Designer TGFβ superfamily ligands with diversified functionality

Transforming Growth Factor--beta (TGFβ) superfamily ligands, including Activins, Growth and Differentiation Factors (GDFs), and Bone Morphogenetic Proteins (BMPs), are excellent targets for protein-based therapeutics because of their pervasiveness in numerous developmental and cellular processes. We developed a strategy termed RASCH (Random Assembly of Segmental Chimera and Heteromer), to engineer chemically-refoldable TGFβ superfamily ligands with unique signaling properties. One of these engineered ligands, AB208, created from Activin-βA and BMP-2 sequences, exhibits the refolding characteristics of BMP-2 while possessing Activin-like signaling attributes. Further, we find several additional ligands, AB204, AB211, and AB215, which initiate the intracellular Smad1-mediated signaling pathways more strongly than BMP-2 but show no sensitivity to the natural BMP antagonist Noggin unlike natural BMP-2. In another design, incorporation of a short N-terminal segment from BMP-2 was sufficient to enable chemical refolding of BMP-9, without which was never produced nor refolded. Our studies show that the RASCH strategy enables us to expand the functional repertoire of TGFβ superfamily ligands through development of novel chimeric TGFβ ligands with diverse biological and clinical values.     

Protein Kinase B Localization and Activation Differentially Affect S6 Kinase 1 Activity and Eukaryotic Translation Initiation Factor 4E-Binding Protein 1 Phosphorylation

Recent studies indicate that phosphatidylinositide-3OH kinase (PI3K)-induced S6 kinase (S6K1) activation is mediated by protein kinase B (PKB). Support for this hypothesis has largely relied on results obtained with highly active, constitutively membrane-localized alleles of wild-type PKB, whose activity is independent of PI3K. Here we set out to examine the importance of PKB signaling in S6K1 activation. In parallel, glycogen synthase kinase 3beta (GSK-3beta) inactivation and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) phosphorylation were monitored as markers of the rapamycin-insensitive and -sensitive branches of the PI3K signaling pathway, respectively. The results demonstrate that two activated PKBalpha mutants, whose basal activity is equivalent to that of insulin-induced wild-type PKB, inhibit GSK-3beta to the same extent as a highly active, constitutively membrane-targeted wild-type PKB allele. However, of these two mutants, only the constitutively membrane-targeted allele of PKB induces S6K1 activation. Furthermore, an interfering mutant of PKB, which blocks insulin-induced PKB activation and GSK-3beta inactivation, has no effect on S6K1 activation. Surprisingly, all the activated PKB mutants, regardless of constitutive membrane localization, induce 4E-BP1 phosphorylation and the interfering PKB mutant blocks insulin-induced 4E-BP1 phosphorylation. The results demonstrate that PKB mediates S6K1 activation only as a function of constitutive membrane localization, whereas the activation of PKB appears both necessary and sufficient to induce 4E-BP1 phosphorylation independently of its intracellular location.     

Epitope down-modulation as a mechanism for the coexistence of competing T-cells

Efficient immune responses against pathogens are frequently characterized by the simultaneous targeting of multiple epitopes. However, it remains unclear how the targeting of multiple epitopes is maintained in the face of competition for antigenic stimulation. Here, we investigate this question by using mathematical models of the population dynamics of a viral pathogen, antigen presentation sites and T-cells. We first show that direct competition for access to antigen presenting sites and indirect competition through killing of the pathogen select for dominance of the T-cell response with the highest affinity for its epitope. We then incorporate in our model that epitopes can become down-modulated following interaction with epitope specific T-cells. We demonstrate that epitope down-modulation leads to differentiation of epitope presentation on antigen presenting sites. This differentiation promotes the coexistence of multiple epitope specific responses. Hence, we propose that the functional relevance of epitope down-modulation may be to enable the persistence of a broad immune response despite competition for antigenic stimulation.     

Xenopus cadherin-6 regulates growth and epithelial development of the retina

Cadherins are crucial for tissue cohesion, separation of cell layers and cell migration during embryogenesis. To investigate the role of classical type II Xcadherin-6 (Xcad-6), we performed loss-of-function studies by morpholino oligonucleotide injections. This resulted in severe eye defects which could be rescued with murine cadherin-6. In the absence of Xcadherin-6, morphological alterations and a decrease in cell proliferation were observed with eye cup formation. Eye field transplantations of Xcadherin-6 depleted donors yielded grafts that failed to form a proper neuroepithelium in a wildtype environment. At later developmental stages Xcadherin-6 deficient eyes showed lamination defects in the outer neural retina, a reduced thickness of the ganglion cell layer (GCL) and a fragmented retina pigment epithelium (RPE). Thus, Xcadherin-6 is essential early in eye development for structural organization and growth of the neuroepithelium before it differentiates into neural retina and RPE.     

Lipid metabolism in cancer

Lipids form a diverse group of water-insoluble molecules that include triacylglycerides, phosphoglycerides, sterols and sphingolipids. They play several important roles at cellular and organismal levels. Fatty acids are the major building blocks for the synthesis of triacylglycerides, which are mainly used for energy storage. Phosphoglycerides, together with sterols and sphingolipids, represent the major structural components of biological membranes. Lipids can also have important roles in signalling, functioning as second messengers and as hormones. There is increasing evidence that cancer cells show specific alterations in different aspects of lipid metabolism. These alterations can affect the availability of structural lipids for the synthesis of membranes, the synthesis and degradation of lipids that contribute to energy homeostasis and the abundance of lipids with signalling functions. Changes in lipid metabolism can affect numerous cellular processes, including cell growth, proliferation, differentiation and motility. This review will examine some of the alterations in lipid metabolism that have been reported in cancer, at both cellular and organismal levels, and discuss how they contribute to different aspects of tumourigenesis.     

Comparing and evaluating process-based ecosystem model predictions of carbon and water fluxes in major European forest biomes

Process‐based models can be classified into: (a) terrestrial biogeochemical models (TBMs), which simulate fluxes of carbon, water and nitrogen coupled within terrestrial ecosystems, and (b) dynamic global vegetation models (DGVMs), which further couple these processes interactively with changes in slow ecosystem processes depending on resource competition, establishment, growth and mortality of different vegetation types. In this study, four models – RHESSys, GOTILWA+, LPJ‐GUESS and ORCHIDEE – representing both modelling approaches were compared and evaluated against benchmarks provided by eddy‐covariance measurements of carbon and water fluxes at 15 forest sites within the EUROFLUX project. Overall, model‐measurement agreement varied greatly among sites. Both modelling approaches have somewhat different strengths, but there was no model among those tested that universally performed well on the two variables evaluated. Small biases and errors suggest that ORCHIDEE and GOTILWA+ performed better in simulating carbon fluxes while LPJ‐GUESS and RHESSys did a better job in simulating water fluxes. In general, the models can be considered as useful tools for studies of climate change impacts on carbon and water cycling in forests. However, the various sources of variation among models simulations and between models simulations and observed data described in this study place some constraints on the results and to some extent reduce their reliability. For example, at most sites in the Mediterranean region all models generally performed poorly most likely because of problems in the representation of water stress effects on both carbon uptake by photosynthesis and carbon release by heterotrophic respiration (R_h). The use of flux data as a means of assessing key processes in models of this type is an important approach to improving model performance. Our results show that the models have value but that further model development is necessary with regard to the representation of the some of the key ecosystem processes.     

Responses of aquatic algae and cyanobacteria to solar UV-B

Continuous depletion of the stratospheric ozone layer has resulted in an increase in solar ultraviolet-B (UV-B; 280–315 nm) radiation reaching the Earth's surface. The consequences for aquatic phototrophic organisms of this small change in the solar spectrum are currently uncertain. UV radiation has been shown to adversely affect a number of photochemical and photobiological processes in a wide variety of aquatic organisms, such as cyanobacteria, phytoplankton and macroalgae. However, a number of photosynthetic organisms counteract the damaging effects of UV-B by synthesizing UV protective compounds such as mycosporine-like amino acids (MAAs) and the cyanobacterial sheath pigment, scytonemin. The aim of this contribution is to discuss the responses of algae and cyanobacteria to solar UV-B radiation and the role of photoprotective compounds in mitigating UV-B damage.     

Receptor based models for spontaneous colour choices in flies and butterflies

Colour is one of several stimuli used by herbivorous insects in host choice. Insects have between 2 and 5 different types of photoreceptors to catch quanta of different wavelengths of the spectrum. Many insects have been shown to possess opponent neural interactions between the receptors that enable them to see colour. I present simple models to describe colour choices as functions of the receptor quantum catches and linear interactions of the receptor types. Models are applied to data sets obtained from own experiments and from the literature, on Pieris brassicae and P. rapae (Lepidoptera, Pieridae), Papilio aegeus (Papilionidae), Dacus oleae (Diptera, Tephritidae) and Eristalis tenax (Syrphidae). In fruit flies, detection of green fruit is based on an inhibitory interaction between a green-sensitive receptor type and a blue-sensitive receptor type. This might explain the preference many herbivorous insects have for yellow over green stimuli. Pollen feeding in hoverflies might have evolved from yellow pollen being a super-normal stimulus for herbivorous insects. In butterflies, an additional red-receptor is involved in the colour choice for an oviposition substratum and leads to them choosing green and not yellow. The models introduced in this study open new perspective for a physiological understanding of the design of visual stimuli for monitoring and trapping pest insects.