High-precision coding in visual cortex

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Underground Vendobionta From Namibia

The late Precambrian fossils from Namibia have generally been regarded as soft-bodied organisms whose three-dimensional preservation resulted from smothering in fluidized sand. The sedimentological context of Pteridinium and Namalia within a sandstone bed, however, allows us to distinguish between two taphocoenoses: (1) winnowed, laterally collapsed, current-transported specimens accumulated as a lag deposit of turbidite-like flows, and (2) specimens 'floating' in the top part of an event bed with their vanes extending upwards to the upper bedding surface. The second taphocoenosis is interpreted as an in situ preserved 'infaunal' community. The immobile underground life habit and the bizarre modes of growth of Pteridinium and Namalia do not fit any extinct or modern group of multicellular organisms. Similar statements can be made for Ernietta and Rangea , thus reviving the Vendobionta hypothesis.     

Peptidomimetic glutathione analogues as novel gammaGT stable GST inhibitors.

Elevated levels of glutathione-S-transferase (GST) isoenzymes are found in many tumor cells and are thought to play a role in the onset of multidrug resistance (MDR). To evaluate the contribution of GST to this process, inhibitors are needed. Glutathione (GSH) conjugates, although good GST inhibitors, cannot be used in vivo, because they are eliminated rapidly. In this paper, we describe the synthesis of a series of novel peptidomimetic glutathione analogues that are stabilized against peptidase mediated breakdown. The peptide bonds in GSH were replaced by isosteres, such as the 'reduced' amide (which was prepared using a novel method), N-methylamide, urethane, and methylene linkages. The in vitro evaluation of the compounds focuses on GST inhibition and stability towards gamma-glutamyl-transpeptidase (gammaGT), the main enzyme involved in GSH breakdown. The compounds were conjugated to the model electrophile ethacrynic acid (EA) to resemble GS-EA, an efficient GST inhibitor. All novel GSH-analogues were shown to inhibit rat liver cytosolic GSTs. Furthermore, peptidomimetic changes of the gamma-glutamyl-cysteine amide bond greatly improved stability towards gammaGT. These compounds may therefore be useful in the design of novel in vivo applicable GST inhibitors.     

Directed mutagenesis of Mycobacterium smegmatis 16S rRNA to reconstruct the in vivo evolution of aminoglycoside resistance in Mycobacterium tuberculosis

Drug resistance in Mycobacterium tuberculosis is a global problem, with major consequences for treatment and public health systems. As the emergence and spread of drug‐resistant tuberculosis epidemics is largely influenced by the impact of the resistance mechanism on bacterial fitness, we wished to investigate whether compensatory evolution occurs in drug‐resistant clinical isolates of M. tuberculosis. By combining information from molecular epidemiology studies of drug‐resistant clinical M. tuberculosis isolates with genetic reconstructions and measurements of aminoglycoside susceptibility and fitness in Mycobacterium smegmatis, we have reconstructed a plausible pathway for how aminoglycoside resistance develops in clinical isolates of M. tuberculosis. Thus, we show by reconstruction experiments that base changes in the highly conserved A‐site of 16S rRNA that: (i) cause aminoglycoside resistance, (ii) confer a high fitness cost and (iii) destabilize a stem‐loop structure, are associated with a particular compensatory point mutation that restores rRNA secondary structure and bacterial fitness, while maintaining to a large extent the drug‐resistant phenotype. The same types of resistance and associated mutations can be found in M. tuberculosis in clinical isolates, suggesting that compensatory evolution contributes to the spread of drug‐resistant tuberculosis disease.     

A Large-Scale Allosteric Transition in Cytochrome P450 3A4 Revealed by Luminescence Resonance Energy Transfer (LRET)

Effector-induced allosteric transitions in cytochrome P450 3A4 (CYP3A4) were investigated by luminescence resonance energy transfer (LRET) between two SH-reactive probes attached to various pairs of distantly located cysteine residues, namely the double-cysteine mutants CYP3A4(C64/C468), CYP3A4(C377/C468) and CYP3A4(C64/C121). Successive equimolar labeling of these proteins with the phosphorescent probe erythrosine iodoacetamide (donor) and the near-infrared fluorophore DY-731 maleimide (acceptor) allowed us to establish donor/acceptor pairs sensitive to conformational motions. The interactions of all three double-labeled mutants with the allosteric activators α-naphthoflavone and testosterone resulted in an increase in the distance between the probes. A similar effect was elicited by cholesterol. These changes in distance vary from 1.3 to 8.5 Å, depending on the position of the donor/acceptor pair and the nature of the effector. In contrast, the changes in the interprobe distance caused by such substrates as bromocriptine or 1-pyrenebutanol were only marginal. Our results provide a decisive support to the paradigm of allosteric modulation of CYP3A4 and indicate that the conformational transition caused by allosteric effectors increases the spatial separation between the beta-domain of the enzyme (bearing residues Cys₆₄ and Cys₃₇₇) and the alpha-domain, where Cys₁₂₁ and Cys₄₆₈ are located.     

Seeded Synthesis of CdSe/CdS Rod and Tetrapod Nanocrystals

We demonstrate a method for the synthesis of multicomponent nanostructures consisting of CdS and CdSe with rod and tetrapod morphologies. A seeded synthesis strategy is used in which spherical seeds of CdSe are prepared first using a hot-injection technique. By controlling the crystal structure of the seed to be either wurtzite or zinc-blende, the subsequent hot-injection growth of CdS off of the seed results in either a rod-shaped or tetrapod-shaped nanocrystal, respectively. The phase and morphology of the synthesized nanocrystals are confirmed using X-ray diffraction and transmission electron microscopy, demonstrating that the nanocrystals are phase-pure and have a consistent morphology. The extinction coefficient and quantum yield of the synthesized nanocrystals are calculated using UV-Vis absorption spectroscopy and photoluminescence spectroscopy. The rods and tetrapods exhibit extinction coefficients and quantum yields that are higher than that of the bare seeds. This synthesis demonstrates the precise arrangement of materials that can be achieved at the nanoscale by using a seeded synthetic approach.     

A statistical approach for detecting genomic aberrations in heterogeneous tumor samples from single nucleotide polymorphism genotyping data

We describe a statistical method for the characterization of genomic aberrations in single nucleotide polymorphism microarray data acquired from cancer genomes. Our approach allows us to model the joint effect of polyploidy, normal DNA contamination and intra-tumour heterogeneity within a single unified Bayesian framework. We demonstrate the efficacy of our method on numerous datasets including laboratory generated mixtures of normal-cancer cell lines and real primary tumours.     

Isolation of Rare Tumor Cells from Blood Cells with Buoyant Immuno-Microbubbles

Circulating tumor cells (CTCs) are exfoliated at various stages of cancer, and could provide invaluable information for the diagnosis and prognosis of cancers. There is an urgent need for the development of cost-efficient and scalable technologies for rare CTC enrichment from blood. Here we report a novel method for isolation of rare tumor cells from excess of blood cells using gas-filled buoyant immuno-microbubbles (MBs). MBs were prepared by emulsification of perfluorocarbon gas in phospholipids and decorated with anti-epithelial cell adhesion molecule (EpCAM) antibody. EpCAM-targeted MBs efficiently (85%) and rapidly (within 15 minutes) bound to various epithelial tumor cells suspended in cell medium. EpCAM-targeted MBs efficiently (88%) isolated frequent tumor cells that were spiked at 100,000 cells/ml into plasma-depleted blood. Anti-EpCAM MBs efficiently (>77%) isolated rare mouse breast 4T1, human prostate PC-3 and pancreatic cancer BxPC-3 cells spiked into 1, 3 and 7 ml (respectively) of plasma-depleted blood. Using EpCAM targeted MBs CTCs from metastatic cancer patients were isolated, suggesting that this technique could be developed into a valuable clinical tool for isolation, enumeration and analysis of rare cells.