Modelling the spatial and temporal constrains of the GABAergic influence on neuronal excitability

GABA (γ-amino butyric acid) is an inhibitory neurotransmitter in the adult brain that can mediate depolarizing responses during development or after neuropathological insults. Under which conditions GABAergic membrane depolarizations are sufficient to impose excitatory effects is hard to predict, as shunting inhibition and GABAergic effects on spatiotemporal filtering of excitatory inputs must be considered. To evaluate at which reversal potential a net excitatory effect was imposed by GABA (E_GABA^Thr), we performed a detailed in-silico study using simple neuronal topologies and distinct spatiotemporal relations between GABAergic and glutamatergic inputs.These simulations revealed for GABAergic synapses located at the soma an E_GABA^Thr close to action potential threshold (E_AP^Thr), while with increasing dendritic distance E_GABA^Thr shifted to positive values. The impact of GABA on AMPA-mediated inputs revealed a complex temporal and spatial dependency. E_GABA^Thr depends on the temporal relation between GABA and AMPA inputs, with a striking negative shift in E_GABA^Thr for AMPA inputs appearing after the GABA input. The spatial dependency between GABA and AMPA inputs revealed a complex profile, with E_GABA^Thr being shifted to values negative to E_AP^Thr for AMPA synapses located proximally to the GABA input, while for distally located AMPA synapses the dendritic distance had only a minor effect on E_GABA^Thr. For tonic GABAergic conductances E_GABA^Thr was negative to E_AP^Thr over a wide range of g_GABA^tonic values. In summary, these results demonstrate that for several physiologically relevant situations E_GABA^Thr is negative to E_AP^Thr, suggesting that depolarizing GABAergic responses can mediate excitatory effects even if E_GABA did not reach E_AP^Thr.     

Trends and challenges in the microbial production of lignocellulosic bioalcohol fuels

Bioalcohols produced by microorganisms from renewable materials are promising substitutes for traditional fuels derived from fossil sources. For several years already ethanol is produced in large amounts from feedstocks such as cereals or sugar cane and used as a blend for gasoline or even as a pure biofuel. However, alcohols with longer carbon chains like butanol have even more suitable properties and would better fit with the current fuel distribution infrastructure. Moreover, ethical concerns contradict the use of food and feed products as a biofuel source. Lignocellulosic biomass, especially when considered as a waste material offers an attractive alternative. However, the recalcitrance of these materials and the inability of microorganisms to efficiently ferment lignocellulosic hydrolysates still prevent the production of bioalcohols from these plentiful sources. Obviously, no known organism exist which combines all the properties necessary to be a sustainable bioalcohol producer. Therefore, breeding technologies, genetic engineering and the search for undiscovered species are promising means to provide a microorganism exhibiting high alcohol productivities and yields, converting all lignocellulosic sugars or are even able to use carbon dioxide or monoxide, and thereby being highly resistant to inhibitors and fermentation products, and easy to cultivate in huge bioreactors. In this review, we compare the properties of various microorganisms, bacteria and yeasts, as well as current research efforts to develop a reliable lignocellulosic bioalcohol producing organism.     

It Takes Two to Tango: Activation of Protein Kinase D by Dimerization

The recent discovery and structure determination of a novel ubiquitin-like dimerization domain in protein kinase D (PKD) has significant implications for its activation. PKD is a serine/threonine kinase activated by the lipid second messenger diacylglycerol (DAG). It is an essential and highly conserved protein that is implicated in plasma membrane directed trafficking processes from the trans-Golgi network. However, many open questions surround its mechanism of activation, its localization, and its role in the biogenesis of cargo transport carriers. In reviewing this field, the focus is primarily on the mechanisms that control the activation of PKD at precise locations in the cell. In light of the new structural findings, the understanding of the mechanisms underlying PKD activation is critically evaluated, with particular emphasis on the role of dimerization in PKD autophosphorylation, and the provenance and recognition of the DAG that activates PKD.     

The third pillar of bacterial signal transduction: classification of the extracytoplasmic function (ECF) σ factor protein family

The ability of a bacterial cell to monitor and adaptively respond to its environment is crucial for survival. After one- and two-component systems, extracytoplasmic function (ECF) σ factors – the largest group of alternative σ factors – represent the third fundamental mechanism of bacterial signal transduction, with about six such regulators on average per bacterial genome. Together with their cognate anti-σ factors, they represent a highly modular design that primarily facilitates transmembrane signal transduction. A comprehensive analysis of the ECF σ factor protein family identified more than 40 distinct major groups of ECF σ factors. The functional relevance of this classification is supported by the sequence similarity and domain architecture of cognate anti-σ factors, genomic context conservation, and potential target promoter motifs. Moreover, this phylogenetic analysis revealed unique features indicating novel mechanisms of ECF-mediated signal transduction. This classification, together with the web tool ECF finder and the information stored in the Microbial Signal Transduction (MiST) database, provides a comprehensive resource for the analysis of ECF σ factor-dependent gene regulation.     

Label-Free Kinetic Studies of Hemostasis-Related Biomarkers Including D-Dimer Using Autologous Serum Transfusion

The objective of this study was to evaluate the elimination kinetics of hemostasis-related biomarkers including the prothrombin activation fragment F1+2, thrombin-antithrombin complex (TAT), plasmin-α₂-antiplasmin complex (PAP), and D-dimer in humans. Autologous serum was used as a biomarker source and infused into 15 healthy volunteers. Serum was prepared from whole blood in the presence of recombinant tissue-type plasminogen activator (final concentration 20 μg/mL) to induce plasmin generation required for PAP and D-dimer formation. Serum transfusions (50 mL/30 min) were well tolerated by all subjects. Endogenous thrombin formation was not induced by serum infusions as measured using a highly sensitive oligonucleotide-based enzyme capture assay. Median peak levels (x-fold increase over baseline) of F1+2, TAT, PAP, and D-dimer of 3.7 nmol/L (28.9), 393 ng/mL (189.6), 3,829 ng/mL (7.0), and 13.4 mg/L (34.2) were achieved at the end of serum infusions. During a 48 h lasting follow-up period all biomarkers showed elimination kinetics of a two-compartment model. Median (interquartile range) terminal half-lives were 1.9 (1.3–3.6) h for F1+2, 0.7 (0.7–2.6) h for TAT, and 10.8 (8.8–11.4) h for PAP. With 15.8 (13.1–23.1) h the D-dimer half-life was about twice as long as previously estimated from radiolabeling studies in animals and small numbers of human subjects. The serum approach presented here allows label-free and simultaneous analysis of the elimination kinetics of various hemostasis-related biomarkers. Based on these data changes in biomarker levels could more precisely used to estimate the activity level of the hemostatic system.     

Therapeutic Implications from Sensitivity Analysis of Tumor Angiogenesis Models

Anti-angiogenic cancer treatments induce tumor starvation and regression by targeting the tumor vasculature that delivers oxygen and nutrients. Mathematical models prove valuable tools to study the proof-of-concept, efficacy and underlying mechanisms of such treatment approaches. The effects of parameter value uncertainties for two models of tumor development under angiogenic signaling and anti-angiogenic treatment are studied. Data fitting is performed to compare predictions of both models and to obtain nominal parameter values for sensitivity analysis. Sensitivity analysis reveals that the success of different cancer treatments depends on tumor size and tumor intrinsic parameters. In particular, we show that tumors with ample vascular support can be successfully targeted with conventional cytotoxic treatments. On the other hand, tumors with curtailed vascular support are not limited by their growth rate and therefore interruption of neovascularization emerges as the most promising treatment target.     

Synthesis and SAR requirements of adamantane-colchicine conjugates with both microtubule depolymerizing and tubulin clustering activities

A series of analogues of conjugate 1, combining an adamantane-based paclitaxel (taxol) mimetic with colchicine was synthesized and tested for cytotoxicity in a cell-based assay with the human lung carcinoma cell line A549. The most active compounds (10 EC(50) 2 ± 1.0 nM, 23 EC(50) 6 ± 1.4 nM, 26 EC(50) 5 ± 1.8 nM, 28 EC(50) 11 ± 1.7 nM, 30 EC(50) 4.8 ± 0.5 nM) were found to interfere with the microtubule dynamics in an interesting manner. Treatment of the cells with these compounds promoted disassembly of microtubules followed by the formation of stable tubulin clusters. Structure-activity relationships for the analogues of 23 revealed the sensitivity of both cytotoxicity and tubulin clustering ability to the linker length. The presence of adamantane (or another bulky hydrophobic and non-aromatic moiety) in 23 was found to play an important role in the formation of tubulin clusters. Structural requirements for optimal activity have been partially explained by molecular modeling.