bluuebunny/crossref_metadata_2025 huggingface dataset:
Tedisamil attenuates foetal transformation of myosin in the hypertrophied rat myocardium
Authors: Turcani Marian , Thormaehlen Dirk , Rupp Heinz
Published: October 2004

Reduction in repolarizing potassium currents has controversial effects on hypertrophic responses in cardiomyocytes of transgenic models and cultured cardiomyocytes. It remains thus unknown whether a blockade of potassium channels with tedisamil (N,N′dicyclopropylmethylene‐9,9‐tetramethylene‐3,7‐diazabicyclo(3.3.1)nonane dihydrochloride) has any effects on cardiac growth during postnatal development or pressure overload.

To test the hypothesis that a treatment with tedisamil affects cardiac growth or protein phenotype, sham‐operated rats and rats with ascending aorta constriction were treated with tedisamil (36 mg kg day−1) for 7 weeks. Left ventricular mass and geometry, relative expression of myosin isoforms, hydroxyproline concentration and isovolumic ventricular function were assessed.

Rats with aortic constriction exhibited a marked increase in left ventricular weight and the diastolic pressure–volume relationship was shifted to smaller volumes. The hydroxyproline concentration remained unaltered. The proportion of α‐myosin heavy chains was, however, reduced (P<0.05). Hypertrophied left ventricles manifested an enhanced overall performance but depressed myocardial contractility.

Administration of tedisamil was associated with decreased heart rate (P<0.05). In contrast, cardiac growth in sham‐operated rats and concentric left ventricular hypertrophy of pressure‐overloaded animals was not significantly altered. Hypertrophied hearts from rats treated with tedisamil expressed more α‐myosin heavy chains (65±4 versus 57±4%; P<0.05). Also, maximal rate of wall stress rise and decline was higher (P<0.05) in tedisamil‐treated pressure‐overloaded rats.

In the rat model of pressure‐overloaded hypertrophy, tedisamil had no effect on cardiac growth but partially corrected myocardial dysfunction. Postulated mechanism of this effect is the phenotype modification of myosin filaments in hypertrophied myocardium.

British Journal of Pharmacology (2004) 143, 561–572. doi:10.1038/sj.bjp.0705992
Glucocorticoid hormone (cortisol) affects axonal transport in human cortex neurons but shows resistance in Alzheimer's disease
Authors: Dai Jiapei , Buijs Ruud , Swaab Dick
Published: October 2004

The changes of tissue sensitivity to glucocorticoids are associated with many pathological states including neurological diseases. In the present study, using a novel in vitro post‐mortem tracing method on human brain slices, we demonstrated that cortisol, a major glucocorticoid hormone in humans, affected axonal transport both in the cortex neurons in four Alzheimer's disease (AD) patients and four nondemented controls.

Cortisol appeared to affect axonal transport of prefrontal cortex (PFC) and temporal cortex (TC) neurons in AD patients and controls in a dose‐dependent way at concentrations of 30, 60, 120 and 240 μg dl−1.

Higher doses of cortisol were needed for TC neurons to achieve a similar axonal transport effect as obtained in PFC neurons in AD patients. The maximum effect (Emax) on axonal transport was achieved in PFC slices at relatively low contraction (30–120 μg dl−1), while in TC slices, a maximum effect was only reached at relatively high concentrations (120–240 μg dl−1).

For PFC and TC slices from nondemented aging subjects, lower doses of cortisol (30–60 μg dl−1) on axonal transport were sufficient to achieve the maximum effect as compared to those used in AD brain slices, while levels of more than 60 μg dl−1 of cortisol mostly depressed axonal transport.

These results suggest that glucocorticoid resistance, which is thought to contribute to the pathogenesis of a number of common human disorders, may exist in AD brains and play an important role in neuropathological mechanisms and dementia.

British Journal of Pharmacology (2004) 143, 606–610. doi:10.1038/sj.bjp.0705995
Novel role for P2X receptor activation in endothelium‐dependent vasodilation
Authors: Harrington Louise S , Mitchell Jane A
Published: October 2004

ATP is an important vasoactive mediator, which acts via two receptor classes: P2X and P2Y. Activation of P2X receptors has traditionally been associated with the well‐characterised vasoconstrictor properties of ATP.

In the current study, we have shown that the P2X1 & 3 receptor ligand, α, β methylene ATP, induces vasodilation of rat isolated mesenteric arteries and that P2X1 receptors are abundantly expressed in the endothelium of these vessels.

Second‐order rat mesenteric arteries were mounted in myographs and vasomotor responses recorded. Both ATP and α, β methylene ATP induced a constriction followed by a vasodilation. The dilator effects of either ATP or α, β methylene ATP were slower in onset than those induced by acetylcholine. By contrast, the traditional vasodilator P2Y ligand, ADP, induced vasodilation without contraction.

Vasodilation induced by α, β methylene ATP was endothelial dependent, but was not affected by treatment of the vessels with l‐NAME plus indomethacin alone. Dilation was, however, partially inhibited by the combination of apamin plus charybdotoxin and blocked by treating vessels with all four drugs.

Using confocal microscopy, P2X1 receptor immunoreactivity was localised to the endothelial, smooth muscle and adventitial layers of mesenteric vessels. P2X1 protein migrated as a primary band at around 50–60 kDa in vascular tissue.

These results show for the first time that P2X1 receptors are expressed on the endothelium and that a selective ligand of this receptor results in vasoconstriction followed by vasodilation. These observations have important implications for our understanding of the role of purines in biological responses.

British Journal of Pharmacology (2004) 143, 611–617. doi:10.1038/sj.bjp.0706004
Tail-anchored protein biosynthesis at the endoplasmic reticulum: the same but different
Authors: High S. , Abell B.M.
Published: October 2004

The post-translational integration of tail-anchored proteins at the endoplasmic reticulum represents a novel and distinct pathway for membrane protein synthesis. Studies of various precursors, exemplified by the synaptobrevins and cytochrome b5, indicate that multiple routes may facilitate their biosynthesis. There is clear evidence that both cytosolic factors and membrane components facilitate the efficient membrane insertion of at least some tail-anchored proteins. However, the nature of these mediators is currently unknown and their identification will be an essential step in defining the molecular basis of tail-anchored protein biogenesis.
Catalysis of disulphide bond formation in the endoplasmic reticulum
Authors: Ellgaard L.
Published: October 2004

Disulphide bonds are critical for the maturation and stability of secretory and cell-surface proteins. In eukaryotic cells, disulphide bonds are introduced in the ER (endoplasmic reticulum), where the redox conditions are optimal to support their formation. Yet, the correct pairing of cysteine residues is not simple and often requires the assistance of redox-active proteins. The enzymes of the thiol-disulphide oxidoreductase family catalyse oxidation, reduction and isomerization, and thereby play important roles for the folding of many proteins. To allow all three redox reactions to take place concurrently in the same compartment, specific protein–protein interactions regulate the function of individual enzymes, while a careful balance of the ER redox environment is maintained. At the same time, the system must be capable of responding to changes in the cellular conditions, caused, for instance, by oxidative stress and protein misfolding. This review presents recent progress in understanding how ER redox conditions are regulated and how protein disulphides are formed in the ER of mammalian cells.
Characterization of the cis-acting element directing perinuclear localization of the metallothionein-1 mRNA
Authors: Chabanon H. , Nury D. , Mickleburgh I. , Burtle B. , Hesketh J.
Published: October 2004

Subcellular localization of mRNAs is a key mechanism for the synthesis of proteins close to their site of function. The mRNA encoding MT-1 (metallothionein-1) is localized in the perinuclear cytoplasm, where it is associated with cytoskeletal-bound polysomes. This localization relies on sequences present in the 3′-UTR (3′-untranslated region). The present study aims to characterize the cis-acting localization element(s) within the 3′-UTR. Using transfected cells expressing tagged MT-1 differing in their 3′-UTRs (deleted or mutated), the section(s) of this region required for directing MT-1 transcripts to the perinuclear cytoplasm has been investigated. Different 3′-UTRs were also used in UV cross-linking experiments that highlighted two distinct regions (nt 26–30 and 66–76) necessary for the binding of a protein of approx. 50 kDa, presumably involved in the mRNA targeting. The poor sequence homology between the MT-1 3′-UTR of various species, together with the bipartite nature of the required cis-element, indicates the involvement of a particular structure in the localization signal. The secondary structure of the MT-1 3′-UTR was investigated using enzymic and chemical probing. Current structural analysis of mutant 3′-UTRs will allow the critical structural features of the MT-1 mRNA perinuclear localization signal to be defined.
p97 and close encounters of every kind: a brief review
Authors: Dreveny I. , Pye V.E. , Beuron F. , Briggs L.C. , Isaacson R.L. , Matthews S.J. , McKeown C. , Yuan X. , Zhang X. , Freemont P.S.
Published: October 2004

The AAA (ATPase associated with various cellular activities) ATPase, p97, is a hexameric protein of chaperone-like function, which has been reported to interact with a number of proteins of seemingly unrelated functions. For the first time, we report a classification of these proteins and aim to elucidate any common structural or functional features they may share. The interactors are grouped into those containing ubiquitin regulatory X domains, which presumably bind to p97 in the same way as the p47 adaptor, and into non-ubiquitin regulatory X domain proteins of different functional subgroups that may employ a different mode of interaction (assuming they also bind directly to p97 and are not experimental artifacts). Future studies will show whether interacting proteins direct p97 to different cellular pathways or a common one and structural elucidation of these interactions will be crucial in understanding these underlying functions.
HAUSP/USP7 as an Epstein–Barr virus target
Authors: Holowaty M.N. , Frappier L.
Published: October 2004

USP7 (also called HAUSP) is a de-ubiquitinating enzyme recently identified as a key regulator of the p53–mdm2 pathway, which stabilizes both p53 and mdm2. We have discovered that the Epstein–Barr nuclear antigen 1 protein of Epstein–Barr virus binds with high affinity to USP7 and disrupts the USP7–p53 interaction. The results have important implications for the role of Epstein–Barr nuclear antigen 1 in the cellular immortalization that is typical of an Epstein–Barr virus latent infection.
X-ray crystallographic studies for ligand–protein interaction changes in rhodopsin
Authors: Okada T.
Published: October 2004

G-protein-coupled receptors constitute the largest transmembrane receptor family in human. They are generally activated on binding their specific ligands at the extracellular side of membranes. The signal carried by an agonist is then transmitted to the intracellular side through a conformational change of the receptor, which becomes competent to catalyse GDP/GTP exchange in the α-subunit of heterotrimeric G-protein. Since most of the G-protein-coupled receptors (rhodopsin-like subfamily) share a set of conserved amino acid residues in the transmembrane domain, it is probable that the ligand-triggered activation process involves a common mechanism of rearrangement of the hepta-helical transmembrane bundle. For understanding the nature of this event that is not yet characterized sufficiently, X-ray crystallographic studies of rhodopsin with or without light stimulation can provide valuable information. In rhodopsin, the initial cis–trans photoisomerization of retinal chromophore triggers the structural changes of transmembrane helices. This activation process has been characterized with some spectroscopically distinct photoreaction intermediates (batho, lumi, Meta I and Meta II). With recent advances in the conditions for crystallographic experiments, the diffraction limit of the rhodopsin crystals has been substantially extended. As a result, it becomes possible to detect small structural changes evoked after photoactivation under cryogenic conditions.
Active and inactive conformations of the epidermal growth factor receptor
Authors: Ferguson K.M.
Published: October 2004

The members of the EGFR (epidermal growth factor receptor) family of RTKs (receptor tyrosine kinases), also known as the ErbB or HER family, have been implicated in many human cancers. Structural studies of the EGFR extracellular region (sEGFR) have led to the proposal of a novel mechanism for ligand-induced receptor dimerization. In this model EGF binding induces a dramatic conformational change in EGFR, exposing a dimerization site that is normally occluded in the inactivated conformation, and thus promoting the formation of an entirely receptor-mediated dimer. It is well established that antibodies against the extracellular region of EGFR that prevent ligand binding and/or receptor signalling can inhibit tumour growth in vivo. At least five such anti-EGFR antibodies are currently in clinical trials and one, C225/cetuximab (Erbitux™), was recently approved in the U.S. and Europe for use in advanced colorectal cancers. Recent structural studies of ErbB2 in complex with anti-ErbB2 antibodies (trastuzumab/Herceptin™ and pertuzumab/Omnitarg™) have provided significant insights into how these drugs function. There have been no such studies for similar EGFR-targeted drugs to date. The implications of this model for the possible mechanisms of antibody-mediated inhibition of EGFR are discussed.
Characterization of simian virus 40 on its infectious entry pathway in cells using fluorescence correlation spectroscopy
Authors: Bernacchi S. , Mueller G. , Langowski J. , Waldeck W.
Published: October 2004

SV40 (simian virus 40) is a double-stranded DNA virus and is strongly oncogenic in experimental animals. SV40 enters cells by atypical endocytosis mediated by caveolae, transporting the virus to its usual destination, namely the endoplasmic reticulum. The cellular mechanisms of capsid disassembly (uncoating) and deliverance of the viral genome into the cellular nucleus remain unknown. Here, we study (i) the formation of caveolae after viral infection and the diffusion of caveosome vesicles in the cytoplasm and (ii) the capsid disassembly and the mobility of the viral genome on its way to the nucleus, using fluorescence correlation spectroscopy. To follow the viral genome and capsids separately, the histone components of SV40 minichromosomes were labelled with enhanced yellow fluorescent protein and the capsid was labelled with a fluorescent red dye, Alexa568. We characterized the diffusion of caveosomes, the capsid disassembly process in the cytoplasm and the mobility of the viral genome in the nucleus, using two kinds of permissive cells.
Activation of the blood–brain barrier by SIV (simian immunodeficiency virus) requires cell-associated virus and is not restricted to endothelial cell activation
Authors: MacLean A.G. , Rasmussen T.A. , Bieniemy D. , Lackner A.A.
Published: October 2004

The primary cell infected during acute HIV neuropathogenesis is the monocyte-derived macrophage. We have demonstrated that there is activation of the BBB (blood–brain barrier) during acute viral infection and at terminal AIDS. However, it has never been determined if there is a requirement for the virus to be carried through the BBB or how these Trojan horses would be induced to cross the BBB. We added SIVmac251-infected (SIV is simian immunodeficiency virus) mononuclear cells (and their supernatants) to the luminal or abluminal compartment of our BBB model. There was activation of both sides of the BBB model, only if viral-infected cells were added to the luminal compartment, as opposed to the addition of cell-free supernatants. This suggests that cell-associated virus is essential for the activation of the BBB. This, in turn, would be expected to lead to further infiltration of cells capable of viral infection. VCAM-1 (vascular cell adhesion molecule 1) staining revealed, for the first time, that there is an absolute requirement for virally infected cells, and not just the presence of virus for the activation of the BBB.
Fluorescence studies of single biomolecules
Authors: Li H. , Ying L. , Ren X. , Balasubramanian S. , Klenerman D.
Published: October 2004

Single-molecule fluorescence has the capability to detect properties buried in ensemble measurements and, hence, provides new insights about biological processes. Ratiometric methods are normally used to reduce the effects of excitation beam inhomogeneity. Fluorescence resonance energy transfer is widely used but there are problems in inserting the fluorophores in the correct position on the biomolecule, particularly if the structure is not known. We have recently developed two-colour coincidence single-molecule fluorescence that addresses this problem. This method can be used to determine quantitatively the multimerization states of biomolecules, in solution without separation. The future prospects of single-molecule fluorescence as applied to biological molecules are discussed.
Single protein molecular dynamics determined with ultra-high precision
Authors: Sasaki Y.C.
Published: October 2004

We have successfully observed dynamical Brownian motions in an individual protein molecule and other biological ones in real-time with one-hundredth the atomic-scale precision (picometer-scale precision) using X-rays of the super photon ring-8 (SPring-8).
Mass spectrometry is a powerful tool for identification of proteins associated with lipid rafts of Jurkat T-cell line
Authors: Pompach P. , Man P. , Novák P. , Havlíček V. , Fišerová A. , Bezouška K.
Published: October 2004

Many proteins involved in signal-transduction pathways are concentrated in membrane microdomains enriched in lipids with distinct physical properties. Since these microdomains are insoluble in non-ionic detergents in cold, proteins associated with them could be efficiently purified by techniques such as sucrose-density gradient centrifugation. The complexity of the resulting protein mixture requires powerful MS technique for its analysis. We have found that successful identification of biologically relevant proteins is critically dependent on the enrichment of the starting material (plasma membranes), and on the extraction procedure. Applying these conditions in combination with microHPLC-ESI (electrospray ionization)-MS/MS, we have identified proteins involved in signalling, cytoskeletal association and cellular adhesion in Jurkat cells that are not stimulated by any antibody incubation.
BCR activation of PI3K is Vav-independent in murine B cells
Authors: Vigorito E. , Clayton E. , Turner M.
Published: October 2004

BCR (B-cell antigen receptor)-induced Ca2+ signalling is initiated by activation of tyrosine kinases, which in concert with adaptor proteins and lipid kinases regulate PLC (phospholipase C) γ2 activation. Vav and PI3K (phosphoinositide 3-kinase) are required for optimal Ca2+ responses, although it has not been established, in primary B-cells, if both proteins are components of the same pathway. In vitro evidence suggests that binding of the PI3K lipid product PIP3 to Vav pleckstrin homology domain contributes to Vav activation. However, pharmacological inhibition of PI3K by wortmannin or deletion of the p110δ catalytic subunit has no effect on Vav activation in response to BCR engagement, suggesting that this mechanism does not operate in vivo. We also show that PI3K recruitment to phosphorylated-tyrosine-containing complexes is Vav-independent. Taken together with our previous observation that protein kinase B phosphorylation is normal in Vav-deficient B-cells, we suggest that PI3K activation is Vav-independent in response to strong signals delivered by multivalent cross-linking.
Glycogen synthase kinase-3 in insulin and Wnt signalling: a double-edged sword?
Authors: Patel S. , Doble B. , Woodgett J.R.
Published: October 2004

Glycogen synthase kinase-3 is an unusual protein serine/threonine kinase that, unlike most of its 500-odd relatives in the genome, is active under resting conditions and is inactivated upon cell stimulation. The two mammalian isoforms, GSK-3α and β, play largely overlapping roles and have been implicated in a variety of human pathologies, including Type II diabetes, Alzheimer's disease, bipolar disorder and cancer. Recently, the modes of regulation of this enzyme have been elucidated through a combination of structural and cell biological studies. A series of relatively selective small molecules have facilitated chemical manipulation of the enzyme in intact cells and tissues, and new roles for the protein kinase in embryonic stem cell differentiation and motility have emerged. Despite these advances, the therapeutic value of this enzyme as a drug target remains clouded by uncertainty over the potential of antagonists to promote tumorigenesis. This article describes the state of understanding of this intriguing enzyme, and weighs current evidence regarding whether there is a therapeutic window for amelioration of diseases in which it is implicated, in the absence of inducing new pathologies.
G-protein-coupled receptor allosterism: the promise and the problem(s)
Authors: Christopoulos A. , May L.T. , Avlani V.A. , Sexton P.M.
Published: October 2004

Allosteric modulators of G-protein-coupled receptors interact with binding sites that are topographically distinct from the orthosteric site recognized by the receptor's endogenous agonist. Allosteric ligands offer a number of advantages over orthosteric drugs, including the potential for greater receptor subtype selectivity and a more ‘physiological’ regulation of receptor activity. However, the manifestations of allosterism at G-protein-coupled receptors are quite varied, and significant challenges remain for the optimization of screening methods to ensure the routine detection and validation of allosteric ligands.
G-protein-coupled receptor accessory proteins: their potential role in future drug discovery
Authors: Presland J.
Published: October 2004

Historically, the activation and inhibition of GPCR (G-protein-coupled receptor) function have been a very successful avenue for drug discovery and development. However, it is clear that receptors do not function in isolation but are impacted by other proteins. These proteins may alter either binding or functional responses. Identification and study of these interactions have grown rapidly in recent years and continue to do so, resulting in a plethora of potential receptor–protein connections. These associations can be regarded as alternative intervention points to modulate GPCR function and may not only provide alternative ways to modify receptor activity but also to exploit new chemical space for drug-like molecules. Such interactions may account for side-effects or undesirable properties associated with otherwise well-validated GPCR targets. Understanding and/or intervening in these interactions may allow scientists to progress those targets that may have been deemed unsuitable for therapeutic intervention. The present study reviews the opportunities for utilizing receptor interacting proteins as potential drug targets and the issues associated with them.
The ubiquitous phosphoinositides
Authors: Parker P.J.
Published: October 2004

There are now known to exist seven phosphoinositides all derived through various metabolic routes from the parent lipid phosphatidylinositol. With one additional metabolite, diacylglycerol, these represent a rich resource of bioactive lipids responsible for recruiting protein effectors and marking membrane compartments. The metabolic map of this pathway and the nature of the binding partner interactions are reviewed.
A molecular code for splicing silencing: configurations of guanosine-rich motifs
Authors: Grabowski P.J.
Published: October 2004

Alternative pre-mRNA splicing is frequently used to expand the protein-coding capacity of genomes, and to regulate gene expression at the post-transcriptional level. It is a significant challenge to decipher the molecular language of tissue-specific splicing because the inherent flexibility of these mechanisms is specified by numerous short sequence motifs distributed in introns and exons. In the present study, we employ the glutamate NMDA (N-methyl-D-aspartate) R1 receptor (GRIN1) transcript as a model system to identify the molecular determinants for a brain region-specific exon silencing mechanism. We identify a set of guanosine-rich motifs that function co-operatively to regulate the CI cassette exon in a manner consistent with its in vivo splicing pattern. Whereas hnRNP (heterogeneous nuclear ribonucleoprotein) A1 mediates silencing of the CI cassette exon in conjunction with the guanosine-rich motifs, hnRNP H functions as an antagonist to silencing. Genome-wide analysis shows that, while this motif pattern is rarely present in human and mouse exons, those exons for which the pattern is conserved are generally found to be skipped exons. The identification of a similar arrangement of guanosine-rich motifs in transcripts of the hnRNP H family of splicing factors has implications for their co-ordinate regulation at the level of splicing.
Transcriptional regulation by the Wilms' tumour suppressor protein WT1
Authors: Wagner K.J. , Roberts S.G.E.
Published: October 2004

Wilms' tumour is a paediatric malignancy of the kidneys and is the most common solid tumour found in children. The Wilms' tumour suppressor protein WT1 is mutated in approx. 15% of Wilms' tumours, and is aberrantly expressed in many others. WT1 can manifest both tumour suppressor and oncogenic activities, but the reasons for this are not yet clear. The Wilms' tumour suppressor protein WT1 is a transcriptional activator, the function of which is under cell-context-specific control. We have previously described a small region at the N-terminus of WT1 (suppression domain) that inhibits the transcriptional activation domain by contacting a co-suppressor protein. We recently identified BASP1 as one of the components of the co-suppressor. Here, we analyse the mechanism of action of the WT1 suppression domain, and discuss its function in the context of the role of WT1 as a regulator of development.
The Flightless I protein and the gelsolin family in nuclear hormone receptor-mediated signalling
Authors: Archer S.K. , Behm C.A. , Claudianos C. , Campbell H.D.
Published: October 2004

The Drosophila melanogaster flightless I protein and its homologues in higher eukaryotes (FliI) are conserved members of the gelsolin family of actin-binding proteins. Members of the gelsolin family generally contain three or six copies of a 125-amino-acid residue gelsolin-related repeating unit, and may contain additional domains including the C-terminal villin-related ‘headpiece’ or N-terminal extensions such as the leucine-rich repeat of the FliI protein. Numerous studies including work done with mouse knockouts for gelsolin, villin and CapG support a role for the family in cytoskeletal actin dynamics. In both fruitfly and mouse, the FliI protein is also essential for early development. Recent studies indicate that supervillin, gelsolin and FliI are involved in intracellular signalling via nuclear hormone receptors including the androgen, oestrogen and thyroid hormone receptors. This unexpected role in signalling has opened a new area in research on the gelsolin family and is providing important new insights into the mechanisms of gene regulation via nuclear receptors.
Redox regulation of phosphatase function
Authors: Leslie N.R. , Lindsay Y. , Ross S.H. , Downes C.P.
Published: October 2004

Although reactive oxygen species play important roles in cellular physiology as signalling molecules, their molecular targets are largely unknown. A probable group of targets for mediating many of the effects of reactive oxygen species on cell signalling is the large diverse family of cysteine-dependent phosphatases, which includes the protein tyrosine phosphatases. Our work and that of others suggest that the oxidative inactivation of protein and lipid phosphatases plays an important part in signalling, downstream of many cellular stimuli. Future studies should give us a clearer picture of the role of phosphatase inactivation in cellular behaviour and explain how specificity is achieved in redox signalling.
Genes controlling the metabolic switch in hibernating mammals
Authors: Andrews M.T.
Published: October 2004

Hibernating mammals have the ability to decrease their metabolic rate and survive up to 6 months without food in an inactive state where body temperatures approach 0°C. In hibernating 13-lined ground squirrels (Spermophilus tridecemlineatus), oxygen consumption holds at 1/30 to 1/50 of the aroused condition and heart rates are as low as 3–10 beats/min, compared with 200–300 beats/min when the animal is active. This seasonal adaptation requires a metabolic shift away from the oxidation of carbohydrates and towards the combustion of stored fatty acids as the primary source of energy. A key element in this fuel switch is the differential expression of the gene encoding pyruvate dehydrogenase kinase isoenzyme 4. Pyruvate dehydrogenase kinase isoenzyme 4 inhibits pyruvate dehydrogenase and thus minimizes carbohydrate oxidation by preventing the flow of glycolytic products into the tricarboxylic acid cycle. Hibernators also exploit the low-temperature activity of PTL (pancreatic triacylglycerol lipase) in both heart and white adipose tissue. Lipolytic activity at body temperatures associated with hibernation was examined using recombinant ground squirrel and human PTL expressed in yeast. Enzymes from both humans and ground squirrel displayed high activity at temperatures as low as 0°C and showed Q10=1.2–1.5 over the temperature range 37–7°C. These studies indicate that low-temperature lipolysis is a general property of PTL and does not require protein modifications unique to mammalian cells and/or the hibernating state.
Kinetics of G-protein-coupled receptor signalling and desensitization
Authors: Krasel C. , Vilardaga J.-P. , Bünemann M. , Lohse M.J.
Published: October 2004

The kinetics of G-protein-coupled receptor activation and deactivation has, so far, been measured only indirectly, most frequently by assessing the production of various second messengers. We have developed methods based on fluorescence resonance energy transfer to quantify the kinetics of receptor activation by agonist (measured as conformational change in the receptor), the kinetics of G-protein activation (measured as G-protein subunit rearrangement) and the kinetics of receptor inactivation by arrestins (measured as receptor–arrestin interaction). Using these methods, we show that receptor activation by agonists and signalling to G-proteins occur on the subsecond time scale, whereas receptor desensitization is limited by receptor phosphorylation and proceeds more slowly.
Small GTP-binding protein-coupled receptors
Authors: Bhattacharya M. , Babwah A.V. , Ferguson S.S.G.
Published: October 2004

Heterotrimeric GPCRs (G-protein-coupled receptors) form the largest group of integral membrane receptor proteins and mediate diverse physiological processes. In addition to signalling via heterotrimeric G-proteins, GPCRs can also signal by interacting with various small G-proteins to regulate downstream effector pathways. The small G-protein superfamily is structurally classified into at least five families: the Ras, Rho/Rac/cdc42, Rab, Sar1/Arf and Ran families. They are monomeric G-proteins with molecular masses over the range 20–30 kDa, which function as molecular switches to control many eukaryotic cell functions. Several studies have provided evidence of crosstalk between GPCRs and small G-proteins. It is well documented that GPCR signalling through heterotrimeric G-proteins can lead to the activation of Ras and Rho GTPases. In addition, RhoA, Rabs, ARFs and ARF GEFs (guanine nucleotide-exchange factors) can associate directly with GPCRs, and GPCRs may also function as GEFs for small GTPases. In this review, we summarize the recent progress made in understanding the interaction between GPCRs and small GTPases, focusing on understanding how the association of small G-proteins with GPCRs and GPCR-regulatory proteins may influence GPCR signalling and intracellular trafficking.
Participation of RGS8 in the ternary complex of agonist, receptor and G-protein
Authors: Benians A. , Nobles M. , Tinker A.
Published: October 2004

The RGS (regulators of G-protein signalling) protein family sharpen signalling kinetics through heterotrimeric G-proteins by enhancing the GTPase activity of the G-protein α subunit. Paradoxically, they also accelerate receptor-stimulated activation. We investigated this paradox using the cloned G-protein gated K+ channel as a reporter of the G-protein cycle, and FRET (fluorescence resonance energy transfer) between cyan and yellow fluorescent protein tagged proteins to detect physical interactions. Our results with the neuronal protein, RGS8, show that the enhancement of activation kinetics is a variable phenomenon determined by receptor type, G-protein isoform and RGS8 expression levels. In contrast, deactivation was consistently accelerated after removal of agonist. FRET microscopy revealed a stable physical interaction between RGS8-yellow fluorescent protein and Go αA-cyan fluorescent protein that occurred in the presence and absence of receptor activation and was not competed away by Gβγ overexpression. FRET was also seen between RGS8 and Gγ, demonstrating that RGS8 binds to the heterotrimeric G-protein as well as G-protein α subunit-GTP and the transition complex. We propose a novel model for the action of RGS proteins on the G-protein cycle involving participation of the RGS in the ternary complex: for certain combinations of agonist, receptor and G-protein, RGS8 expression improves upon the ‘kinetic efficacy’ of G-protein activation.
Oscillations in transcription factor dynamics: a new way to control gene expression
Authors: Nelson D.E. , Sée V. , Nelson G. , White M.R.H.
Published: October 2004

Oscillations in second-messenger signalling (e.g. calcium) have previously been shown to be important in the control of transcription. More recently, oscillations in localization and absolute levels of transcription factors and their regulators have been identified. Here we discuss the role of network motifs such as the negative feedback loop and their role in oscillatory signalling, and how oscillations in components of the nuclear factor κB signalling pathway are important to the dynamic control of transcription in response to a cytokine stimulus.
The importance of butyrate transport to the regulation of gene expression in the colonic epithelium
Authors: Cuff M.A. , Shirazi-Beechey S.P.
Published: October 2004

Butyrate is a naturally occurring monocarboxylate, produced in the lumen of the colon by microbial fermentation of complex carbohydrates that escape digestion in the small intestine. It serves as the principal metabolic fuel for colonic epithelial cells, and exerts a variety of effects important to intestinal health and function. This brief discussion focuses on the route, role and regulation of butyrate transport in the large intestine, with particular emphasis on the significance of butyrate transport to the ability of butyrate to modulate expression of genes important to the processes maintaining colonic tissue homoeostasis.
The telling tail of L-selectin
Authors: Ivetic A. , Ridley A.J.
Published: October 2004

L-selectin is constitutively expressed on the surface of most leucocytes and is important for tethering and subsequent rolling of leucocytes on endothelial cells, facilitating their migration into secondary lymphoid organs (e.g. naive T cells) and sites of inflammation (e.g. neutrophils). Previous studies have shown that the 17-amino-acid L-selectin cytoplasmic tail is important for its function in cell adhesion and, hence, identifying binding partners will provide insight into how L-selectin is regulated in leucocytes. This review describes currently known binding partners of the L-selectin tail and how their associations affect L-selectin function.
The aggregation and membrane-binding properties of an α-synuclein peptide fragment
Authors: Madine J. , Doig A.J. , Middleton D.A.
Published: October 2004

α-Synuclein is a 140 amino acid protein, which is associated with presynaptic membranes in the brain, and is the major component of protein aggregates produced during the progression of many neurodegenerative diseases. It has been shown that a central hydrophobic region of α-synuclein comprising residues 71–82 is required for aggregation of the protein into the fibrillar form found in pathogenic aggregates [Giasson, Murray, Trojanowski and Lee (2001) J. Biol. Chem. 276, 2380–2386]. In the present study, we used 2H NMR and electron microscopy to investigate the aggregation and membrane-binding properties of a synthetic peptide corresponding to this region. Results indicate that this region associates with phospholipid bilayers but also forms amyloid-like fibrils in the absence of lipid membranes.
Structural and magnetic properties of insulating Zn1−xCoxO thin films
Authors: Yin Zhigang , Chen Nuofu , Chai Chunlin , Yang Fei
Published: October 2004

Cobalt-doped ZnO (Zn1−xCoxO) thin films were fabricated by reactive magnetron cosputtering. The processing conditions were carefully designed to avoid the occurrence of Co precipitations. The films are c-axis oriented, and the solubility limit of Co in ZnO is less than 17%, determined by x-ray diffraction. X-ray photoemission spectroscopy measurements show Co ions have a chemical valance of 2+. In this paper, hysteresis loops were clearly observed for Zn1−xCoxO films at room temperature. The coercive field, as well as saturation magnetization per Co atom, decreases with increasing Co content, within the range of 0.07<x<0.17. Most intriguing, the Zn1−xCoxO films are nonconductive as x is no more than 17%. Our results clearly demonstrate that ferromagnetism can be realized in Zn1−xCoxO without carrier incorporation.
Effect of the substrate temperature on the properties of RE–Fe–B thin film magnets
Authors: Castaldi L. , Gibbs M. R. J. , Davies H. A.
Published: October 2004

We report a systematic study of the structural, morphological, and magnetic properties of RE–Fe–B films prepared by magnetron sputtering on thermally oxidized Si substrates heated to temperatures Ts in the range between Ts∼470 and ∼800°C during the deposition. The combination of crystallographic texture and mechanical and hard magnetic properties was observed for Ts∼470°C, which results in better mechanical, textural, and hard magnetic properties in terms of perpendicular remanence (0.97T), coercivity (420kA∕m), and maximum energy product (136kJ∕m3).
Third order aberration theory of double Wien filters
Authors: Ioanoviciu D. , Tsuno K. , Martinez G.
Published: October 2004

The second and the third order aberration theory for a double Wien filter have been analytically developed. A new second order aberration-free condition is found at the image plane of the second filter. This condition is met when b2=−1/4, e2=−1/2, and b3−e3=−1/8, where b2=B2R/B1, e2=E2R/E1, b3=B3R2/B1, and e3=E3R2/E1. Here, R is the cyclotron radius and E1, B1, E2, B2, E3, and B3 are the dipole, quadrupole, and hexapole components of electric and magnetic fields, respectively. This condition is different from the second order aberration-free condition for a single Wien filter, which is satisfied when b2=−3/4, e2=−1, and b3−e3=−3/8. The geometrical second order aberration-free condition has also been found, and requires that e3−b3=(m−1)/8, e2=−m/4, and b2=(1−m)/4. This last set is sufficient to satisfy the above two sets of conditions as well. Residual third order aberrations are calculated for various m. The third order aberrations at the second focus are very small when the new aberration-free condition is fulfilled.
Nonlocal electrostatics in heterogeneous suspensions using a point-dipole model
Authors: von Pfeil Karl , Klingenberg Daniel J.
Published: October 2004

The electrostatic dipole moment distribution in heterogeneous suspensions is determined via a self-consistent, point-dipole model, which incorporates nonlocal electrostatics. Predictions agree qualitatively with previous asymptotic results for discontinuous concentration profiles. For small fluctuations in concentration, the dipole strength can be expressed as an expansion in gradients of the concentration. This expansion is incorporated into a linearized continuum model for structure evolution in sheared electrorheological suspensions. Prior stability analysis of a fully local continuum model predicts the formation of concentrated particle stripes oriented in the flow direction, in agreement with experimental observations. Incorporating nonlocal electrostatics suppresses the growth of high wave number fluctuations, providing a more realistic finite rate of growth of fluctuations. Incorporating nonlocal electrostatics in the full nonlinear continuum model produces a single particulate stripe at steady state.
Residual gas analysis of a dc plasma for carbon nanofiber growth
Authors: Cruden Brett A. , Cassell Alan M. , Hash David B. , Meyyappan M.
Published: October 2004

We report the analysis of a plasma enhanced chemical vapor deposition process for carbon nanofiber growth. A direct current (dc) plasma is employed with a mixture of acetylene and ammonia. Residual gas analysis is performed on the downstream plasma effluent to determine degrees of precursor dissociation and high molecular weight species formation. Results are correlated to growth quality obtained in the plasma as a function of dc voltage∕power, gas mixture, and pressure. Behaviors in plasma chemistry are understood through application of a zero-dimensional model.
X-ray diffraction on laterally modulated (InAs)n∕(AlAs)m short-period superlattices
Authors: Caha O. , Křápek V. , Holý V. , Moss S. C. , Li J. H. , Norman A. G. , Mascarenhas A. , Reno J. L. , Stangl J. , Meduňa M.
Published: October 2004

Lateral composition modulation in InAs∕AlAs short-period superlattices was investigated by x-ray grazing-incidence diffraction and coplanar x-ray diffraction at a “normal” wavelength and at an anomalous wavelength, for which diffraction from the (200) planes does not exhibit a chemical contrast. The experimental data were compared with theoretical simulations assuming that the interfaces consist of a periodic sequence of monoatomic steps. The displacement field in the superlattice was calculated by continuum elasticity and using a valence-force field method. From the fit of the experimental data to the theory, the lengths of individual atomic terraces were determined.
Effect of impurity incorporation on crystallization in AlN sublimation epitaxy
Authors: Kakanakova-Georgieva A. , Gueorguiev G. K. , Yakimova R. , Janzén E.
Published: October 2004

We have implemented graphite, graphite-tantalum (Ta), and Ta growth environment to the sublimation epitaxy of aluminum nitride (AlN) and have studied development, morphological, and cathodoluminescence emission properties of AlN crystallites. Three apparently different types of crystallites form in the three different types of growth environment, which presumably manifests the relationship between crystallite-habit-type and impurities. Comparison between the cathodoluminescence spectra reveals certain dynamics in the incorporation into AlN of the main residual dopants, oxygen and carbon, when the growth environment changes. At high temperatures, in addition to Al and N2, which constitute the vapor over AlN, vapor molecules of CN, NO, Al2C, and many more can be present in the vapor from which AlN grows and both oxygen and carbon can be incorporated into AlN in varying ratios. Involving calculations of the cohesive energy per atom of such vapor molecules and also of Ta containing molecules, we have considered possible mechanisms how oxygen and carbon get incorporated into AlN and how this kinetics interferes with the growth environment. The positive effect of Ta consists in the marked reduction of residual oxygen and carbon impurities in the vapor from which AlN is growing. However, on the account of this reduction, the overall composition of the vapor changes. We speculate that during AlN nucleation stage small impurity levels may be beneficial in order to provide a better balance between the AlN crystallites development and impurity incorporation issues. We have shown that some impurity containing vapor molecules are acting as essential transport agents and suppliers of nitrogen for the AlN growth.
Dynamical properties of the Q-controlled atomic force microscope
Authors: Kokavecz János , Horváth Zoltán L. , Mechler Ádám
Published: October 2004

In intermittent contact mode atomic force microscopy (AFM), the quality factor (Q) of the oscillating probe is believed to account for the imaging speed and sensitivity. Q control is a method to artificially modify the quality factor of the probe. Here, we present a comprehensive study of the dynamics of the Q-controlled AFM. By comparing the analytical solutions of the force equations, we prove that the Q-controlled and non-Q-controlled systems are equivalent in the absence of surface forces. We also determine the conditions for the numerical simulation. In order to study the mechanism of contrast enhancement, we simulate the normal AFM operation including the surface forces. We found that there is a maximal probe sensitivity which cannot be exceeded even with Q control. Consistently, Q control enhances sensitivity only when imaging soft samples. Finally, we show that the phase signal of the Q-controlled system is more sensitive to the changes of the sample properties than in case of non-Q-controlled AFMs.
Optimized GaAs∕AlGaAs light-emitting diodes and high efficiency wafer-fused optical up-conversion devices
Authors: Ban D. , Luo H. , Liu H. C. , Wasilewski Z. R. , SpringThorpe A. J. , Glew R. , Buchanan M.
Published: October 2004

Achieving a high internal quantum efficiency in GaAs∕AlGaAs based light-emitting diodes (LEDs) for room-temperature operation at low current-density injection is crucial for applications such as optical up-converters based on the integration of LEDs and photodetectros. We report the experimental results as well as the theoretical analyses of the internal quantum efficiency of GaAs∕AlGaAs LEDs as a function of the p-doping concentration of the active region for low current injection operation. By optimizing the doping concentration, we have achieved a close to 100% internal quantum efficiency for room-temperature operation of LEDs in the low injection current-density range, i.e., around 0.1A∕cm2. An optical up-converter was fabricated using wafer-fusion technology by integrating the optimized GaAs∕AlGaAs LED with an InGaAs∕InP photodetector. The internal up-conversion quantum efficiency was measured to be 76%.
Characteristics of silicone fluid as a pressure transmitting medium in diamond anvil cells
Authors: Shen Yongrong , Kumar Ravhi S. , Pravica Michael , Nicol Malcolm F.
Published: October 2004

The properties of a silicone fluid with initial viscosity of 1 cst as a pressure transmitting medium for diamond anvil cells have been determined by ruby R1 line broadening and R1-R2 separation measurements to 64 GPa at ambient temperature. By these criteria, the silicone fluid is as good a pressure medium as a 4:1 methanol:ethanol mixture at low pressures to about 20 GPa, and is better than the mixture at higher pressures. Although argon media are better than the silicone at pressures to 30 GPa, this silicone behaves as well as argon at higher pressures. Furthermore, the silicone is easier to load than argon and is almost chemically inert.