Macromolecular interaction on a cAMP responsive region in the urokinase-type plasminogen activator gene: a role of protein phosphorylation.

We have studied the regulation of urokinase-type plasminogen activator gene expression by cAMP in LLC-PK1 cells. We found a cAMP responsive region 3.4 kb upstream of the transcription initiation site, which comprised three protein-binding domains designated A, B, and C. Domains A and B both contain a sequence, TGACG, homologous to a consensus cAMP response element (CRE; TGACGTCA). Effective cAMP-mediated induction was achieved when these two domains were linked with domain C, which by itself did not confer cAMP responsiveness to a heterologous promoter nor contained CRE-like sequence, suggesting a functional cooperation among these domains. Results of competition studies using gel retardation and DNase I footprinting assays suggest that there is a protein-protein interaction between a CRE binding protein and a domain C binding protein. In gel retardation assays, binding of a nuclear protein to domains A and B was strongly augmented by addition of the catalytic subunit of cAMP-dependent protein kinase, whereas the protein binding to domain C was slightly inhibited, suggesting that protein phosphorylation is involved in the regulation of protein-DNA interaction

Effect of Si-doping on InAs nanowire transport and morphology

The effect of Si-doping on the morphology, structure, and transport properties of nanowires was investigated. The nanowires were deposited by selective-area metal organic vapor phase epitaxy in an N-2 ambient. It is observed that doping systematically affects the nanowire morphology but not the structure of the nanowires. However, the transport properties of the wires are greatly affected. Room-temperature four-terminal measurements show that with an increasing dopant supply the conductivity monotonously increases. For the highest doping level the conductivity is higher by a factor of 25 compared to only intrinsically doped reference nanowires. By means of back-gate field-effect transistor measurements it was confirmed that the doping results in an increased carrier concentration. Temperature dependent resistance measurements reveal, for lower doping concentrations, a thermally activated semiconductor-type increase of the conductivity. In contrast, the nanowires with the highest doping concentration show a metal-type decrease of the resistivity with decreasing temperature. (C) 2011 American Institute of Physics. [doi: 10.1063/1.3631026

Multiple nuclear factors interact with promoter sequences of the urokinase-type plasminogen activator gene.

To characterize proteins that bind to the cyclic AMP inducible promoter of the urokinase-type plasminogen activator gene, we performed a DNAase I footprinting analysis. Within 500 nucleotides upstream of the transcription start site we found eight protected regions due to at least four different binding proteins. Among these is a single binding site for the transcription factor CTF/NF1, which is flanked on each side by two conserved binding sites for the transcription factor Sp1. A region at -380, which shares a similarity with sequences observed in the corresponding regions of other cyclic AMP regulated genes, was protected. This binding site contains a sequence of ten nucleotides which is repeated further upstream at -480 and also protected against DNAase I digestion. Comparisons of extracts from four different cell lines revealed that all DNA binding factors are present in nuclei of uPA expressing and nonexpressing cells. Mechanism underlying hormonal regulation of the gene is discussed

Magnetic and electrokinetic manipulations on a microchip device for bead-based immunosensing applications

9 páginas, 5 figuras.The combination of electrophoretic and magnetic manipulations with electrochemical detection for a versatile microfluidic and bead-based biosensing application is demonstrated. Amperometric detection is performed in an off-channel setup by means of a voltammetric cell built at the microchannel outlet and using a gold working electrode. Superparamagnetic particles are introduced and handled inside the channel by means of an external permanent magnet in combination with the electrogenerated flow which allows reproducible loading. The specific detection of phenol as electroactive alkaline phosphatase product is used in this study as proof of concept for a sensitive protein quantification. Characterizations and optimization of different parameters have been carried out in order to achieve the best detection signal. The applicability of the device has been finally demonstrated by the detection of rabbit IgG as model protein after an immunoassay performed on magnetic particles as immobilization platform. A comparison between the electrochemical detection using the developed device and the optical standard detection revealed similar performances with, however, extremely lower amount of reagent used and shorter analysis time. The developed electrophoretic- and magnetic-based chip may open the way to several other biosensing applications with interest not only for other proteins but also for DNA analysis, cell counting, and environmental control.The authors acknowledge funding from the ‘‘Ministerio de Ciencia e Innovación’’ (Madrid, Spain) for Projects MAT 2008-03079/NAN and CSD2006-00012 ‘‘NANOBIOMED’’ (Consolider-Ingenio 2010).Peer reviewe

On the influence of gas inlet configuration with respect to homogeneity in a horizontal single wafer reactor

In this paper two different gas inlet modifications for a horizontal single-wafer MOVPE reactor were studied: the conventional one ensured gas mixing of groups III and V precursors by forcibly merging both gases in a gas mixing unit, the newer one allowed mixing of groups III and V precursors solely by diffusion after the separation plate. AlAs/ GaAs Bragg reflector structures were deposited in nitrogen atmosphere for both gas inlet types and the uniformity determined by X-ray diffraction. Three-dimensional detailed numerical modeling was employed to explain the results. They establish that the observed strong non-uniformity of growth for the new inlet as well as the unusual concave instead of the common convex growth profile found is due to the incomplete mixing of precursor gases in a highly dense gas phase. By adjusting the groups III/V gas stream ratio an effect could be taken on the diffusion processes. Furthermore the total flow in the reactor was optimized for the new gas inlet. By taking advantage of changed diffusion properties in nitrogen atmosphere, however, perfect uniformity with an absolute deviation smaller than 1% was obtained. The modeling results are presented and agree very well with the experimental data. (C) 2001 Elsevier Science B.V. All rights reserved