bluuebunny/crossref_metadata_2025 huggingface dataset:
Three-dimensional Culture Regulates Raf-1 Expression to Modulate Fibronectin Matrix Assembly
Authors: Winters B. S. , Raj B. K. Mohan , Robinson E. E. , Foty R. A. , Corbett S. A.
Published: May 2006

Oncogenic transformation has been associated with decreased fibronectin (FN) matrix assembly. For example, both the HT-1080 fibrosarcoma and MAT-LyLu cell lines fail to assemble a FN matrix when grown in monolayer culture (2-dimensional [2D] system). In this study, we show that these cells regain the ability to assemble a FN matrix when they are grown as aggregates (3-dimensional [3D] system). FN matrix assembly in 3D correlates with decreased Raf-1 protein expression compared with cells grown in monolayer culture. This effect is associated with reduced Raf-1 mRNA levels as determined by quantitative RT-PCR and not proteasome-mediated degradation of endogenous Raf-1. Interestingly, transient expression of a Raf-1 promoter-reporter construct demonstrates increased Raf-1 promoter activity in 3D, suggesting that the transition to 3D culture may modulate Raf-1 mRNA stability. Finally, to confirm that decreased Raf-1 expression results in increased FN matrix assembly, we used both pharmacological and small interfering RNA knockdown of Raf-1. This restored the ability of cells in 2D culture to assemble a FN matrix. Moreover, overexpression of Raf-1 prevented FN matrix assembly by cells cultured in 3D, resulting in decreased aggregate compaction. This work provides new insight into how the cell microenvironment may influence Raf-1 expression to modulate cell–FN interactions in 3D.
The Detection of Micromolar Pericellular ATP Pool on Lymphocyte Surface by Using Lymphoid Ecto-Adenylate Kinase as Intrinsic ATP Sensor
Authors: Yegutkin Gennady G. , Mikhailov Andrey , Samburski Sergei S. , Jalkanen Sirpa
Published: May 2006

Current models of extracellular ATP turnover include transient release of nanomolar ATP concentrations, triggering of signaling events, and subsequent ectoenzymatic inactivation. Given the high substrate specificity for adenylate kinase for reversible reaction (ATP + AMP ↔ 2ADP), we exploited lymphoid ecto-adenylate kinase as an intrinsic probe for accurate sensing pericellular ATP. Incubation of leukemic T- and B-lymphocytes with [3H]AMP or [α-32P]AMP induces partial nucleotide conversion into high-energy phosphoryls. This “intrinsic” AMP phosphorylation occurs in time- and concentration-dependent fashions via nonlytic supply of endogenous γ-phosphate–donating ATP, remains relatively resistant to bulk extracellular ATP scavenging by apyrase, and is diminished after lymphocyte pretreatment with membrane-modifying agents. This enzyme-coupled approach, together with confocal imaging of quinacrine-labeled ATP stores, suggests that, along with predominant ATP accumulation within cytoplasmic granules, micromolar ATP concentrations are constitutively retained on lymphoid surface without convection into bulk milieu. High basal levels of inositol phosphates in the cells transfected with ATP-selective human P2Y2-receptor further demonstrate that lymphocyte-surrounding ATP is sufficient for triggering purinergic responses both in autocrine and paracrine fashions. The ability of nonstimulated lymphocytes to maintain micromolar ATP halo might represent a novel route initiating signaling cascades within immunological synapses and facilitating leukocyte trafficking between the blood and tissues.
Targeting of Transmembrane Protein Shrew-1 to Adherens Junctions Is Controlled by Cytoplasmic Sorting Motifs
Authors: Jakob Viktor , Schreiner Alexander , Tikkanen Ritva , Starzinski-Powitz Anna
Published: May 2006

We recently identified transmembrane protein shrew-1 and showed that it is able to target to adherens junctions in polarized epithelial cells. This suggested shrew-1 possesses specific basolateral sorting motifs, which we analyzed by mutational analysis. Systematic mutation of amino acids in putative sorting signals in the cytoplasmic domain of shrew-1 revealed three tyrosines and a dileucine motif necessary for basolateral sorting. Substitution of these amino acids leads to apical localization of shrew-1. By applying tannic acid to either the apical or basolateral part of polarized epithelial cells, thereby blocking vesicle fusion with the plasma membrane, we obtained evidence that the apically localized mutants were primarily targeted to the basolateral membrane and were then redistributed to the apical domain. Further support for a postendocytic sorting mechanism of shrew-1 was obtained by demonstrating that μ1B, a subunit of the epithelial cell-specific adaptor complex AP-1B, interacts with shrew-1. In conclusion, our data provide evidence for a scenario where shrew-1 is primarily delivered to the basolateral membrane by a so far unknown mechanism. Once there, adaptor protein complex AP-1B is involved in retaining shrew-1 at the basolateral membrane by postendocytic sorting mechanisms.
Distinct Roles for Tsg101 and Hrs in Multivesicular Body Formation and Inward Vesiculation
Authors: Razi M. , Futter C. E.
Published: May 2006

In mammalian cells, epidermal growth factor (EGF) stimulation promotes multivesicular body (MVB) formation and inward vesiculation within MVB. Annexin 1 is required for EGF-stimulated inward vesiculation but not MVB formation, demonstrating that MVB formation (the number of MVBs/unit cytoplasm) and inward vesiculation (the number of internal vesicles/MVB) are regulated by different mechanisms. Here, we show that EGF-stimulated MVB formation requires the tumor susceptibility gene, Tsg101, a component of the ESCRT (endosomal sorting complex required for transport) machinery. Depletion of Tsg101 potently inhibits EGF degradation and MVB formation and causes the vacuolar domains of the early endosome to tubulate. Although Tsg101 depletion inhibits MVB formation and alters the morphology of the early endosome in unstimulated cells, these effects are much greater after EGF stimulation. In contrast, depletion of hepatocyte growth factor receptor substrate (Hrs) only modestly inhibits EGF degradation, does not induce tubulation of the early endosome, and causes the generation of enlarged MVBs that retain the ability to fuse with the lysosome. Together, these results indicate that Tsg101 is required for the formation of stable vacuolar domains within the early endosome that develop into MVBs and Hrs is required for the accumulation of internal vesicles within MVBs and that both these processes are up-regulated by EGF stimulation.
Association of Bcl-2 with Misfolded Prion Protein Is Linked to the Toxic Potential of Cytosolic PrP
Authors: Rambold Angelika S. , Miesbauer Margit , Rapaport Doron , Bartke Till , Baier Michael , Winklhofer Konstanze F. , Tatzelt Jörg
Published: May 2006

Protein misfolding is linked to different neurodegenerative disorders like Alzheimer’s disease, polyglutamine, and prion diseases. We investigated the cytotoxic effects of aberrant conformers of the prion protein (PrP) and show that toxicity is specifically linked to misfolding of PrP in the cytosolic compartment and involves binding of PrP to the anti-apoptotic protein Bcl-2. PrP targeted to different cellular compartments, including the cytosol, nucleus, and mitochondria, adopted a misfolded and partially proteinase K–resistant conformation. However, only in the cytosol did the accumulation of misfolded PrP induce apoptosis. Apoptotic cell death was also induced by two pathogenic mutants of PrP, which are partially localized in the cytosol. A mechanistic analysis revealed that the toxic potential is linked to an internal domain of PrP (amino acids 115–156) and involves coaggregation of cytosolic PrP with Bcl-2. Increased expression of the chaperones Hsp70 and Hsp40 prevented the formation of PrP/Bcl-2 coaggregates and interfered with PrP-induced apoptosis. Our study reveals a compartment-specific toxicity of PrP misfolding that involves coaggregation of Bcl-2 and indicates a protective role of molecular chaperones.
PAR1b Promotes Cell–Cell Adhesion and Inhibits Dishevelled-mediated Transformation of Madin-Darby Canine Kidney Cells
Authors: Elbert Maya , Cohen David , Müsch Anne
Published: May 2006

Mammalian Par1 is a family of serine/threonine kinases comprised of four homologous isoforms that have been associated with tumor suppression and differentiation of epithelial and neuronal cells, yet little is known about their cellular functions. In polarizing kidney epithelial (Madin-Darby canine kidney [MDCK]) cells, the Par1 isoform Par1b/MARK2/EMK1 promotes the E-cadherin–dependent compaction, columnarization, and cytoskeletal organization characteristic of differentiated columnar epithelia. Here, we identify two functions of Par1b that likely contribute to its role as a tumor suppressor in epithelial cells. 1) The kinase promotes cell–cell adhesion and resistance of E-cadherin to extraction by nonionic detergents, a measure for the association of the E-cadherin cytoplasmic domain with the actin cytoskeleton, which is critical for E-cadherin function. 2) Par1b attenuates the effect of Dishevelled (Dvl) expression, an inducer of wnt signaling that causes transformation of epithelial cells. Although Dvl is a known Par1 substrate in vitro, we determined, after mapping the PAR1b-phosphorylation sites in Dvl, that PAR1b did not antagonize Dvl signaling by phosphorylating the wnt-signaling molecule. Instead, our data suggest that both proteins function antagonistically to regulate the assembly of functional E-cadherin–dependent adhesion complexes.
THE SOURCES OF GROWTH AT DIFFERENT STAGES OF DEVELOPMENT
Authors: OSBORNE EVAN
Published: May 2006

Cross‐country growth regressions have become an increasingly common tool in empirical development research. But these regressions typically do not attempt to distinguish among countries in different stages of development. Three empirical methods are used to test for such differences. Several of the factors known to affect economic growth are shown to operate differently for countries in different portions of the global income distribution. The results have implications for the role of financial markets, openness and factor accumulation in promoting growth. (JEL O150, O160, O400)
X-ray crystallographic structures of enamine and amine Schiff bases of pyridoxal and its 1:1 hydrogen-bonded complexes with benzoic acid derivatives: evidence for coupled inter- and intramolecular proton transfer
Authors: Sharif Shasad , Powell Douglas R. , Schagen David , Steiner Thomas , Toney Michael D. , Fogle Emily , Limbach Hans-Heinrich
Published: May 2006

Crystal structures of Schiff bases containing pyridoxal (PL), N-(pyridoxylidene)-tolylamine, C15H16N2O2 (I), N-(pyridoxylidene)-methylamine, C9H12N2O2 (III), and their 1:1 adduct with 2-nitrobenzoic acid, (I)+ C7H4NO_4^- (II), and 4-nitrobenzoic acid, (III)+ C7H4NO_4^- (IV), serve as models for the coenzyme pyridoxal-5′-phosphate (PLP) in its PLP-dependent enzymes. These models allow the study of the intramolecular OHN hydrogen bond of PL/PLP Schiff bases and the H-acceptor properties of their pyridine rings. The free base (I) forms hydrogen-bonded chains involving the hydroxyl side groups and the rings of adjacent molecules, whereas (III) forms related hydrogen-bonded cyclic dimers. The adducts (II)/(IV) consist of 1:1 hydrogen-bonded complexes, exhibiting strong intermolecular bonds between the carboxylic groups of the acids and the pyridine rings of (I)/(III). In conclusion, the proton in the intramolecular O—H...N hydrogen bond of (I)/(III) is located close to oxygen (enolamine form). The added acids protonate the pyridine ring in (II)/(IV), but only in the latter case does this protonation lead to a shift of the intramolecular proton towards the nitrogen (ketoimine form). All crystallographic structures were observed in the open form. In contrast, the formation of the pyridinium salt by dissolving (IV) leads to the cyclic aminal form.
S,S-1,2-Dicyclohexylethane-1,2-diol and its racemic compound: a striking exception to Wallach's rule
Authors: Patrick Brian O. , Brock Carolyn Pratt
Published: May 2006

The structures of enantiopure S,S-1,2-dicyclohexylethane-1,2-diol and its racemic compound (rac-S,S-1,2-dicyclohexylethane-1,2-diol) have been determined at 295 and 173 K. The crystals of the enantiopure material are more than 4% denser than the crystals of the racemic compound, but the melting points indicate that the crystals of the less dense racemic compound are considerably more stable than those of the racemic conglomerate. This apparent exception to the correlation of crystal density and melting point is explained. The enantiopure crystals have four molecules in the asymmetric unit (Z′ = 4). Two of the molecules have the conformation observed for the one independent molecule of the racemic compound and two have a higher energy conformation; the overall P21 structure is a perturbed version of a P212121 structure with Z′ = 2. The enantiopure and racemic crystals have the same hydrogen-bonding motif, but the motif in the former appears to be significantly strained. A reason why crystals of enantiopure material might be systematically less dense than crystals of its racemic compound and to be more likely to have Z′ > 1 is suggested.
Structure prediction of ordered and disordered multiple octahedral cation perovskites using SPuDS
Authors: Lufaso Michael W. , Barnes Paris W. , Woodward Patrick M.
Published: May 2006

The software package SPuDS has previously been shown to accurately predict crystal structures of AMX
3 and A
1 − x

A′
x

MX
3 perovskites that have undergone octahedral tilting distortions. This paper describes the extension of this technique and its accuracy for A
2
MM′X
6 ordered double perovskites with the aristotype Fm\overline 3m cubic structure, as well as those that have undergone octahedral tilting distortions. A survey of the literature shows that roughly 70% of all ordered double perovskites undergo octahedral tilting distortions. Of the 11 distinct types of octahedral tilting that can occur in ordered perovskites, five tilt systems account for ∼97% of the reported structures. SPuDS can calculate structures for the five dominant tilt systems, Fm\overline 3m (a
0
a
0
a
0), I4/m (a
0
a
0
c
−), R\overline 3 (a
−
a
−
a
−), I2/m (a
0
b
−
b
−) and P21/n (a
−
a
−
b
+), as well as two additional tilt systems, Pn\overline 3 (a
+
a
+
a
+) and P4/mnc (a
0
a
0
c
+). Comparison with reported crystal structures shows that SPuDS is quite accurate at predicting distortions driven by octahedral tilting. The favored modes of octahedral tilting in ordered double perovskites are compared and contrasted with those in AMX
3 perovskites. Unit-cell pseudosymmetry in Sr- and Ca-containing double perovskites is also examined. Experimentally, Sr2
MM′O6 compounds show a much stronger tendency toward pseudosymmetry than do Ca2
MM′O6 compounds with similar tolerance factors.
Cis/trans isomers of PtX
2
L
2 (X = halogen, L = neutral ligand); the crystal structure of trans-dichlorobis(dimethyl sulfide)platinum(II) and the pressure dependence of its unit-cell dimensions
Authors: Hansson Christian , Carlson Stefan , Giveen Deborah , Johansson Maria , Yong Shen , Oskarsson Åke
Published: May 2006

trans-PtCl2(dms)2 (dms is dimethyl sulfide) crystallizes in the space group P21/n and adopts the molecular point group C

i
, which is the most frequently occurring point group for trans-PtX
2
L
2 complexes (78%), as observed in the Cambridge Structural Database (CSD; 2005 release), followed by C
1 (16%). Density functional theory calculations show that the observed geometry for trans-PtCl2(dms)2 has slightly higher energy than the most favorable geometry in the point group C
2h
, but this geometry would require a space group that hampers close packing, thus showing that intermolecular forces determine the point group for the title compound. High-pressure powder diffraction studies of trans-PtCl2(dms)2 show no phase transformation up to 8.0 GPa. The bulk modulus is 8.1 (6) GPa and the pressure derivative 8.1 (4). In the CSD, the number of cis- and trans-PtX
2
L
2 compounds are almost equal, viz. 156 cis and 160 trans compounds, showing no preference for either isomer in the solid state.
Chemical bonding in pentaerythritol at very low temperature or at high pressure: an experimental and theoretical study
Authors: Zhurova Elizabeth A. , Tsirelson Vladimir G. , Zhurov Vladimir V. , Stash Adam I. , Pinkerton A. Alan
Published: May 2006

Chemical bonding in the pentaerythritol crystal based on the experimental electron density at 15 (1) K, and theoretical calculations at the experimental molecular geometries obtained at room and low (15 K) temperatures have been analyzed and compared in terms of the topological analysis. Topological electron-density features corresponding to the high-pressure (1.15 GPa) geometry are also reported. In addition to the bond critical points (CPs) within the molecular layers, CPs between the atoms of different molecular layers have been located and the bonding character of these relatively weak interactions discussed. Atomic charges and energies have been integrated over the atomic basins delimited by the zero-flux surfaces, and the intermolecular interaction energies have been calculated. The interaction between molecular layers in the crystal becomes stronger both at very low temperature and high pressure, as demonstrated by the more negative intermolecular interaction energies, higher electron density and energy density values at the CPs, and sharper electronic-energy density profiles.
The incommensurately modulated structure of a tricyclic natural-product-like compound of empirical formula C22H20O3
Authors: Guiblin Nicolas , Fuhrer Cyril A. , Häner Robert , Stoeckli-Evans Helen , Schenk Kurt , Chapuis Gervais
Published: May 2006

3,4-Diphenyl-2a,5a,6,7,8,8a,8b-heptahydro-furo[4,3,2-de]chromen-2-one (1) was prepared as part of a project aimed at the synthesis of polycyclic natural-product-like scaffolds. X-ray analysis of crystals grown from ethanol revealed an incommensurately modulated structure. Data include main and satellite reflections up to second order. The modulation vector was refined using the NADA program. The modulated character of the structure of the organic compound C22H20O3 is interpreted in terms of the intermolecular C—H...O hydrogen bonds and close-contact approximation.
Experimental versus expected halide-ion size differences; structural changes in three series of isotypic bismuth chalcogenide halides
Authors: Keller Egbert , Krämer Volker
Published: May 2006

Experimentally determined halide-ion size differences are compared with expected size differences in the three series of isotypic bismuth chalcogenide halide compounds, KBi6O9
X (X = Cl, Br and I), BiOX (X = F, Cl, Br and I) and BiSX (X = Cl, Br and I). The strong deviations observed can be assigned to steric strain caused by the heterogenity of the bond-valence pattern and (for BiOX) to anion–anion repulsion and a change in the connectivity scheme. Some special features of the BiOF structure and the question of `isotypism' within the BiOX series are briefly discussed. Structural changes within the BiSX series are analysed.
Estimation of polyhedral compressibilities and structural evolution of GdFeO3-type perovskites at high pressures
Authors: Zhao J. , Ross N. L. , Angel R. J.
Published: May 2006

A new approach based on the bond-valence matching relation is developed to predict the detailed structural evolution of GdFeO3-type perovskites at high pressure from knowledge of the room-pressure structure and the high-pressure unit-cell parameters alone. The evolution of perovskite structures estimated in this way is in good agreement with the structure refinements available from high-pressure single-crystal diffraction measurements of a number of perovskites. The method is then extended to predict the structure of MgSiO3 perovskite at pressures for which no single-crystal structural data are available and the results are compared to ab initio quantum calculations of MgSiO3 perovskite up to 120 GPa.
Crystal structures of the trifluoromethyl sulfonates M(SO3CF3)2 (M = Mg, Ca, Ba, Zn, Cu) from synchrotron X-ray powder diffraction data
Authors: Dinnebier Robert , Sofina Natalia , Hildebrandt Lars , Jansen Martin
Published: May 2006

The crystal structures of divalent metal salts of trifluoromethyl sulfonic acid (`trifluoromethyl sulfonates') M(SO3CF3)2 (M = Mg, Ca, Ba, Zn, Cu) were determined from high-resolution X-ray powder diffraction data. Magnesium, calcium and zinc trifluoromethyl sulfonate crystallize in the rhombohedral space group R\bar3. Barium trifluoromethyl sulfonate crystallizes in the monoclinic space group I2/a(C2/c) and copper trifluoromethyl sulfonate crystallizes in the triclinic group P\bar1. Within the crystal structures the trifluoromethyl sulfonate anions are arranged in double layers with the apolar CF3 groups pointing towards each other. The cations are located next to the SO3 groups. The symmetry relations between the different crystal structures have been analysed.
Structural complexity in minerals: twinning, polytypism and disorder in the crystal structure of polybasite, (Ag,Cu)16(Sb,As)2S11
Authors: Evain Michel , Bindi Luca , Menchetti Silvio
Published: May 2006

The crystal structures of 222- and 221-polybasite [(Ag,Cu)16(Sb,As)2S11] crystals have been solved and refined by means of X-ray diffraction data (collected at 100 and 120 K, respectively) from twinned crystals. Both structures consist of the stacking of [(Ag,Cu)6Sb2S7]2− and [Ag9CuS4]2+ module layers in which Sb forms isolated SbS3 pyramids typically occurring in sulfosalts; copper links two S atoms in a linear coordination and silver occupies sites with coordination ranging from quasi-linear to almost tetrahedral. An Ag → Cu substitution in the [(Ag,Cu)6Sb2S7]2− module layer is observed in both structures, the substitution amount being larger in the 221- than in the 222-polybasite. A pattern of the possible mechanism regulating the type of unit cell that is stabilized is proposed: starting from the hypothetical stoichiometric and fully ordered Ag15CuSb2S11 222-polybasite structure, with a low C2/c monoclinic symmetry and a large 222 supercell, the disorder introduced by the substitution of Cu for Ag increases the symmetry with a cell reduction along the c axis yielding the 221 supercell and a trigonal crystal system. A further increase of the substitution gives rise to a folding of the cell along the a and b axes and the 111-pearceite structure, space group P\bar3m1.
Compressibility of the nitridosilicate SrYb[Si4N7] and the oxonitridoaluminosilicates MYb[Si4−x
Al
x
O
x
N7−x] (x = 2; M = Sr, Ba)
Authors: Juarez-Arellano E. A. , Friedrich A. , Knorr K. , Lieb A. , Winkler B. , Amboage M. , Hanfland M. , Schnick W.
Published: May 2006

The compressibilities of the nitridosilicate SrYb[Si4N7] and the oxonitridoaluminosilicates MYb[Si4−x
Al
x
O
x
N7−x
] (x = 2; M = Sr, Ba) were investigated by in situ high-pressure X-ray powder diffraction. Pressures up to 42 GPa were generated using the diamond–anvil cell technique. The title compounds are structurally stable to the highest pressure obtained. A fit of a third-order Birch–Murnaghan equation-of-state to the p–V data results in V
0 = 302.91 (6) Å3, B
0 = 176 (2) GPa and B′ = 4.4 (2) for SrYb[Si4N7]; V
0 = 310.4 (1) Å3, B
0 = 161 (2) GPa and B′ = 4.6 (2) for SrYb[Si4−x
Al
x
O
x
N7−x
]; and V
0 = 317.3 (5) Å3, B
0 = 168 (2) GPa and B′ = 4.7 (2) for BaYb[Si4−x
Al
x
O
x
N7−x
]. While the linear compressibilities of the a and c axes of BaYb[Si4−x
Al
x
O
x
N7−x
] are very similar up to 30 GPa, distinct differences were observed for SrYb[Si4N7] and SrYb[Si4−x
Al
x
O
x
N7−x
], with the c axis being the most compressible axis. In all of the investigated compounds the bulk compressibility is dominated by the compression behaviour of the tetrahedral network, while the size of the substituted cation plays a minor role.
A comparative crystallochemical analysis of binary compounds and simple anhydrous salts containing pyramidal anions LO3 (L = S, Se, Te, Cl, Br, I)
Authors: Blatov V. A. , Peskov M. V.
Published: May 2006

A comparative analysis of binary compounds and 61 simple anhydrous salts M

y
(LO3)
z
(L = S, Se, Te, Cl, Br, I) was performed using the crystallochemical program package TOPOS. A topological similarity was found between the salts and six types of binary compounds (NaCl, NiAs, PoCl2, Tl2S2, ZnTe, rutile). It is notable that these structure relationships are typical for other groups of inorganic salts: borates, carbonates, nitrates, orthophosphates, orthoarsenates, sulfates, selenates, perchlorates, molybdates and halogenides of d-metals. For all the M

y
(LO3)
z
compounds the topology and uniformity of the ion arrays were investigated. It has been established that in 36 out of the 61 salts at least one ion array has the topology of close packing or the body-centred cubic lattice. The results obtained have allowed us to come to conclusions about the structure-forming role of the arrays of various chemical composition.
17 salts of ephedrine: crystal structures and packing analysis
Authors: Collier Edwin A. , Davey Roger J. , Black Simon N. , Roberts Ron J.
Published: May 2006

The structures of two neutral and 17 salt forms of the base (1R, 2S)-(−)-ephedrine are reported. These structures are discussed in the light of the conformers of the ephedrine moiety, the existence of bilayers and the structure determining role of the counterions. Overall, most of the salt structures are essentially derived from the observed packing of the neutral base and are dominated by the amphiphilic nature of the ephedrine molecular structure. In a few cases the size and hydrophobicity of the counterion disrupts this behaviour.
When are alternative stable states more likely to occur?
Authors: Didham Raphael K. , Norton David A.
Published: May 2006

T. Fukami and W. G. Lee argue that the logical expectation from ecological theory is that competitively‐structured assemblages will be more likely to exhibit alternative stable states than abiotically‐structured assemblages. We suggest that there are several important misinterpretations in their arguments, and that the substance of their hypothesis has both a weak basis in ecological theory and is not supported by empirical evidence which shows that alternative stable states occur more frequently in natural systems subject to moderate‐ to harsh abiotic extremes. While this debate is founded in ecological theory, it has important applied implications for restoration management. Sound theoretical predictions about when to expect alternative stable states can only aid more effective restoration if theoretical expectations can be shown to translate into predictable empirical outcomes. If strongly abiotically‐ or disturbance‐structured systems are more likely to exhibit catastrophic phase shifts in community structure that can be resilient to management efforts, then restoration ecologists will need to treat these systems differently in terms of the types of management inputs that are required.
A human DNA polymerase η complex containing Rad18, Rad6 and Rev1; proteomic analysis and targeting of the complex to the chromatin‐bound fraction of cells undergoing replication fork arrest
Authors: Yuasa Mayumi S. , Masutani Chikahide , Hirano Akihiko , Cohn Martin A. , Yamaizumi Masaru , Nakatani Yoshihiro , Hanaoka Fumio
Published: May 2006

DNA polymerase eta (Polη) is responsible for efficient translesion synthesis (TLS) past cis‐syn cyclobutane thymine dimers (TT dimers), the major DNA lesions induced by UV irradiation. Loss of human Polη leads to xeroderma pigmentosum variant syndrome, clearly indicating that Polη plays a vital role in preventing skin cancer caused by exposure to sunlight. To further examine Polη functions and the mechanisms that regulate this important protein, Polη complexes were purified from HeLa cells over‐expressing epitope‐tagged Polη, and polypeptides associated with Polη, including Rad18, Rad6 and Rev1, were identified by a combination of mass spectrometry and Western blot analysis. The chromatin‐bound fractions of cells subjected to UV irradiation, S phase synchronization, or S phase arrest were specifically enriched in such complexes. These results suggest that arrested replication forks strengthen interactions among Polη, Rad18/Rad6 and Rev1, consistent with the requirement for effective TLS by Polη at sites of DNA lesions.
Reconstitution ofSaccharomyces cerevisiaeprereplicative complex assemblyin vitro
Authors: Kawasaki Yasuo , Kim Hee‐Dai , Kojima Akihiro , Seki Takashi , Sugino Akio
Published: May 2006

The assembly of the prereplicative complex (pre‐RC) at the origin of replication in eukaryotes is a highly regulated and highly conserved process that plays a critical role in preventing multiple rounds of DNA replication per cell division cycle. This study analyzes the molecular dynamics of the assembly ofSaccharomyces cerevisiaepre‐RCin vitrousingARS1plasmid DNA and yeast whole cell extracts. In addition, pre‐RC assembly was reconstitutedin vitrousingARS1DNA and purified origin‐recognition complex (ORC), Cdc6p and Cdt1p‐Mcm2‐7p. The results reveal sequential recruitment of ORC, Cdc6p, Cdt1p and Mcm2‐7p on toARS1DNA. When Mcm2‐7p is maximally loaded, Cdc6p and Cdt1p are released, suggesting that these two proteins are co‐ordinately regulated during pre‐RC assembly. In extracts fromsid2‐21mutant cells that are deficient inCDT1, ORC and Cdc6p bind toARS1but Cdt1p and Mcm2‐7p do not. However, Mcm2‐7p does bind in the presence of exogenous Cdt1p or Cdt1p‐Mcm2‐7p complex. Cdt1p‐Mcm2‐7p complex, which was purified from G1‐, early S or G2/M‐arrested cells, exhibits structure‐specific DNA binding, interacting only with bubble‐ or Y‐shape‐DNA, but the biological significance of this observation is not yet known.
Adrenomedullin regulates expressions of transforming growth factor‐β1 and β1‐induced matrix metalloproteinase‐2 in hepatic stellate cells
Authors: Wang Yi , Zhang Jin‐Sheng , Qian Jin , Huang Guang‐Cun , Chen Qi
Published: May 2006

Adrenomedullin (AM), a peptide isolated from human pheochromocytoma, can be produced and secreted by various types of cells including hepatic stellate cells (HSCs), and its possible role in HSCs is not clear now. In the present study, the interactive regulation between transforming growth factor (TGF)‐β1 and AM and the effect of AM on TGF‐β1‐induced matrix metalloproteinase (MMP)‐2 expression in HSCs were investigated. TGF‐β1 and AM inhibited gene transcript level mutually (real‐time reverse transcription‐polymerase chain reaction). AM suppressed the protein expression level of TGF‐β1 (Western blot), but TGF‐β1 might have no effect on AM secretion level. MMP‐2 protein expression in HSCs was increased in response to TGF‐β1, and upregulation of MMP‐2 expression stimulated with TGF‐β1 was suppressed by AM in dose‐dependent manner (Western blot). AM decreased the phosphorylation level of extracellular signal‐regulated kinase (ERK) in HSCs treated with TGF‐β1, and TGF‐β1‐induced MMP‐2 expression was suppressed by adding Mitogen‐activated protein Kinase/ERK (MEK) inhibitor U0126 (Western blot). Our results suggest that AM may intervene the activation of HSCs by inhibiting TGF‐β1 production and TGF‐β1‐induced MMP‐2 expression; AM may suppress the upregulation of MMP‐2 expression induced by TGF‐β1 partially through ERK pathway.
Effects of transforming growth factor‐β in the development of inflammatory pseudotumour‐like lesions in a murine model
Authors: Guariniello Luciana Doria , Correa Mariangela , Jasiulionis Miriam Galvonas , Machado Joel , Silva José Antônio , Pesquero João Bosco , Carneiro Célia Regina Whitaker
Published: May 2006

Alterations in transforming growth factor (TGF)‐β signalling have been frequently implicated in human cancer, and an important mechanism underlying its pro‐oncogenic nature is suppression of the host antitumour immune response. Considering the immunosuppressive effect of TGF‐β, we asked whether human tumour cells, known to secrete TGF‐β in culture, would survive and grow when implanted into the peritoneal cavity of immunocompetent mice. Therefore, we developed a xenogeneic model where mice were intraperitoneally (i.p.) injected with a TGF‐β‐secreting human colorectal adenocarcinoma cell line, LISP‐A10. Although animals did not develop macroscopic tumours, the recovery and isolation of human tumour cells was achieved when an inflammatory environment was locally induced by the administration of complete Freund's adjuvant (CFA). This procedure significantly increased TGF‐β concentrations in the peritoneal fluid and was accompanied by impaired activation of the host‐specific immune response against LISP‐A10 cells. Furthermore, inflammatory lesions resembling human inflammatory pseudotumours (IPTs) were observed on the surface of i.p. organs. These lesions could be induced by either injection of LISP‐A10 cells, cells‐conditioned medium or recombinant TGF‐β but only after administration of CFA. In addition, host cyclooxygenase‐2 and kinin receptors played an important role in the induction of TGF‐β‐mediated IPT‐like lesions in our experimental model.
Heart failure alters MyoD and MRF4 expressions in rat skeletal muscle
Authors: Carvalho Robson Francisco , Cicogna Antonio Carlos , Campos Gerson Eduardo Rocha , Lopes Francis da Silva , Sugizaki Mário Mateus , Nogueira Célia Regina , Pai‐Silva Maeli Dal
Published: May 2006

Heart failure (HF) is characterized by a skeletal muscle myopathy with increased expression of fast myosin heavy chains (MHCs). The skeletal muscle‐specific molecular regulatory mechanisms controlling MHC expression during HF have not been described. Myogenic regulatory factors (MRFs), a family of transcriptional factors that control the expression of several skeletal muscle‐specific genes, may be related to these alterations. This investigation was undertaken in order to examine potential relationships between MRF mRNA expression and MHC protein isoforms in Wistar rat skeletal muscle with monocrotaline‐induced HF. We studied soleus (Sol) and extensor digitorum longus (EDL) muscles from both HF and control Wistar rats. MyoD, myogenin and MRF4 contents were determined using reverse transcription‐polymerase chain reaction while MHC isoforms were separated using polyacrylamide gel electrophoresis. Despite no change in MHC composition of Wistar rat skeletal muscles with HF, the mRNA relative expression of MyoD in Sol and EDL muscles and that of MRF4 in Sol muscle were significantly reduced, whereas myogenin was not changed in both muscles. This down‐regulation in the mRNA relative expression of MRF4 in Sol was associated with atrophy in response to HF while these alterations were not present in EDL muscle. Taken together, our results show a potential role for MRFs in skeletal muscle myopathy during HF.
Trypanosoma cruzi: clones isolated from the Colombian strain, reproduce the parental strain characteristics, with ubiquitous histotropism
Authors: Camandaroba Edson , Thé Torriceli S. , Pessina Daniel Huber , Andrade Sonia G.
Published: May 2006

Clonal histotropism and biological characters of five clones isolated during the early acute phase of the infection of Swiss mice with the Colombian strain of Trypanosoma cruzi (T. cruzi I), Biodeme Type III, were investigated. Clones were isolated from mice at the 10th and the 30th day of infection with the Colombian strain. Isolation was performed by micromanipulation and injection of one trypomatigote blood form into newborn mice, followed by passages into suckling mice for obtaining the inocula for the experimental groups. Mice infected with parental strain were also studied. All the clones have shown the basic characteristics of Biodeme Type III, with the same patterns of parasitemia, tissue tropism, morphological characters and isoenzymic profiles, such as the parental strain. Histotropism was most intense to myocardium and skeletal muscles, with intense lesions found in the advanced phase (20th to 30th day of infection). Both parental strain and the clones were seen to parasitize several organs and tissues; amastigote nests were identified in the cytoplasm of macrophages, adipose cells, smooth muscle of intestinal wall and Auerbach's neuronal plexus. The findings of the present study confirm the homology of the clones isolated from the Colombian strain, with predominance of a ‘principal clone’ and an ubiquitous distribution of parasites belonging to a same clone.
Developmental abnormalities in multiple proliferative tissues of ApcMin/+ mice
Authors: You Shaojin , Ohmori Masami , Peña Maria Marjorette O. , Nassri Basel , Quiton Jovelyn , Al‐Assad Ziad A. , Liu Lucy , Wood Patricia A. , Berger Sondra H. , Liu ZhiJian , Wyatt Michael D. , Price Robert L. , Berger Franklin G. , Hrushesky William J. M.
Published: May 2006

Germ‐line mutation of the Apc gene has been linked to familial adenomatous polyposis (FAP) that predisposes to colon cancer. ApcMin/+ mice, heterozygous for the Apc gene mutation, progressively develop small intestinal tumours in a manner that is analogous to that observed in the colon of patients with FAP (Su et al. 1992; Fodde et al. 1994; Moser et al. 1995). We have studied the effects of Apc gene mutation on murine intestinal and extra‐intestinal, proliferatively active tissues. We have contrasted the histology to that of the age‐ and sex‐matched wild‐type C57BL/6 mice. Histological assessment of the normal appearing intestinal mucosa demonstrates minimal change in size of crypts. In contrast, villi are longer in the ileum of ApcMin/+ mice relative to C57BL/6 mice at 12 and 15 weeks of age. Vigorous splenic haematopoiesis in ApcMin/+ mice was seen at 12 and 15 weeks of age, as reflected by marked splenomegaly, increased splenic haematopoietic cells and megakaryocytes. Peripheral blood counts, however, did not differ between C57BL/6 and ApcMin/+ mice at 15 weeks of age. Lymphoid depletion in ApcMin/+ mice was characterized by diminished numbers of splenic lymphoid follicles and small intestinal Peyer's patches. The ovaries of 12‐ and 15‐week‐old ApcMin/+ mice exhibited increased numbers of atretic follicles, and estrous cycling by serial vaginal smears showed tendency of elongation in the mutant mice during these age ranges. The testicles of 10‐week‐old ApcMin/+ mice showed increased numbers of underdeveloped seminiferous tubules. Collectively, these data suggest that, in addition to its obvious effects upon intestinal adenoma formation, Apc gene mutation causes impairment of developmental and apparent differentiation blockade in proliferative tissues, including those of the haematopoietic system, lymphoid and reproductive tract.
Tumour‐associated macrophages and melanoma tumourigenesis: integrating the complexity
Authors: Hussein Mahmoud R.
Published: May 2006

When the body discovers a tumour cell (foreign antigen), several kinds of mechanisms and cells operate in what is called an immune response. The latter has evolved into two mechanisms: non‐specific immunity and specific immunity, which are closely linked to and influence each other. The former represents the first line of defence against neoplastic cells. The adaptive (specific) immunity is orchestrated by antigen‐specific T and B lymphocytes. The effector cells of innate immunity include granulocytes, macrophages and natural killer cells. Among these cells, macrophages represent the most important part of innate immunity against tumours. Tumour‐associated macrophages (TAMs) are important antigen‐presenting cells and as such an understanding of their interactions with tumour cells gives insights into novel therapeutic strategies. In tumours, the effect of TAMs is the outcome of their two concomitantly competing interactions: tumour growth reduction and tumour growth promotion. The macrophage (TAMs) content of melanoma ranges from 0 to 30% and their density increases with increasing tumour thickness. The melanoma cells and TAMs seem to interact with each other through the release of soluble factors that either prevent or enhance tumour growth. For instance, syngeneic macrophages from tumour‐bearing mice can inhibit melanoma growth in the nude mice more than the control macrophages. Alternatively, metastatic B16 melanoma cells can produce some macrophage cytotoxic substances that help tumour cells not only escape the host immunosurveillance system but also prevent distant metastasis. Together, these observations suggest opposing effects for these soluble factors in melanoma. To date, little is available in the literature about the interactions between TAMs and melanoma cells. This viewpoint not only tries to examine these interactions but also provides relevant speculations.
TGFβ1 induces epithelial–mesenchymal transition, but not myofibroblast transdifferentiation of human kidney tubular epithelial cells in primary culture
Authors: Forino Monica , Torregrossa Rossella , Ceol Monica , Murer Luisa , Vella Manuela Della , Prete Dorella Del , D'Angelo Angela , Anglani Franca
Published: May 2006

The origin and fate of renal interstitial myofibroblasts (MFs), the effector cells of renal fibrosis, are still debated. Experimental evidence suggests that renal MFs derive from tubular epithelial cells throughout the epithelial–mesenchymal transition (EMT) process. Primary human tubular epithelial cells (HUTECs) were cultured for 4 and 6 days on plastic or type I collagen‐coated plates with 1, 5, 10 and 50 ng/ml of transforming growth factor β1 (TGFβ1). The EMT process was monitored by morphology and immunophenotyping for αSMA, cytokeratin 8–18, E‐cadherin, vimentin and collagen III. Quantitative comparative RT/PCR and real‐time PCR were used to evaluate the expression of collagen III and IV, fibronectin, tenascin, MMP‐2, CTGF, E‐cadherin and cadherin 11 genes, as well as those of the Smad signalling pathway. TGFβ1 was found capable of reactivating the mesenchymal programme switched off during tubulogenesis, but it induced no de novo expression of αSMA gene or myofibroblast phenotype. We demonstrate that the EMT process is conditioned by the extracellular matrix and characterized by TGFβ1‐driven Smad3 downregulation. Our study results suggest that TGFβ1 could function as a classic embryonal inducer, initiating a cascade of de‐differentiating events that might be further controlled by other factors in the cellular environment.
Morphological evaluation of the radioprotective effects of melatonin against X‐ray‐induced early and acute testis damage in Albino rats: an animal model
Authors: Hussein Mahmoud R. , Abu‐Dief Eman E. , Abou El‐Ghait Amal T. , Adly Mohamed A. , Abdelraheem Mohamad H.
Published: May 2006

Irradiation has profound effects on the reproductive function. Our knowledge about radioprotective effects of melatonin against X‐ray‐induced testis damage is rudimentary. In this investigation, we hypothesized that melatonin can minimize germ‐cell depletion and morphological features of cell damage in testis following X‐ray irradiation (XRI). To examine these effects, and to test our hypothesis, an animal model comprised of 60 Albino rats was established. The animals were divided into five groups: Group 1, non‐irradiated; Group 2, X‐ray irradiated (XRI, 8 Grays); Group 3, XRI pretreated with solvent (ethanol and phosphate‐buffered saline); Group 4, non‐irradiated group treated with melatonin and Group 5, XRI pretreated with melatonin. The testes were evaluated for both histological (light microscopy) and ultrastructural changes (transmission electron microscopy). Histologically, there were marked depletions (66%) of the germinal epithelial cells, in XRI group (Groups 2 and 3), whereas these changes were almost absent in XRI testis of animals pretreated with melatonin (Group 5). The number of spermatogenic cells in XRI testis of animals pretreated with melatonin (Group 5) was comparable (95%) to that of non‐irradiated group (Groups 1 and 4) but significantly (P < 0.05) higher than those in XRI testis (34%, Groups 2 and 3). Ultrastructurally, XRI testis (Groups 2 and 3) showed features of apoptosis (condensation of the nuclei, vacuolization of the cytoplasm, increased cytoplasmic density and apoptotic bodies). These features were absent in XRI testis of animals pretreated with melatonin (Group 5). Also, this Group showed features of an increased metabolic activity (large acrosomal vesicle, prominent Golgi, increased mitotic activity, increased complement of cytoplasmic organelles and appearance of nucleoli‐like bodies). There was a minimal depletion of the Sertoli and Leydig cells following XRI. Also, morphological features of apoptosis were infrequent in these cells. Administration of melatonin (MEL) prior to irradiation can protect testis against its destructive effects. The protective effects include amelioration of germ‐cell depletion and apoptotic changes. The clinical ramifications of these observations mandate further studies.
Regulation ofEscherichia coli hchA, a stress‐inducible gene encoding molecular chaperone Hsp31
Authors: Mujacic Mirna , Baneyx François
Published: May 2006

Escherichia coliHsp31 is a homodimeric member of the ThiI/DJ‐1/PfpI superfamily that combines molecular chaperone and aminopeptidase activities. Although it was originally identified on the basis of its induction by heat shock, little is known about the regulation ofhchA, the structural gene encoding Hsp31. Here, we show thathchAis transcribed from dual promoters recognized by the σD(σ70) and σS(σ38) subunits of RNA polymerase (E). In exponentially growing cells, the nucleoid‐binding protein H‐NS downregulates Hsp31 synthesis, andhchAthermal induction primarily relies on the relief of H‐NS‐mediated silencing of EσD‐dependent transcription. This uncommon alternative to the use of a heat‐shock sigma factor guarantees that Hsp31 concentration remains high throughout the length of the high temperature exposure phase. Entry into stationary phase leads to enhancedhchAtranscription from its EσS‐dependent promoter. Consistent with a role of Hsp31 in the management of starvation,hchAnull mutants exhibit a decrease ability to survive in deep stationary phase relative tohchA+cells. Based onhchAheat‐inducibility and membership in the σSgeneral stress regulon, we propose that Hsp31 performs a protective function under a wide range of stress conditions.
A mechanical role for the chemotaxis system in swarming motility
Authors: Mariconda Susana , Wang Qingfeng , Harshey Rasika M.
Published: May 2006

The chemotaxis system, but not chemotaxis, is essential for swarming motility in Salmonella enterica serovar Typhimurium. Mutants in the chemotaxis pathway exhibit fewer and shorter flagella, downregulate class 3 or ‘late’ motility genes, and appear to be less hydrated when propagated on a surface. We show here that the output of the chemotaxis system, CheY∼P, modulates motor bias during swarming as it does during chemotaxis, but for a distinctly different end. A constitutively active form of CheY was found to promote swarming in the absence of several upstream chemotaxis components. Two point mutations that suppressed the swarming defect of a cheY null mutation mapped to FliM, a protein in the motor switch complex with which CheY∼P interacts. A common property of these suppressors was their increased frequency of motor reversal. These and other data suggest that the ability to switch motor direction is important for promoting optimal surface wetness. If the surface is sufficiently wet, exclusively clockwise or counterclockwise directions of motor rotation will support swarming, suggesting also that the bacteria can move on a surface with flagellar bundles of either handedness.