needs of specific wounds.

The roles of flux- and decompression melting and their respective fractionation lines for continental crust formation: Evidence from the Kohistan arc

Delamination and foundering of the lower continental crust (LCC) into the mantle is part of the crust-forming mechanism. However, knowledge of the composition and mineralogy of the preserved or delaminated LCC over geological timescales remains scarce. We provide a synopsis of recent research within the Kohistan arc (Pakistan) and demonstrate that hydrous and less hydrous liquid lines of descent related to flux assisted and decompression mantle melting, respectively, produce compositionally different lower crustal rocks. The argument refers to two lower crustal sections exposed in Kohistan, the older Southern Plutonic Complex (SPC) and the younger Chilas Complex. The SPC typifies a hydrous, high-pressure fractionation sequence of olivine-pyroxenes-garnet-Fe/Ti-oxide-amphibole-plagioclase. The Chilas Complex illustrates a less hydrous fractionation sequence of olivine-clinopyroxene-orthopyroxene-plagioclase-amphibole. Despite the similarity of the Chilas Complex rocks to proposed lower crust compositions, the less hydrous fractionation results in unrealistically small volumes of silica-rich rocks, precluding the Chilas Complex gabbros to represent the magmatic complement to the upper crust. The composition of the SPC lower crust differs markedly from bulk lower crust estimates, but is complementary to silica-rich rocks exposed along this section and in the Kohistan batholith. These observations inspire a composite model for the formation of continental crust (CC) where the negatively buoyant delaminated and the buoyant preserved lower continental crusts (LCC) differ in genesis, mineralogy, and composition. We propose that the upper, non-sedimentary subsequent removal of the complementary, negatively buoyant garnet-pyroxene-amphibole-plagioclase-rich cumulates. In contrast, the LCC, which is buoyant and preserved over geological timescales, is formed by less hydrous parental mantle melts. We suggest that the bulk continental crust composition is related to mixing of these petrologically not directly related end members. Published by Elsevier B.V.

Xenopus peroxisome proliferator activated receptors: genomic organization, response element recognition, heterodimer formation with retinoid X receptor and activation by fatty acids.

Peroxisome proliferator activated receptors are ligand activated transcription factors belonging to the nuclear hormone receptor superfamily. Three cDNAs encoding such receptors have been isolated from Xenopus laevis (xPPAR alpha, beta, and gamma). Furthermore, the gene coding for xPPAR beta has been cloned, thus being the first member of this subfamily whose genomic organization has been solved. Functionally, xPPAR alpha as well as its mouse and rat homologs are thought to play an important role in lipid metabolism due to their ability to activate transcription of a reporter gene through the promoter of the acyl-CoA oxidase (ACO) gene. ACO catalyzes the rate limiting step in the peroxisomal beta-oxidation of fatty acids. Activation is achieved by the binding of xPPAR alpha on a regulatory element (DR1) found in the promoter region of this gene, xPPAR beta and gamma are also able to recognize the same type of element and are, as PPAR alpha, able to form heterodimers with retinoid X receptor. All three xPPARs appear to be activated by synthetic peroxisome proliferators as well as by naturally occurring fatty acids, suggesting that a common mode of action exists for all the members of this subfamily of nuclear hormone receptors.

De la conception à la pratique Reconfiguration d'un dispositif de formation des enseignants aux technologies de l'information et de la communication.

Constatant que la formation des enseignants aux technologies de l'information et de la communication (TIC) pose des problèmes spécifiques, cet article a pour but d'engager une réflexion sur ce thème. Nos réflexions s'appuient sur une expérience de formation réalisée dans 23 écoles professionnelles suisses engagées dans un programme national de soutien à l'utilisation pédagogique des TIC. L'article décrit la manière dont le dispositif de formation prévu a été mis en pratique, les difficultés auxquelles il a donné lieu et les réponses que les acteurs ont élaborées pour faire face à ces difficultés. Nous montrons qu'une analyse du déroulement effectif d'un dispositif de formation et des transformations qu'il subit constitue un élément essentiel pour orienter les futures actions de formation et penser la place des apprenants dans un dispositif de formation.

Edar/Eda interactions regulate enamel knot formation in tooth morphogenesis.

tabby and downless mutant mice have apparently identical defects in teeth, hair and sweat glands. Recently, genes responsible for these spontaneous mutations have been identified. downless (Dl) encodes Edar, a novel member of the tumour necrosis factor (TNF) receptor family, containing the characteristic extracellular cysteine rich fold, a single transmembrane region and a death homology domain close to the C terminus. tabby (Ta) encodes ectodysplasin-A (Eda) a type II membrane protein of the TNF ligand family containing an internal collagen-like domain. As predicted by the similarity in adult mutant phenotype and the structure of the proteins, we demonstrate that Eda and Edar specifically interact in vitro. We have compared the expression pattern of Dl and Ta in mouse development, taking the tooth as our model system, and find that they are not expressed in adjacent cells as would have been expected. Teeth develop by a well recorded series of epithelial-mesenchymal interactions, similar to those in hair follicle and sweat gland development, the structures found to be defective in tabby and downless mice. We have analysed the downless mutant teeth in detail, and have traced the defect in cusp morphology back to initial defects in the structure of the tooth enamel knot at E13. Significantly, the defect is distinct from that of the tabby mutant. In the tabby mutant, there is a recognisable but small enamel knot, whereas in the downless mutant the knot is absent, but enamel knot cells are organised into a different shape, the enamel rope, showing altered expression of signalling factors (Shh, Fgf4, Bmp4 and Wnt10b). By adding a soluble form of Edar to tooth germs, we were able to mimic the tabby enamel knot phenotype, demonstrating the involvement of endogenous Eda in tooth development. We could not, however, reproduce the downless phenotype, suggesting the existence of yet another ligand or receptor, or of ligand-independent activation mechanisms for Edar. Changes in the structure of the enamel knot signalling centre in downless tooth germs provide functional data directly linking the enamel knot with tooth cusp morphogenesis. We also show that the Lef1 pathway, thought to be involved in these mutants, functions independently in a parallel pathway.

A single point mutation in the pore region of the epithelial Na+ channel changes ion selectivity by modifying molecular sieving.

The epithelial Na+ channel (ENaC) belongs to a new class of channel proteins called the ENaC/DEG superfamily involved in epithelial Na+ transport, mechanotransduction, and neurotransmission. The role of ENaC in Na+ homeostasis and in the control of blood pressure has been demonstrated recently by the identification of mutations in ENaC beta and gamma subunits causing hypertension. The function of ENaC in Na+ reabsorption depends critically on its ability to discriminate between Na+ and other ions like K+ or Ca2+. ENaC is virtually impermeant to K+ ions, and the molecular basis for its high ionic selectivity is largely unknown. We have identified a conserved Ser residue in the second transmembrane domain of the ENaC alpha subunit (alphaS589), which when mutated allows larger ions such as K+, Rb+, Cs+, and divalent cations to pass through the channel. The relative ion permeability of each of the alphaS589 mutants is related inversely to the ionic radius of the permeant ion, indicating that alphaS589 mutations increase the molecular cutoff of the channel by modifying the pore geometry at the selectivity filter. Proper geometry of the pore is required to tightly accommodate Na+ and Li+ ions and to exclude larger cations. We provide evidence that ENaC discriminates between cations mainly on the basis of their size and the energy of dehydration.

Low molecular weight carboxylic acids in oxidizing porphyry copper tailings

The distribution of low molecular weight carboxylic acids (LMWCA) was investigated in pore water profiles from two porphyry copper tailings impoundments in Chile (Piuquenes at La Andina and Cauquenes at El Teniente mine). The objectives of this study were (1) to determine the distribution of LMWCA, which are interpreted to be the metabolic byproducts of the autotroph microbial community in this low organic carbon system, and (2) to infer the potential role of these acids in cycling of Fe and other elements in the tailings impoundments. The speciation and mobility of iron, and potential for the release of H+ via hydrolysis of the ferric iron, are key factors in the formation of acid mine drainage in sulfidic mine wastes. In the low-pH oxidation zone of the Piuquenes tailings, Fe(III) is the dominant iron species and shows high mobility. LMWCA, which occur mainly between the oxidation front down to 300 cm below the tailings surface at both locations (e.g., max concentrations of 0.12 mmol/L formate, 0.17 mmol/L acetate, and 0.01 mmol/L pyruvate at Piuquenes and 0.14 mmol/L formate, 0.14 mmol/L acetate, and 0.006 mmol/L pyruvate at Cauquenes), are observed at the same location as high Fe concentrations (up to 71.2 mmol/L Fe(II) and 16.1 mmol/L Fe(III), respectively). In this zone, secondary Fe(111) hydroxides are depleted. Our data suggest that LMWCA may influence the mobility of iron in two ways. First, complexation of Fe(III), through formation of bidentate Fe(III)-LMWCA complexes (e.g., pyruvate, oxalate), may enhance the dissolution of Fe(III) (oxy)hydroxides or may prevent precipitation of Fe(III) (oxy)hydroxides. Soluble Fe(III) chelate complexes which may be mobilized downward and convert to Fe(II) by Fe(III) reducing bacteria. Second, monodentate LMWCA (e.g., acetate and formate) can be used by iron-reducing bacteria as electron donors (e.g., Acidophilum spp.), with ferric iron as the electron acceptor. These processes may, in part, explain the low abundances of secondary Fe(III) hydroxide precipitates below the oxidation front and the high concentrations of Fe(II) observed in the pore waters of some low-sulfide systems. The reduction of Fe(III) and the subsequent increase of iron mobility and potential acidity transfer (Fe(II) oxidation can result in the release of H+ in an oxic environment) should be taken in account in mine waste management strategies.