Monitoring protein-protein interactions between the mammalian integral membrane transporters and PDZ-interacting partners using a modified split-ubiquitin membrane yeast two-hybrid system

PDZ-binding motifs are found in the C-terminal tails of numerous integral membrane proteins where they mediate specific protein-protein interactions by binding to PDZ-containing proteins. Conventional yeast two-hybrid screens have been used to probe protein-protein interactions of these soluble C termini. However, to date no in vivo technology has been available to study interactions between the full-length integral membrane proteins and their cognate PDZ-interacting partners. We previously developed a split-ubiquitin membrane yeast two-hybrid (MYTH) system to test interactions between such integral membrane proteins by using a transcriptional output based on cleavage of a transcription factor from the C terminus of membrane-inserted baits. Here we modified MYTH to permit detection of C-terminal PDZ domain interactions by redirecting the transcription factor moiety from the C to the N terminus of a given integral membrane protein thus liberating their native C termini. We successfully applied this "MYTH 2.0" system to five different mammalian full-length renal transporters and identified novel PDZ domain-containing partners of the phosphate (NaPi-IIa) and sulfate (NaS1) transporters that would have otherwise not been detectable. Furthermore this assay was applied to locate the PDZ-binding domain on the NaS1 protein. We showed that the PDZ-binding domain for PDZK1 on NaS1 is upstream of its C terminus, whereas the two interacting proteins, NHERF-1 and NHERF-2, bind at a location closer to the N terminus of NaS1. Moreover NHERF-1 and NHERF-2 increased functional sulfate uptake in Xenopus oocytes when co-expressed with NaS1. Finally we used MYTH 2.0 to demonstrate that the NaPi-IIa transporter homodimerizes via protein-protein interactions within the lipid bilayer. In summary, our study establishes the MYTH 2.0 system as a novel tool for interactive proteomics studies of membrane protein complexes.

Driving the refactoring of Java Enterprise applications by evaluating the distance between application elements

Java Enterprise Applications (JEAs) are complex systems composed using various technologies that in turn rely on languages other than Java, such as XML or SQL. Given the complexity of these applications, the need to reverse engineer them in order to support further development becomes critical. In this paper we show how it is possible to split a system into layers and how is possible to interpret the distance between application elements in order to support the refactoring of JEAs. The purpose of this paper is to explore ways to provide suggestions about the refactoring operations to perform on the code by evaluating the distance between layers and elements belonging those layers. We split JEAs into layers by considering the kinds and the purposes of the elements composing the application. We measure distance between elements by using the notion of the shortest path in a graph. Also we present how to enrich the interpretation of the distance value with enterprise pattern detection in order to refine the suggestion about modifications to perform on the code.

Split-mouth designs in orthodontics: an overview with applications to orthodontic clinical trials

SUMMARY Split-mouth designs first appeared in dental clinical trials in the late sixties. The main advantage of this study design is its efficiency in terms of sample size as the patients act as their own controls. Cited disadvantages relate to carry-across effects, contamination or spilling of the effects of one intervention to another, period effects if the interventions are delivered at different time periods, difficulty in finding similar comparison sites within patients and the requirement for more complex data analysis. Although some additional thought is required when utilizing a split-mouth design, the efficiency of this design is attractive, particularly in orthodontic clinical studies where carry-across, period effects and dissimilarity between intervention sites does not pose a problem. Selection of the appropriate research design, intervention protocol and statistical method accounting for both the reduced variability and potential clustering effects within patients should be considered for the trial results to be valid.

Split for the cure: VEGF, PDGF-BB and intussusception in therapeutic angiogenesis

Therapeutic angiogenesis is an attractive strategy to treat patients suffering from ischaemic conditions and vascular endothelial growth factor-A (VEGF) is the master regulator of blood vessel growth. However, VEGF can induce either normal or aberrant angiogenesis depending on its dose localized in the microenvironment around each producing cell in vivo and on the balanced stimulation of platelet-derived growth factor-BB (PDGF-BB) signalling, responsible for pericyte recruitment. At the doses required to induce therapeutic benefit, VEGF causes new vascular growth essentially without sprouting, but rather through the alternative process of intussusception, or vascular splitting. In the present article, we briefly review the therapeutic implications of controlling VEGF dose on one hand and pericyte recruitment on the other, as well as the key features of intussusceptive angiogenesis and its regulation.

Marital split-up and widowhood in old age: Differential impact on psychological and social well-being

Marital split-up and spousal loss are among the most stressful critical life events. Numerous studies have documented their detrimental effects on well-being, yet the large individual differences in psychological adaptation are still not well understood. Whereas in old age bereavement is normative and can be anticipated, divorce is an “off-time” transition for this age group. In contrast to bereavement which has been amply studied, research on later life divorce is still missing despite the increasing relevance of the topic due to the significant increase of divorces in older age. Based on a modified and extended view of Amato’s divorce-stress-adjustment model (2000), the aim of this contribution is to explore the differential impact of marital split-up and widowhood in older age on psychological (life satisfaction) and social well-being (social loneliness), and the adaptation to these critical life events. Our analyses are based on data gathered in a questionnaire study, which is part of the Swiss National Centre of Competence in Research LIVES. In a first step we compared three groups of individuals aged 60 to 75 years: a sample of 251 persons with a marital split-up (127 women; 123 men), a sample of 270 widowed persons (170 women; 100 men), and a group of 221 continuously married people (110 women; 111 men), which served as control group. In a second step, we investigated the role of socio-demographic variables, intrapersonal and interpersonal resources and variables of the context of loss as predictors for the psychological adaptation to a marital break-up and loss in old age. First results by ANCOVA indicate significant differences with regard to life satisfaction among the three groups, with divorced persons with the lowest scores, followed by the bereaved ones, and the married controls with the highest. Regarding social loneliness, divorced individuals report higher social loneliness than the bereaved group and the married controls (no significant difference between widowed and the married). In both loss groups, financial and intrapersonal resources, as well as the emotional valence of the loss are the most important predictors for the psychological and social adaptation. However, happiness in the past relationship is an important resource regarding the indicators for adaptation for the widowers, but not for individuals with a marital dissolution.

Effects of PEG-Induced Water Deficit in Solanum nigrum on Zn and Ni Uptake and Translocation in Split Root Systems

Drought strongly influences root activities in crop plants and weeds. This paper is focused on the performance of the heavy metal accumulator Solanum nigrum, a plant which might be helpful for phytoremediation. The water potential in a split root system was decreased by the addition of polyethylene glycol (PEG 6000). Rubidium, strontium and radionuclides of heavy metals were used as markers to investigate the uptake into roots, the release to the shoot via the xylem, and finally the basipetal transport via the phloem to unlabeled roots. The uptake into the roots (total contents in the plant) was for most makers more severely decreased than the transport to the shoot or the export from the shoot to the unlabeled roots via the phloem. Regardless of the water potential in the labeling solution, 63Ni and 65Zn were selectively redistributed within the plant. From autoradiographs, it became evident that 65Zn accumulated in root tips, in the apical shoot meristem and in axillary buds, while 63Ni accumulated in young expanded leaves and roots but not in the meristems. Since both radionuclides are mobile in the phloem and are, therefore, well redistributed within the plant, the unequal transfer to shoot and root apical meristems is most likely caused by differences in the cell-to-cell transport in differentiation zones without functional phloem (immature sieve tubes).

Giant Electric Field Enhancement in Split Ring Resonators Featuring Nanometer-Sized Gaps

Today's pulsed THz sources enable us to excite, probe, and coherently control the vibrational or rotational dynamics of organic and inorganic materials on ultrafast time scales. Driven by standard laser sources THz electric field strengths of up to several MVm−1 have been reported and in order to reach even higher electric field strengths the use of dedicated electric field enhancement structures has been proposed. Here, we demonstrate resonant electric field enhancement structures, which concentrate the incident electric field in sub-diffraction size volumes and show an electric field enhancement as high as ~14,000 at 50 GHz. These values have been confirmed through a combination of near-field imaging experiments and electromagnetic simulations.

N budget and NH3 exchange of a grass/clover crop at two levels of N application

Atmospheric ammonia (NH3) exchange during a single growing season was measured over two grass/clover fields managed by cutting and treated with different rates of mineral nitrogen (N) fertilizer. The aim was to quantify the total NH3 exchange of the two systems in relation to their N budget, the latter was split into N derived from symbiotic fixation, from fertilization, and from the soil. The experimental site was located in an intensively managed agricultural area on the Swiss plateau. Two adjacent fields with mixtures of perennial ryegrass (Lolium perenne L.), cocks foot (Dactylis glomerata L.), white clover (Trifolium repens L.) and red clover (Trifolium pratense L.) were used. These were treated with either 80 or 160 kg N ha−1 applied as NH4NO3 fertilizer in equal portions after each of four cuts. Continuous NH3 flux measurements were carried out by micrometeorological techniques. To determine the contribution of each species to the overall NH3 canopy compensation point, stomatal NH3 compensation points of the individual plant species were determined on the basis of NH4+ + NH3 (NHx) concentrations and pH in the apoplast. Symbiotic N2 fixation was measured by the 15N dilution method.