Genomic organization of the Neurospora crassa gsn gene: possible involvement of the STRE and HSE elements in the modulation of transcription during heat shock

Glycogen synthase, an enzyme involved in glycogen biosynthesis, is regulated by phosphorylation and by the allosteric ligand glucose-6-phosphate (G6P). In addition, enzyme levels can be regulated by changes in gene expression. We recently cloned a cDNA for glycogen synthase (gsn) from Neurospora crassa, and showed that gsn transcription decreased when cells were exposed to heat shock (shifted from 30degreesC to 45degreesC). In order to understand the mechanisms that control gsn expression, we isolated the gene, including its 5' and 3' flanking regions, from the genome of N. crassa. An ORF of approximately 2.4 kb was identified, which is interrupted by four small introns (II-V). Intron I (482 bp) is located in the 5'UTR region. Three putative Transcription Initiation Sites (TISs) were mapped, one of which lies downstream of a canonical TATA-box sequence (5'-TGTATAAA-3'). Analysis of the 5'-flanking region revealed the presence of putative transcription factor-binding sites, including Heat Shock Elements (HSEs) and STress Responsive Elements (STREs). The possible involvement of these motifs in the negative regulation of gsn transcription was investigated using Electrophoretic Mobility Shift Assays (EMSA) with nuclear extracts of N. crassa mycelium obtained before and after heat shock, and DNA fragments encompassing HSE and STRE elements from the 5'-flanking region. While elements within the promoter region are involved in transcription under heat shock, elements in the 5'UTR intron may participate in transcription during vegetative growth. The results thus suggest that N. crassa possesses trans-acting elements that interact with the 5'-flanking region to regulate gsn transcription during heat shock and vegetative growth.

On non-linear dynamics and a periodic control design applied to the potential of membrane action

The Fitzhugh-Nagumo (fn) mathematical model characterizes the action potential of the membrane. The dynamics of the Fitzhugh-Nagumo model have been extensively studied both with a view to their biological implications and as a test bed for numerical methods, which can be applied to more complex models. This paper deals with the dynamics in the (FH) model. Here, the dynamics are analyzed, qualitatively, through the stability diagrams to the action potential of the membrane. Furthermore, we also analyze quantitatively the problem through the evaluation of Floquet multipliers. Finally, the nonlinear periodic problem is controlled, based on the Chebyshev polynomial expansion, the Picard iterative method and on Lyapunov-Floquet transformation (L-F transformation).

Linear readout of dynamic phase modulation index in an optical voltage sensor using the new J(1) ... J(3) spectral method

The J(1)...J(3) is a recent optical method for linear readout of dynamic phase modulation index in homodyne interferometers. In this work, the J(1)... J(3) method is applied to measure voltage in an optical voltage sensor. Based on the classical J(1)...J(4) method, the J(1)... J(3) technique shows to be more stable to phase drift and simpler for implementation than the original one. The sensor dynamic range is enhanced. The agreement between theoretical and experimental results, based on 1/f noise, is demonstrated.

CHOLESTEROL MODULATION OF LIPID PROTEIN INTERACTIONS IN LIVER MICROSOMAL MEMBRANE - A SPIN LABEL STUDY

ESR spectra of spin probes were used to monitor lipid-protein interactions in native and cholesterol-enriched microsomal membranes. In both systems composite spectra were obtained, one characteristic of bulk bilayer organization and another due to a motionally restricted population, which was ascribed to lipids in a protein microenvironment. Computer spectral subtractions revealed that cholesterol modulates the order/mobility of both populations in opposite ways, i.e., while the lipid bilayer region gives rise to more anisotropic spectra upon cholesterol enrichment, the spectra of the motionally restricted population become indicative of increased mobility and/or decreased order. These events were evidenced by measurement of both effective order parameters and correlation times. The percentages of the motionally restricted component were invariant in native and cholesterol-enriched microsomes. Variable temperature studies also indicated a lack of variation of the percentages of both spectral components, suggesting that the motionally restricted one was not due to protein aggregation. The results correlate well with the effect of cholesterol enrichment on membrane-bound enzyme kinetics and on the behavior of fluorescent probes [Castuma & Brenner (1986) Biochemistry 25, 4733-4738]. Several hypothesis are put forward to explain the molecular mechanism of the cholesterol-induced spectral changes.

Titanocene modulation of cytokine imbalance induced by Ehrlich ascites tumour progression

In the present work, we have studied the effects of two titanocenes, biscyclopentadienyldichlorotitanium IV (DDCT) and its derivative, biscyclopentadienylditiocianatetitanium IV (BCDT), on the production of cytokines [interferon-gamma (IFN-gamma), interelukin-1, interleukin (IL) 2, IL-4, and IL-10] by concanavalin A (Con A)-stimulated T cells obtained from Ehrlich ascites tumour (EAT)-bearing BALB/c mice. The treatment consisted of intraperitoneal (i.p) administration of 15 mg/kg/day DDCT for 2 days or 10 mg/kg/day BCDT for 3 days. We observed that the levels of IFN-gamma, but not IL-2, were dramatically increased in the early phase of EAT development. With tumour evolution, however, a sharp and progressive decrease in the levels of both IFN-gamma and IL-2 was found concomitantly to an enhancement in the levels of IL-10. Treatment of these mice with both titanocene compounds demonstrated that DDCT is more effective in modulating the cytokine imbalance induced by the tumour since it could prevent the early enhancement of IFN-gamma, the late decline of IFN-gamma and IL-2, and the increase in the IL-10. The administration of BCDT, in spite of preventing early IFN-gamma enhancement and increase in IL-10, did not produce any change in the IL-2 levels and did not prevent the decline of IFN-gamma levels during tumour evolution. Collectively, these results reveal that the ability of titanocenes to reverse tumour-induced immunosuppression and delay tumour growth is more evident in the DDCT compound, thus indicating that the substitution of the halides halogens by pseudohalogens, present in the molecular structure of BCDT, leads to a less effective antitumoral compound. (C) 2004 Elsevier B.V. All rights reserved.

Changeover during in situ compositional modulation of hexacyanoferrate (Prussian Blue) material

This paper describes the importance of (H2O)(6) clusters in controlling the properties of hexacyanoferrate (Prussian Blue) materials. A careful in situ study of compositional changes by using electrogravimetric techniques (in ac and dc modes) in hexacyanoferrates containing K+ alkali metals reveals the existence of a changeover in the properties of these films in a narrow potential range. Control of the compositional variation of the changeover is dependent on the K+ stoichiometric number in the compound structure. However, a specific K+ occupation in the compound structure activates the occupation of the (H2O)(6) cluster by H3O+ and/or H+, causing the changeover in the properties of hexacyanoferrate film. Thus, the information thus obtained is very useful for understanding the mechanisms involved in the electrochemical reversible switch between ferrimagnetism/paramagnetism, semiconductor/metal and electroluminescence/nonelectroluminescence properties of molecular cyanide materials.

Immune modulation induced by tuberculosis DNA vaccine protects non-obese diabetic mice from diabetes progression

We have described previously the prophylactic and therapeutic effect of a DNA vaccine encoding the Mycobacterium leprae 65 kDa heat shock protein (DNA-HSP65) in experimental murine tuberculosis. However, the high homology of this protein to the corresponding mammalian 60 kDa heat shock protein (Hsp60), together with the CpG motifs in the plasmid vector, could trigger or exacerbate the development of autoimmune diseases. The non-obese diabetic (NOD) mouse develops insulin-dependent diabetes mellitus (IDDM) spontaneously as a consequence of an autoimmune process that leads to destruction of the insulin-producing beta cells of the pancreas. IDDM is characterized by increased T helper 1 (Th1) cell responses toward several autoantigens, including Hsp60, glutamic acid decarboxylase and insulin. In the present study, we evaluated the potential of DNA-HSP65 injection to modulate diabetes in NOD mice. Our results show that DNA-HSP65 or DNA empty vector had no diabetogenic effect and actually protected NOD mice against the development of severe diabetes. However, this effect was more pronounced in DNA-HSP65-injected mice. The protective effect of DNA-HSP65 injection was associated with a clear shift in the cellular infiltration pattern in the pancreas. This change included reduction of CD4(+) and CD8(+) T cells infiltration, appearance of CD25(+) cells influx and an increased staining for interleukin (IL)-10 in the islets. These results show that DNA-HSP65 can protect NOD mice against diabetes and can therefore be considered in the development of new immunotherapeutic strategies.

Periodic collisions between the moon Prometheus and Saturn's F ring

Saturn's F ring, which lies 3,400 km beyond the edge of the main ring system, was discovered by the Pioneer 11 spacecraft(1) in 1979. It is a narrow, eccentric ring which shows an unusual 'braided' appearance in several Voyager 1 images' obtained in 1980, although it appears more regular in images from Voyager 2 obtained nine months later(3). The discovery of the moons Pandora and Prometheus orbiting on either side of the ring provided a partial explanation for some of the observed features(4). Recent observations of Prometheus(5,6) by the Hubble Space Telescope show, surprisingly, that it is lagging behind its expected position by similar to 20 degrees. By modelling the dynamical evolution of the entire Prometheus-F ring-Pandora system, we show here that Prometheus probably encountered the core of the F ring in 1994 and that it may still be entering parts of the ring once per orbit. Collisions with objects in the F ring provide a plausible explanation for the observed lag and imply that the mass of the F ring is probably less than 25% that of Prometheus.

Structural and electronic properties of PbTiO3 slabs: a DFT periodic study

Structural and electronic properties of the bulk and relaxed surfaces (TiO2 and PbO terminated) of cubic PbTiO3 are investigated by means of periodic quantum-mechanical calculations based on density functional theory. It is observed that the difference in surface energies is small and relaxations effects are most prominent for Ti and Ph surface atoms. The electronic structure shows a splitting of the lowest conduction bands for the TiO2 terminated surface and of the highest valence bands for the PbO terminated slab. The calculated indirect band gap is: 3.18, 2.99 and 3.03 eV for bulk, TiO2 and PbO terminations, respectively. The electron density maps show that the Ti-O bond has a partial covalent character, whereas the Pb-O bonds present a very low covalency. (C) 2004 Elsevier B.V. All rights reserved.

Modulation of charge-density waves by superlattice structures

We discuss the interplay between electronic correlations and an underlying superlattice structure in determining the period of charge density waves (CDW's), by considering a one-dimensional Hubbard model with a repeated (nonrandom) pattern of repulsive (U > 0) and free (U=0) sites. Density matrix renormalization group diagonalization of finite systems (up to 120 sites) is used to calculate the charge-density correlation function and structure factor in the ground state. The modulation period can still be predicted through effective Fermi wave vectors k(F)(*) and densities, and we have found that it is much more sensitive to electron (or hole) doping, both because of the narrow range of densities needed to go from q(*)=0 to pi, but also due to sharp 2k(F)(*)-4k(F)(*) transitions; these features render CDW's more versatile for actual applications in heterostructures than in homogeneous systems.

Non-periodic bifurcations of one-dimensional maps

We prove that a 'positive probability' subset of the boundary of '{uniformly expanding circle transformations}' consists of Kupka-Smale maps. More precisely, we construct an open class of two-parameter families of circle maps (f(alpha,theta))(alpha,theta) such that, for a positive Lebesgue measure subset of values of alpha, the family (f(alpha,theta))(theta) crosses the boundary of the uniformly expanding domain at a map for which all periodic points are hyperbolic (expanding) and no critical point is pre-periodic. Furthermore, these maps admit an absolutely continuous invariant measure. We also provide information about the geometry of the boundary of the set of hyperbolic maps.

Modulation of intercellular communication in macrophages: possible interactions between GAP junctions and P2 receptors

Gap junctions are connexin-formed channels that play an important role in intercellular communication in most cell types. In the immune system, specifically in macrophages, the expression of connexins and the establishment of functional gap junctions are still controversial issues. Macrophages express P2X(7) receptors that, once activated by the binding of extracellular ATP, lead to the opening of transmembrane pores permeable to molecules of up to 900 Da. There is evidence suggesting an interplay between gap junctions and P2 receptors in different cell systems. Thus, we used ATP-sensitive and -insensitive J774.G8 macrophage cell lines to investigate this interplay. To study junctional communication in J774-macrophage-like cells, we assessed cell-to-cell communication by microinjecting Lucifer Yellow. Confluent cultures of ATP-sensitive J774 cells (ATP-s cells) are coupled, whereas ATP-insensitive J774 cells (ATP-i cells), derived by overexposing J774 cells to extracellular ATP until they do not display the phenomenon of ATP-induced permeabilization, are essentially uncoupled. Western-blot and reverse-transcription polymerase chain reaction assays revealed that ATP-s and ATP-i cells express connexin43 (Cx43), whereas only ATP-s cells express the P2X(7) receptor. Accordingly, ATP-i cells did not display any detectable ATP-induced current under whole-cell patch-clamp recordings. Using immunofluorescence microscopy, Cx43 reactivity was found at the cell surface and in regions of cell-cell contact of ATP-s cells, whereas, in ATP-i cells, Cx43 immunoreactivity was only present in cytosolic compartments. Using confocal microscopy, it is shown here that, in ATP-s cells as well as in peritoneal macrophages, Cx43 and P2X(7) receptors are co-localized to the membrane of ATP-s cells and peritoneal macrophages.

Band-edge states of the zero(th)-order gap in quasi-periodic photonic superlattices

The photonic modes of Thue-Morse and Fibonacci lattices with generating layers A and B, of positive and negative indices of refraction, are calculated by the transfer-matrix technique. For Thue-Morse lattices, as well for periodic lattices with AB unit cell, the constructive interference of reflected waves, corresponding to the zero(th)-order gap, takes place when the optical paths in single layers A and B are commensurate. In contrast, for Fibonacci lattices of high order, the same phenomenon occurs when the ratio of those optical paths is close to the golden ratio. In the long wavelength limit, analytical expressions defining the edge frequencies of the zero(th) order gap are obtained for both quasi-periodic lattices. Furthermore, analytical expressions that define the gap edges around the zero(th) order gap are shown to correspond to the = 0 and = 0 conditions.

Serotonergic mechanisms on breathing modulation in the rat locus coeruleus

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)