A comprehensive, non-invasive visualization of primordial germ cell development in mice by the Prdm1-mVenus and Dppa3-ECFP double transgenic reporter

The ability to monitor the development of a given cell lineage in a non-invasive manner by fluorescent markers both in vivo and in vitro provides a great advantage for the analysis of the lineage of interest. To date, a number of transgenic or knock-in mouse strains, in which developing germ cells are marked with fluorescent reporters, have been generated. We here describe a novel double transgenic reporter mouse strain that expresses membrane-targeted Venus (mVenus), a brighter variant of yellow fluorescent protein (YFP), under the control of Prdm1 (Blimp1) regulatory elements and enhanced cyan fluorescent protein (ECFP) under the control of Dppa3 (Stella/Pgc7). The double transgenic strain unambiguously marked Prdm1 expression in the lineage-restricted precursors of primordial germ cells (PGCs) in the proximal epiblast at embryonic day (E) 6.25 and specifically illuminated Prdm1- and Dppa3-positive migrating PGCs after E8.5. The double transgenic reporter also precisely recapitulated dynamic embryonic expression of Prdm1 outside the germ cell lineage. Moreover, we derived ES cells that bore both transgenes. These cells made a robust contribution both to the germ and somatic cell lineages in chimeras with accurate Prdm1-mVenus and Dppa3-ECFP expression. The transgenic strain and the ES cells will serve as valuable experimental materials not only for analyzing the origin and properties of the germ cell lineage in vivo, but also for establishing a culture system to efficiently induce proper germ cells with temporally coordinated Prdm1 and Dppa3 expression in vitro.     

Liposome and lipid bilayer arrays towards biosensing applications

Sensitive and selective biosensors for high-throughput screening are having an increasing impact in modern medical care. The establishment of robust protein biosensing platforms however remains challenging, especially when membrane proteins are involved. Although this type of proteins is of enormous relevance since they are considered in >60% of the pharmaceutical drug targets, their fragile nature (i.e., the requirement to preserve their natural lipid environment to avoid denaturation and loss of function) puts strong additional prerequisites onto a successful biochip. In this review, the leading approaches to create lipid membrane-based arrays towards the creation of membrane protein biosensing platforms are described. Liposomes assembled in micro- and nanoarrays and the successful set-ups containing functional membrane proteins, as well as the use of liposomes in networks, are discussed in the first part. Then, the complementary approaches to create cell-mimicking supported membrane patches on a substrate in an array format will be addressed. Finally, the progress in assembling free-standing (functional) lipid bilayers over nanopore arrays for ion channel sensing will be reported. This review illustrates the rapid pace by which advances are being made towards the creation of a heterogeneous biochip for the high-throughput screening of membrane proteins for diagnostics, drug screening, or drug discovery purposes.     

Study of cysteine residues in the {alpha} subunit of Escherichia coli tryptophan synthase. 2. Role in enzymatic function

To understand the functional roles of Cys residues in the subunitof tryptophan synthase from Escherichia coli, single mutantsof the subunit, in which each of the three Cys residues wassubstituted with Ser, Gly, Ala or Val, were constructed by site-directedmutagenesis. The effects of the substitutions on the functionof tryptophan synthase were investigated by activity measurements,calorimetric measurements of association with the ßsubunit and steadystate kinetic analysis of catalysis. Althoughthe three Cys residues are located away from the apparentlyimportant parts for enzymatic activity, substitutions at position81 by Ser, Ala or Val caused decreases in the intrinsic activityof the subunit. Furthermore, Cys81Ser and Cys81Val reducedstimulation activities in the and ß reactions dueto formation of a complex with the ß subunit. Thelower stimulation activities of the mutant proteins were notcorrelated with their abilities to associate with the ßsubunit but were correlated with decreases in k_cat. The presentresults suggest that position 81 plays an indirectly importantrole in the activity of the subunit itself and the mutual activationmechanism of the complex.     

Crystallographic Data of a New Phase of Dicalcium Silicate

Unit-cell parameters and the space group of a new phase of dicalcium silicate (Ca₂SiO₄) were determined by using powder X-ray diffractometry and selected-area electron diffraction techniques. This phase could be synthesized via the dissociation of hydrothermally synthesized alpha-Ca₂(SiO₄H)OH at temperatures of ∼500°-920°C. Crystallographic data for the sample synthesized at 600°C were as follows: Ca₂SiO₄, monoclinic; P 2₁/ c space group; lattice parameters of a = 0.82147(9) nm, b = 0.9808(1) nm, c = 0.9741(1) nm, and β= 94.642(7)°; cell volume ( V ) of 0.7857(1) nm³; Z = 8 (where Z is the number of chemical formula units in a unit cell); and a density of 2.91 g/cm³. The crystallographic data for samples synthesized at 800°C had slightly different unit-cell parameters of a = 0.82124(6) nm, b = 0.97348(7) nm, c = 0.97935(7) nm, β= 94.831(5)°, and V = 0.7849(1) nm³. Structural relationships of the new phase with the other dicalcium silicates are discussed.     

Regulatory mechanisms of periodontal regeneration

The periodontal ligament, located between the cementum and the alveolar bone, has a width ranging from 0.15 to 0.38 mm. Regeneration and homeostasis of the periodontal ligament are highly significant functions in relation to periodontal therapy, tooth transplantation or replantation, and orthodontic tooth movement. The purpose of this review is to discuss the regulatory mechanisms of regenerative and homeostatic functions in the periodontal ligament based on currently published studies and also on our own experimental data. We consider the capability of the ligament tissue to promote or to suppress calcification in connection with bone and cementum formation and the maintenance of the periodontal ligament space. Also discussed are the involvement of the periodontal ligament tissue in the regenerative ability, cell proliferation, growth and differentiation factors, extracellular matrix proteins, homeostatic phenomena, function of Malassez epithelial rests, tooth movement, or occlusal loading. Regulatory mechanisms for regeneration and homeostasis of the periodontal ligament are hypothetically proposed.     

Corn Plants Treated with Jasmonic Acid Attract More Specialist Parasitoids, Thereby Increasing Parasitization of the Common Armyworm

We investigated whether corn plants treated with jasmonic acid (JA) increases the ability of the parasitic wasp, Cotesia kariyai, to find and control the common armyworm (Mythimna separata) under laboratory conditions. The rank order of attractiveness increased from intact plants treated with distilled water (DW) (2 d), JA-treated intact plants (2 d), DW-treated infested plants (2 d) to JA-treated infested plants (2 d). Single JA-treatment to either infested or uninfested plants increased attractiveness to C. kariyai over a period lasting at least 10 d. We then showed that the increase in attractiveness of infested corn plants by JA-treatment resulted in increased parasitism by C. kariyai. These results hold a promise for field application of JA-treatment. First, JA-treatment not only promotes the attractiveness of uninfested plants, but also armyworm-infested plants. Thus, parasitoids are not likely to waste time on JA-treated uninfested plants when JA-treated infested plants are available. Second, the effect of JA-treatment is lasting for at least 10 d, a result now obtained in two independent studies.     

Fluoride and apatite formation in vivo and in vitro

In recent years, the biomineralization process has attracted much interest from academics and industries for potential technological application. The rule in biomineralization is to have a variety of interfaces and surfaces which can act as nucleators. The ultimate step in any biomineralization process, i.e. the deposition of mineral, must conform to the driving forces operating on the system. A new paradigm in the assessment of the driving force for biomineralization is that a variety of ions existing in the mineralizing milieu are not a bystander, but are instead an active player that directly regulates the precipitation process and nature of biogenic apatites. Thus, the most putative stoichiometric model of a biomineral is (Ca)(5-x)(Mg)q(Na)u(HPO4)v(CO3)w(PO4)(3-y)(OH,F)(1-z). Fluoride participates in many aspects of calcium phosphate formation in vivo and has enormous effects on its process and on the nature and properties of the final products. In the development of biogenic apatites, fluoride ion in the mineralizing media is supposed to accelerate the hydrolysis of acidic precursor(s) and increase the growth rates by augmenting the driving force for precipitation. Inhibitory activities of ions and molecules are related to their adsorption onto the apatite surfaces. From theoretical and practical points of view, it is of paramount importance to elucidate and predict the effect and outcome of fluoride (accelerator) and inhibitors of biological relevance, because of their use in combination for healthcare in dentistry and medicine, e.g. prevention of dental caries and calculus deposition and in the formulation of antiosteoporosis treatments.