Molecular drift of the bride of sevenless (boss) gene in Drosophila

DNA sequences were determined for three to five alleles of the bride-of- sevenless (boss) gene in each of four species of Drosophila. The product of boss is a transmembrane receptor for a ligand coded by the sevenless gene that triggers differentiation of the R7 photoreceptor cell in the compound eye. Population parameters affecting the rate and pattern of molecular evolution of boss were estimated from the multinomial configurations of nucleotide polymorphisms of synonymous codons. The time of divergence between D. melanogaster and D. simulans was estimated as approximately 1 Myr, that between D. teissieri and D. yakuba as approximately 0.75 Myr, and that between the two pairs of sibling species as approximately 2 Myr. (The boss genes themselves have estimated divergence times approximately 50% greater than the species divergence times.) The effective size of the species was estimated as approximately 5 x 10(6), and the average mutation rate was estimated as 1-2 x 10(-9)/nucleotide/generation. The ratio of amino acid polymorphisms within species to fixed differences between species suggests that approximately 25% of all possible single-step amino acid replacements in the boss gene product may be selectively neutral or nearly neutral. The data also imply that random genetic drift has been responsible for virtually all of the observed differences in the portion of the boss gene analyzed among the four species.      

Molecular dynamics studies of the HIV-1 TAR and its complex with argininamide

The dynamic behavior of HIV-1 TAR and its complex with argininamide is investigated by means of molecular dynamics simulations starting from NMR structures, with explicit inclusion of water and periodic boundary conditions particle mesh Ewald representation of the electrostatic energy. During simulations of free and argininamide-bound TAR, local structural patterns, as determined by NMR experiments, were reproduced. An interdomain motion was observed in the simulations of free TAR, which is absent in the case of bound TAR, leading to the conclusion that the free conformation of TAR is intrinsically more flexible than the bound conformation. In particular, in the bound conformation the TAR–argininamide interface is very well ordered, as a result of the formation of a U·A·U base triple, which imposes structural constraints on the global conformation of the molecule. Free energy analysis, which includes solvation contributions, was used to evaluate the influence of van der Waals and electrostatic terms on formation of the complex and on the conformational rearrangement from free to bound TAR.     

Quasi-Monopolar Stimulation: A Novel Electrode Design Configuration for Performance Optimization of a Retinal Neuroprosthesis

In retinal neuroprostheses, spatial interaction between electric fields from various electrodes – electric crosstalk – may occur in multielectrode arrays during simultaneous stimulation of the retina. Depending on the electrode design and placement, this crosstalk can either enhance or degrade the functional characteristics of a visual prosthesis. To optimize the device performance, a balance must be satisfied between the constructive interference of crosstalk on dynamic range and power consumption and its negative effect on artificial visual acuity. In the present computational modeling study, we have examined the trade-off in these positive and negative effects using a range of currently available electrode array configurations, compared to a recently proposed stimulation strategy – the quasi monopolar (QMP) configuration – in which the return current is shared between local bipolar guards and a distant monopolar electrode. We evaluate the performance of the QMP configuration with respect to the implantation site and electrode geometry parameters. Our simulation results demonstrate that the beneficial effects of QMP are only significant at electrode-to-cell distances greater than the electrode dimensions. Possessing a relatively lower activation threshold, QMP was found to be superior to the bipolar configuration in terms of providing a relatively higher visual acuity. However, the threshold for QMP was more sensitive to the topological location of the electrode in the array, which may need to be considered when programming the manner in which electrode are simultaneously activated. This drawback can be offset with a wider dynamic range and lower power consumption of QMP. Furthermore, the ratio of monopolar return current to total return can be used to adjust the functional performance of QMP for a given implantation site and electrode parameters. We conclude that the QMP configuration can be used to improve visual information-to-stimulation mapping in a visual prosthesis, while maintaining low power consumption.     

A Proteinase K Inhibitor Using α,β-Unsaturated (Dehydro) Residues: A Presumptive Model

Abstract

Enzymes of the subtilisin family, of which proteinase K is a member, have been studied extensively on account of their numerous biological applications. Specific inhibitors of the proteinases are of immense importance in regulating their activity so as to protect the cells against uncontrolled proteolysis. Using the specific design principles of peptides containing dehydro-Alanine (ΔAla), generated by our theoretical calculations, we present here the design of an inhibitor of proteinase K. Energy minimization and molecular modeling of the interaction of the designed tetrapeptide with the recognition site of proteinase K indicate that it is an effective inhibitor.     

Saikosaponin-d,a novel SERCA inhibitor,induces autophagic cell death in apoptosis-defective cells

Autophagy is an important cellular process that controls cells in a normal homeostatic state by recycling nutrients to maintain cellular energy levels for cell survival via the turnover of proteins and damaged organelles. However, persistent activation of autophagy can lead to excessive depletion of cellular organelles and essential proteins, leading to caspase-independent autophagic cell death. As such, inducing cell death through this autophagic mechanism could be an alternative approach to the treatment of cancers. Recently, we have identified a novel autophagic inducer, saikosaponin-d (Ssd), from a medicinal plant that induces autophagy in various types of cancer cells through the formation of autophagosomes as measured by GFP-LC3 puncta formation. By computational virtual docking analysis, biochemical assays and advanced live-cell imaging techniques, Ssd was shown to increase cytosolic calcium level via direct inhibition of sarcoplasmic/endoplasmic reticulum Ca²⁺ ATPase pump, leading to autophagy induction through the activation of the Ca²⁺/calmodulin-dependent kinase kinase–AMP-activated protein kinase–mammalian target of rapamycin pathway. In addition, Ssd treatment causes the disruption of calcium homeostasis, which induces endoplasmic reticulum stress as well as the unfolded protein responses pathway. Ssd also proved to be a potent cytotoxic agent in apoptosis-defective or apoptosis-resistant mouse embryonic fibroblast cells, which either lack caspases 3, 7 or 8 or had the Bax-Bak double knockout. These results provide a detailed understanding of the mechanism of action of Ssd, as a novel autophagic inducer, which has the potential of being developed into an anti-cancer agent for targeting apoptosis-resistant cancer cells.     

Nanoformulation of curcumin protects HUVEC endothelial cells against ionizing radiation and suppresses their adhesion to monocytes: potential in prevention of radiation-induced atherosclerosis

Objectives

To investigated the potential of a novel dendrosomal nanoformulation of curcumin (DNC) in blocking radiation-induced changes in irradiated human umbilical vein endothelial cells (HUVECs), and their adhesion to human THP-1 monocytoid cells.

Results

Co⁶⁰ gamma rays reduced viability, raised the expression of adhesion molecules, ICAM-1, VCAM-1 and E-selectin (mRNA and protein), augmented the adhesion of THP-1 cells to HUVECs, activated NF-κB binding, increased the release of pro-inflammatory cytokines (IL-6, IL-8 and MCP-1) and induced oxidative damage (reduced glutathione declined, while 8-OHdG and TBARS increased). 5 µM DNC significantly inhibited these radiation-induced changes, activated the Nrf-2 pathway, and effectively suppressed THP-1 adhesion to HUVECs, implicating p38 MAPK signaling.

Conclusion

DNC treatment is a potential preventive method against inflammation and vascular damage from ionizing radiation.

     

Stage-specific expression of a Schistosoma mansoni polypeptide similar to the vertebrate regulatory protein stathmin

The ubiquitous vertebrate protein stathmin is expressed and phosphorylated in response to a variety of external and internal signals. Stathmin, in turn, controls cell growth and differentiation through its capacity to regulate microtubule assembly dynamics. This is the first report on the molecular cloning and characterization of a stathmin-like protein (SmSLP) in an invertebrate, the human blood fluke Schistosoma mansoni. SmSLP is first synthesized at high levels in the intermediate molluscan host and completely disappears 48 h after penetration into the mammalian host. The protein is preferentially iodinated in intact immature parasites using the Bolton-Hunter reagent, can be quantitatively extracted in high salt buffers, and remains soluble after boiling. Native SmSLP was partially sequenced, and its complete structure was derived from the cloning and sequencing of its cDNA. The sequence is up to 26% identical to vertebrate stathmin sequences and contains two potential phosphorylation sites. Native SmSLP is indeed phosphorylated because phosphatase digestion shifts its mobility in electrofocusing gels. SmSLP associates with tubulin, as suggested by immune co-precipitation results. In vitro experiments demonstrated that SmSLP inhibits tubulin assembly and causes the depolymerization of preassembled microtubules, thus probably fulfilling regulatory roles in critical steps of schistosome development.     

Risk-related standards of competence: continuing the debate over risk-related standards of competence

This discussion paper addresses Ian Wilks' defence of the risk-related standard of competence that appears in Bioethics 11 . Wilks there argues that the puzzle posed by Mark Wicclair in Bioethics 5 against Dan Brock's argument in favour of a risk-related standard of competence — namely that Brock's argument allows for situations of asymmetrical competence — is not a genuine problem for a risk-related standard of competence. To show this, Wilks presents what he believes to be two examples of real situations in which asymmetrical competence arises.
I argue that insofar as Wilks equivocates two senses of competence in his examples — namely, competence to perform a task and competence in performing a task — Wilks is unable to illustrate the existence of real situations of asymmetrical competence. By examining the way in which Wilks equivocates two senses of competence in his examples, and by applying the results of this examination to the problem of patient competency within the medical field, I argue that not only does Wilks fail to show that situations of asymmetrical competence exist, but he is also unable to provide a foundation for understating how the risk-related standard of competence can strike a balance between an individual's autonomy and benevolent intervention.
I thus conclude that insofar as Wilks fails to answer the objections raised by Wicclair and others against the risk-related standard of competence, the risk-related standard of competence continues to be undermined by the problem of asymmetrical competence.     

DNA detection by integrable electronics

This paper presents a new electronic methodology to detect DNA hybridization for rapid identification of diseases, as well as food and environmental monitoring on a genetic base. The proposed solution exploits a new (electrical) capacitive measurement circuit, not requiring any prior labeling of the DNA (as it is often the case with the commonly employed optical detection). The sensitivity, the reliability, and the reproducibility of this device have been evaluated by experiments performed with a (non-integrated) prototype implementation, easily integrable in IC and/or micro-fabricated lab-on-a-chip.