bluuebunny/crossref_metadata_2025 huggingface dataset:
tRNA–ribosome interactions
Authors: Ehrenberg Måns , Bilgin Nese , Dincbas Vildan , Karimi Reza , Hughes Diarmaid , Abdulkarim Farhad
Published: December 2009

Direct measurements of the rates of dissociation of dipeptidyl-tRNA from the ribosome show that hyperaccurate SmP and SmD ribosomes have unstable A-site binding of peptidyl-tRNA, while P-site binding is extremely stable in relation to the wild type. Error-prone Ram ribosomes, on the other hand, have stable A-site and unstable P-site binding of peptidyl-tRNA. At least for these mutant ribosomes, we conclude that stabilization of peptidyl-tRNA in one site destabilizes binding in the other. Elongation factor Tu (EF-Tu) undergoes a dramatic structural transition from its GDP-bound form to its active GTP-bound form, in which it binds aa-tRNA (aminoacyl-fRNA) in ternary complex. The effects of substitution mutations at three sites in domain I of EF-Tu, Gln124, Leu120, and Tyr160, all of which point into the domain I – domain III interface in both the GTP and GDP conformations of EF-Tu, were examined. Mutations at each position cause large reductions in aa-tRNA binding. An attractive possibility is that the mutations alter the domain I – domain III interface such that the switching of EF-Tu between different conformations is altered, decreasing the probability of aa-tRNA binding. We have previously found that two GTPs are hydrolyzed per peptide bond on EF-Tu, the implication being that two molecules of EF-Tu may interact on the ribosome to catalyze the binding of a single aa-tRNA to the A-site. More recently we found that ribosomes programmed with mRNA constructs other than poly(U), including the sequence AUGUUUACG, invariably use two GTPs per peptide bond in EF-Tu function. Other experiments measuring the protection of aa-tRNA from deacylation or from RNAse A attack show that protection requires two molecules of EF-Tu, suggesting an extended ternary complex. To remove remaining ambiguities in the interpretion of these experiments, we are making direct molecular weight determinations with neutron scattering and sedimentation–diffusion techniques.Key words: ribosome, EF-Tu, accuracy, in vitro translation.
Translational regulation of infC operon in Bacillus stearothermophilus
Authors: Pediconi Dario , Spurio Roberto , Teana Anna La , Jemiolo David , Gualerzi Claudio O. , Pon Cynthia L.
Published: December 2009

A Bacillus stearothermophilus in vitro translational system has been developed to study the expression of the three cistrons (infC, rpmI, and rplT) constituting the infC operon of this bacterium. When directed by homologous in vitro transcribed infC tricistronic mRNA, this system, which consists of partially purified and purified components of the B. stearothermophilus translational apparatus, synthesizes with high efficiency and specificity the three gene products (IF3, L35, and L20) in a ratio similar to that found in vivo (i.e., about 1:6:6). The three cistrons are translationally coupled and expressed in a specific temporal order: a low level of IF3 synthesis stimulates the expression of L35 which, in turn, greatly stimulates the synthesis of L20 and IF3. Protein L20 and an excess of IF3 were found to act as translational feedback inhibitors of the entire operon. The synthesis of IF3 displayed a strong dependence on IF2. This dependence as well as the repressibility by excess IF3 were found to be due to the presence of the rare AUU initiation triplet at the beginning of infC.Key words: translational coupling, IF3, IF2, L35, L20.
A pseudoknot is required for efficient translational initiation and regulation of theEscherichia coli rpsOgene coding for ribosomal protein S15
Authors: Ehresmann Chantal , Philippe Claude , Westhof E. , Ehresmann Bernard , Bénard Lionel , Portier Claude
Published: December 2009

Escherichia coli ribosomal protein S15 down regulates its own synthesis by binding to its mRNA in a region overlapping the ribosome binding site, called the translational operator. This binding stabilizes a pseudoknot structure that exists in equilibrium with two stem–loop structures. When synthesized in excess over 16S rRNA, S15 binds to its translational operator and traps the ribosome on its loading site in a transient state, preventing the formation of the active ternary (30S–mRNA–rRNAfMet) complex. This inhibition can be suppressed by 16S rRNA, which displaces S15 from the mRNA. An extensive mutational analysis showed that the pseudoknot is the structural element required for S15 recognition and in vivo translational control. Specific sequence determinants are located in limited regions of the structure formed by the pseudoknot. An unexpected result is that the pseudoknot can exist in a variety of topologically equivalent structures recognizable and shapable by S15. Based on footprinting experiments and computer graphic modelling, S15 shields the two stems of the pseudoknot, sitting in the major groove of the coaxial stack.Key words: ribosomes, translational control, r-protein S15, pseudoknot, RNA–protein recognition.
Fungal ribotoxins: a family of naturally engineered targeted toxins?
Authors: Kao Richard , Davies Julian
Published: December 2009

α-Sarcin, mitogillin, and restrictocin are small (~17 kDa) basic ribosome-inactivating proteins (RIPs) produced by the Aspergilli that catalytically inactivate the large ribosomal subunits of all organisms tested to date. These three fungal ribotoxins act as specific ribonucleases by hydrolyzing one single phosphodiester bond in the universally conserved α-sarcin domain of 23–28S rRNAs and are among the most potent inhibitors of protein synthesis known. Previous molecular studies of ribotoxins indicated that they belong to the superfamily of ribonucleases and analysis of the mitogillin gene employing PCR-mediated site-specific mutagenesis suggests that certain domains in ribotoxins, which share homologies with motifs in ribosome-related proteins, may be responsible for the targeting of ribotoxins to the ribosome. The applications of the ribotoxins as tools in research and their uses as therapeutic and diagnostic agents are also reviewed in this paper.Key words: ribotoxins, mitogillin, α-sarcin loop, protein synthesis, aspergillosis, immunotoxins.
Antibiotic resistance mechanisms of mutant EF-Tu species inEscherichia coli
Authors: Kraal Barend , Zeef Leo A. H. , Mesters Jeroen R. , Boon Kathy , Vorstenbosch Erik L. H. , Bosch Leendert , Anborgh Pieter H. , Parmeggiani Andrea , Hilgenfeld Rolf
Published: December 2009

Analysis of antibiotic-resistant EF-Tu mutants has revealed a connection between resistance and structural elements that participate in the GTPase switching mechanism. Both random and site-directed mutagenesis methods have yielded sets of purified mutant EF-Tu resistant to kirromycin (kirr) or pulvomycin (pulr). All kirrmutations cluster in the interface of domains 1 and 3 of EF-Tu in its GTP-bound conformation, not in that of EF-Tu∙GDP. Other evidence also suggests that kirromycin binds to the interface of wild-type EF-Tu, thereby jamming the GTPase switch. Various functional studies reveal two subsequent resistance mechanisms. The first hinders kirromycin binding to EF-Tu∙GTP and the second occurs after GTP hydrolysis by rejection of bound kirromycin. All pulrmutations cluster in the three-domain junction interface of EF-Tu∙GTP (which is an open hole in EF-Tu∙GDP) and destabilize a salt-bridge network. Pulvomycin may bind nearby and overlap with tRNA binding. Mutations show that a D99–R230 salt bridge is not essential for the transduction of the GTPase switch signal from domain 1. In vivo and in vitro studies reveal that pulvomycin sensitivity is dominant over resistance. This demands a revision of the current view of the mechanism of pulvomycin inhibition of protein synthesis and may support a translation model with two EF-Tus on the ribosome. Several mutant EF-Tu species display altered behaviour towards aminoacyl-tRNA with interesting effects on translational accuracy. KirrEF-Tu(A375T) is able to reverse the streptomycin-dependent phenotype of a ribosomal protein S12 mutant strain to streptomycin sensitivity.Key words: kirromycin, pulvomycin, streptomycin, GTPase switch, aminoacyl-tRNA.
Elongation factor Tu, a GTPase triggered by codon recognition on the ribosome: mechanism and GTP consumption
Authors: Rodnina Marina V. , Pape Tillmann , Fricke Rainer , Wintermeyer Wolfgang
Published: December 2009

The mechanism of elongation factor Tu (EF-Tu) catalyzed aminoacyl-tRNA (aa-tRNA) binding to the A site of the ribosome was studied. Two types of complexes of EF-Tu with GTP and aa-tRNA, EF-Tu∙GTP∙aa-tRNA (ternary) and (EF-Tu∙GTP)2∙aa-tRNA (quinternary), can be formed in vitro depending on the conditions. On interaction with the ribosomal A site, generally only one molecule of GTP is hydrolysed per aa-tRNA bound and peptide bond formed. The second GTP molecule from the quinternary complex is hydrolyzed only during translation of an oligo(U) tract in the presence of EF-G. The first step in the interaction between the ribosome and the ternary complex is the codon-independent formation of an initial complex. In the absence of codon recognition, the aa-tRNA–EF-Tu complex does not enter further steps of A site binding and remains in the initial binding state. Despite the rapid formation of the initial complex, the rate constant of GTP hydrolysis in the noncognate complex is four orders of magnitude lower compared with the cognate complex. This, together with the results of time-resolved fluorescence measurements, suggests that codon recognition by the ternary complex on the ribosome initiates a series of structural rearrangements that result in a conformational change of EF-Tu, presumably involving the effector region, which, in turn, triggers GTP hydrolysis and the subsequent steps of A site binding.Key words: translation, A site, codon recognition, fluorescence, stopped-flow.
Peptide hormone evolution: functional heterogeneity within GnRH and CRF families
Authors: Lovejoy David A.
Published: December 2009

Recent investigations indicate that the gonadotropin-releasing hormone (GnRH) and corticotropin-releasing factor (CRF) family of peptides are each composed of at least two functionally discrete paralogous lineages. [His5Trp7Tyr8]GnRH (chicken GnRH-II) is associated with brain neuromodulatory and possibly peripheral endocrine activity, whereas [Arg8]GnRH (mammal GnRH) and its orthologues play major roles as hypothalamic releasing factors. Similarly, CRF appears to be the primary vertebrate ACTH-releasing peptide, whereas the paralogous lineage of urotensin-I-sauvagine has been associated with a variety of diverse peripheral activities. In phylogenetically older species, representatives of both GnRH and CRF family lineages have been characterized. Structural and functional conservation of these peptide systems in vertebrates suggest that additional GnRH-like and CRF-like peptides will be found in the mammal brain.Key words: neuropeptides, evolution, reproduction, HPA axis, stress, receptor–ligand interactions.
Radiation inactivation and in situ renaturation of protein tyrosine kinases reveal a major 50-kDa enzyme as part of a membrane complex present in dividing but not in resting prostatic epithelial cells
Authors: Nguyen Linh The , Durocher Yves , Beauregard Guy , Tessier Sylvain , Atfi Azeddine , Allard Pierre , Chapdelaine Alcide , Potier Michel , Chevalier Simone
Published: December 2009

Because protein tyrosine kinases play a crucial role in the regulation of cell division and carcinogenesis, we have herein measured such enzyme activities (specific activity and subcellular distribution) and compared their characteristics with respect to hydrodynamic properties and radiation inactivation sizes as well as renaturation after electrophoresis in denaturing conditions in canine prostatic epithelial cells either in a resting (freshly isolated) or in a dividing (cultured cells) state. In quiescent cells, most protein tyrosine kinase activity was expressed by soluble proteins with a Stokes' radius (Rs) of 3.05 nm, a sedimentation coefficient (S20,w) of 4.0 S, and a molecular mass of 50 kDa. By contrast, in dividing cells (three days in primary culture), the specific activity was higher and the enzyme was mainly membrane bound. The use of a detergent (Triton X-100) allowed the extraction of most of that enzyme; its partial specific volume, S20,w, and Rswere then 0.883 cm3/g, 4.0 S, and 5.6 nm, respectively, hence yielding a molecular mass of 215 kDa, which decreased to 125–145 kDa when corrected for detergent binding. Probing these chromatography-peak fractions, 50 kDa from cytosol of resting cells and 215 kDa from membrane extracts of dividing cells, with a phosphotyrosine antibody following their incubation with ATP and electrophoresis in denaturing conditions revealed the presence of a common 50-kDa phosphotyrosylated protein along with three other bands (130, 75, and 40 kDa) in the high-Mr peak of enzyme. However, the radiation inactivation size for protein tyrosine kinases expressed in both resting and dividing cells were similar, 47.2 ± 8.7 and 44.5 ± 6.1 kDa, respectively. Furthermore, by renaturation after electrophoresis in denaturing conditions, major protein tyrosine kinase polypeptides of 50 kDa were identified in both cell populations. Taken together, these results indicate that, in dividing prostatic epithelial cells, membrane-bound protein tyrosine kinases of low molecular weight with properties similar to those of monomeric soluble forms present in quiescent cells are part of high-molecular weight complexes. This activation process may be critical for hormone-independent proliferation of prostatic epithelial cells.Key words: protein tyrosine kinase, kinase renaturation, cell division, prostate, radiation inactivation.
A polysaccharide–peptide complex from cultured mycelia of the mushroom Tricholoma mongolicum with immunoenhancing and antitumor activities
Authors: Wang H. X. , Ooi V. E. C. , Chang S. T. , Ng T. B. , Liu W. K.
Published: December 2009

A polysaccharide–peptide complex with immunoenhancing and antitumor activities was obtained from the mycelial culture of Tricholoma mongolicum, an edible mushroom found in Northern China. The polysaccharide–peptide complex had a molecular mass of 15.5 kDa, as estimated by gel filtration, and a carbohydrate–protein ratio of about 8:1 and was not adsorbed on DEAE – Sepharose CL-6B. It possessed the activities of activating macrophages, stimulating macrophage antigen-presenting activity, which in turn enhanced proliferation of T-cells, and inhibiting the growth of sarcoma 180 cells that had been implanted in mice.Key words: Tricholoma mongolicum, immunoenhancing activity, antitumor activity, polysaccharide–peptide complex.
Purification and metal ion requirements of a candidate matrix metalloproteinase: a 41 kDa gelatinase activity in the sea urchin embryo
Authors: Mayne Janice , Robinson John J.
Published: December 2009

Using substrate gel zymography, the sea urchin embryo was found to express a dynamic pattern of gelatinase activities with a 41 kDa species persisting throughout the course of embryonic development. We have purified to near homogeneity the 41 kDa gelatinase in the sea urchin egg. In both qualitative and quantitative assays, the 41 kDa gelatinase activity was inhibited by ethylenediaminetetracetic acid but not the serine protease inhibitor, phenylmethylsulfonylfluoride, or the chelating agent, 1,10-phenanthroline. Activity could be restored to the inactivated gelatinase by each of several divalent cations: Ca2+ > Mn2+ > Mg2+ > Cu2+. Cadmium and Zn2+were largely ineffective at reconstituting the inactivated enzyme. In metal ion binding assays, the relative apparent affinities of the metal ions for binding to the gelatinase were determined to be Zn2+ ≥ Cd2+ ≥ Ca2+ > Mn2+ > Mg2+ > Cu2+. While the gelatinase is clearly a metalloproteinase, metal ion binding per se is not sufficient for activity. The 41 kDa gelatinase exhibited selective substrate utilization, being most active with gelatin, substantially less active with casein, and inactive towards bovine haemoglobin and bovine serum albumin as substrates. The substrate specificity and metal ion requirements suggest that this species is a member of the matrix metalloproteinase class of extracellular matrix remodelling enzymes.Key words: gelatinase, metalloproteinase, sea urchin.
Ferriheme and ferroheme are isosteric inhibitors of fatty acid binding to rat liver fatty acid binding protein
Authors: Stewart John M. , Slysz Gordon W. , Pritting Mary Anne , Muller-Eberhard Ursula
Published: December 2009

In addition to fatty acids, liver fatty acid binding protein (L-FABP) also interacts with ferriheme, which it binds with an affinity approximately one order of magnitude greater than that for oleic acid. We have, therefore, examined the effect of ferroheme and ferriheme on the binding of oleate to rat L-FABP, also called heme-binding protein. Both oxidation states of heme behaved as isosteric inhibitors for the binding of the fatty acid confirming a common binding site. The reduced form of heme (Fe(II)) is a threefold better competitor of oleate binding than ferriheme. To show whether the diffusion of heme would be affected by the presence of the binding protein, we measured the effect of the fatty acid binding protein on the diffusional flux of a water-soluble heme derivative, iron-deuteroporphyrin. The diffusional flux of iron-deuteroporphyrin did not change in the presence of the protein. This suggested that the binding affinity of fatty acid binding protein for iron-deuteroporphyrin is too great to allow rapid equilibrium between bound and unbound ligand across the system in an appropriate time frame.Key words: fatty acid binding protein, heme, diffusion, liver, oleic acid.
Glucose inhibits phenobarbital-induced δ-aminolevulinate synthase expression in normal but not in diabetic rat hepatocytes
Authors: Varone Cecilia L. , Cánepa Eduardo T. , Llambías Elena B. C. , Grinstein Moisés
Published: December 2009

In the present work, we demonstrate the presence of a glucose inhibitory effect on the phenobarbital-mediated induction of the δ-aminolevulinate synthase mRNA in normal rat hepatocytes, consistent with the results obtained with the δ-aminolevulinate synthase activity previously reported. This "glucose effect" can be prevented by adding cAMP, adenylate cyclase activators, or a phosphodiesterase inhibitor. δ-Aminolevulinate synthase mRNA half-life is not modified in the presence of phenobarbital or glucose. When the same experiments are performed using diabetic cells, no glucose effect is observed, even when the endogenous cAMP content is lowered to normal levels. The results obtained in this study suggest that glucose decreases δ-aminolevulinate synthase biosynthesis by acting at a pretranslational step. Assuming that the glucose effect operates by a repression mechanism exerted by metabolites derived from or related to glucose, the present results may reflect a derangement in the formation of these metabolites as a result of the abnormal metabolism operating in the diabetic state.Key words: glucose, δ-aminolevulinate synthase expression, diabetic rat hepatocytes, phenobarbital, cAMP.
Mitogenic stimulation of primary cultures of lung epithelial cells by linoleic acid
Authors: Usha S. , Maya A. , Balakrishnan A.
Published: December 2009

The role of linoleic acid (18:2 n-6) in stimulating proliferation of normal lung epithelial cells in vitro is investigated. When 18:2 n-6 is present with insulin (I) and cholera toxin (CT), growth is stimulated synergistically. In the presence of indomethacin (10 μM), an inhibition of proliferation is observed in I, CT, and 18:2 n-6, which can be reversed by the addition of exogenous prostaglandin E2(PGE2). Incorporation of [14C]18:2 n-6 with lipid-independent I, CT, and cortisol and lipid-dependent I, CT, and 18:2 n-6 conditions suggests differences in mobilization of 18:2 n-6 from the phospholipid (PL) fractions between 2 and 8 days. The decline of [14C]18:2 n-6 in PL fractions with lipid-dependent condition suggests that free 18:2 n-6 may be available for metabolism by the cyclooxygenase pathway. In non-proliferative cultures, an accumulation of the label in the PL fraction is observed. Proliferation in lipid-dependent conditions appears to be due to the mobilization of 18:2 n-6 whereas proliferation in lipid-independent conditions appears to be independently controlled.Key words: linoleate metabolism, proliferation, lung epithelial cell cultures.
Signal transduction and TGF-β superfamily receptors
Authors: Kolodziejczyk Steven M. , Hall Brian K.
Published: December 2009

The TGF-β superfamily includes a large number of related growth and differentiation factors expressed in virtually all phyla. Superfamily members bind to specific cell surface receptors that activate signal transduction mechanisms to elicit their effects. Candidate receptors fall into two primary groups, termed type I and type II receptors. Both types are serine/threonine kinases. Upon activation by the appropriate ligand, type I and type II receptors physically interact to form hetero-oligomers and subsequently activate intracellular signaling cascades, ultimately regulating gene transcription and expression. In addition, TGF-β binds to a third receptor class, type III, a membrane-anchored proteoglycan lacking the kinase activity typical of signal transducing molecules. Type III receptors appear to regulate ligand availability to type I and type II receptors. Although a number of transduction mechanisms may be available to TGF-β superfamily members, evidence gathered through the use of specific kinase and G-protein inhibitors and through assays measuring activation and levels of signaling intermediates suggests that at least one signaling pathway interacts with Ras and Raf proteins via a G-protein intermediate. Raf begins the cytoplasmic kinase cascade that leads to gene regulation. The myriad responses regulated by TGF-β superfamily members makes the understanding of signal transduction mechanisms utilized by these proteins of great interest to a wide range of biological disciplines.Key words: TGF-β superfamily, serine/threonine kinase receptors, G-proteins, Ras, cytoplasmic kinase cascade.
The yeast nucleoporin Nsp1 binds nuclear localization sequences in vitro
Authors: Barth Werner , Stochaj Ursula
Published: December 2009

Facilitated transport of proteins into the nucleus requires nuclear localization sequences (NLSs) be present in the protein destined for the nucleus. The specific binding of NLSs by components of the nuclear transport apparatus is essential for these targeting reactions. We now report that the yeast nucleoporin Nsp1 binds specifically nuclear localization sequences in vitro. This nucleoporin recognizes several NLSs that are functional for nuclear targeting in vivo, including the NLS of SV40 T-antigen and of the yeast transcription factor Gal4. Nsp1 is organized into three domains, and we have located NLS binding sites to the N-terminal portion and the middle repetitive region of the protein. For the interaction between the NLS of SV40 T-antigen and Nsp1, we obtained association constants of 1.2 × 107 M−1and 5 × 107 M−1. An association constant of 5 × 107 M−1was determined for NLS binding to the repetitive domain of Nsp1. We analyzed binding of Nsp1 and its domains to a mutant version of the NLS derived from SV40 T-antigen, which poorly functions for nuclear targeting in vivo. The affinity for the mutant signal was about two orders of magnitude lower than for the wild-type NLS.Key words: Nsp1, nuclear pore complex, nucleoporin, nuclear localization sequence, protein targeting, yeast.
Integrity of the homophilic binding site is required for the preferential localization of NCAM in intercellular contacts
Authors: Sandig Martin , Rao Yong , Siu Chi-Hung , Kalnins Vitauts I.
Published: December 2009

The neural cell adhesion molecule NCAM is a member of the immunoglobulin (Ig) superfamily. NCAM can undergo homophilic binding and heterophilic interactions with cell surface components and is often concentrated at sites of intercellular contact. To investigate the molecular basis of this biased surface distribution, we examined L cell transfectants expressing wild-type or mutant forms of chick NCAM-140 by laser scanning confocal microscopy. Mutant NCAMs that lacked Ig-like domains 1, 2, 4, or 5 were preferentially localized in contact regions. However, the relative concentration of these mutant NCAMs in contact sites was substantially reduced compared with wild-type NCAM. In contrast, NCAM redistribution to intercellular contacts was abolished in cells expressing mutant NCAMs that either lacked Ig-like domain 3 or contained mutations in the homophilic binding site in this domain. In heterotypic contacts between PC12 cells and L cell transfectants, colocalization of rat NCAM and chick NCAM was again dependent on the integrity of the homophilic binding site of the NCAM expressed on L cells. These results provide evidence that homophilic binding is the main mechanism by which NCAM becomes redistributed to intercellular contacts. They also implicate a role for other Ig-like domains in the accumulation of NCAM at cell–cell contacts.Key words: cell–cell adhesion, adhesion molecule, NCAM, homophilic binding, surface distribution.
Neuronal Cdc2-like kinase: from cell cycle to neuronal function
Authors: Tang Damu , Matsuura Isao , Wang Jerry H. , Lee Ki-Young , Qi Zhong
Published: December 2009

Neuronal Cdc2-like kinase, Nclk, is a heterodimer of cyclin-dependent protein kinase 5 (Cdk5) and a 25-kDa essential regulatory subunit that is derived from a 35-kDa brain- and neuron-specific protein. This protein is called neuronal Cdk5 activator, p25/35nck5a. Nclk is one of the best characterized Cdc2 family kinases whose primary function is not cell cycle related. It has been suggested that this protein kinase plays important roles in neurocytoskeleton dynamics and its loss of regulation has been implicated in Alzheimer pathology. As a member of the Cdc2-like kinase family, Nclk shares many common properties with other members of the Cdc2-like kinase family. It also possesses unique characteristics that may be related to its distinct and noncell cycle related functions. The regulatory and functional properties of Nclk are reviewed in this communication.Key words: Cdc2 kinase, Cdk5, neuronal Cdk5 activator.
Phosphorylation of the carboxyl-terminal region of dystrophin
Authors: Michalak Marek , Fu Susan Y. , Milner Rachel E. , Busaan Jody L. , Hance Jacqueline E.
Published: December 2009

Dystrophin is a protein product of the gene responsible for Duchenne and Becker muscular dystrophy. The protein is localized to the inner surface of sarcolemma and is associated with a group of membrane (glyco)proteins. Dystrophin links cytoskeletal actins via the dystrophin-associated protein complex to extracellular matrix protein, laminin. This structural organization implicates the role of dystrophin in stabilizing the sarcolemma of muscle fibers. Precisely how dystrophin functions is far from clear. The presence of an array of isoforms of the C-terminal region of dystrophin suggests that dystrophin may have functions other than structural. In agreement, many potential phosphorylation sites are found in the C-terminal region of dystrophin, and the C-terminal region of dystrophin is phosphorylated both in vitro and in vivo by many protein kinases, including MAP kinase, p34cdc2kinase, CaM kinase, and casein kinase, and is dephosphorylated by calcineurin. The C-terminal domain of dystrophin is also a substrate for hierarchical phosporylation by casein kinase-2 and GSK-3. These observations, in accordance with the finding that the cysteine-rich region binds to Ca2+, Zn2+, and calmodulin, suggest an active involvement of dystrophin in transducing signals across muscle sarcolemma. Phosphorylation–dephosphorylation of the C-terminal region of dystrophin may play a role in regulating dystrophin–protein interactions and (or) transducing signal from the extracellular matrix via the dystrophin molecule to the cytoskeleton.Key words: Duchenne muscular dystrophy, protein phosphorylation, protein kinases, calcineurin, cytoskeleton.
Regulation of transcription factor activity during cellular aging
Authors: Wheaton Keith , Atadja Peter , Riabowol Karl
Published: December 2009

Several lines of evidence suggest that the limited replication potential of normal human cells is due to the presence of an intrinsic genetic programme. This "senescence programme" is believed to reduce the incidence of cancer by limiting the growth of most of the transformed cells arising in vivo, although some cells do escape senescence becoming both immortalized and transformed. Here we review the literature that describes the senescence process in terms of gene expression and the regulation of gene expression by a variety of mechanisms affecting transcription factor activity. We focus on regulation of the c-fos gene through posttranslational modification of the serum response factor (SRF) as an example of altered gene expression during cellular aging.Key words: cellular aging, transcription, Fos, SRF, phosphorylation.
Cyclic AMP phosphodiesterase: a regulator of forward motility initiation during epididymal sperm maturation
Authors: Jaiswal Bijay S. , Majumder Gopal C.
Published: December 2009

The concentrations of cAMP, cAMP phosphodiesterase (PDE) activity, and the effect of theophylline in vitro on the forward motility (FM) of maturing goat epididymal sperm have been analyzed. cAMP levels increased slowly during transit of the cells from the caput to the proximal cauda, although they acquired a minimal degree of forward progression. The last phase of sperm transit (proximal to distal cauda) was associated with a concomitant sharp rise in the level of both c AMP as well as flagellar motility. PDE activity progressively decreased (approximately threefold) during epididymal maturation, being minimal in mature cauda sperm. Theophylline (30 mM), a specific inhibitor of PDE, markedly activated (10-fold or greater) motility of the sperm derived from proximal-corpus, mid-corpus, distal-corpus, and proximal-cauda epididymides. FM of the native mature caudal sperm was similar to that of the theophylline-treated proximal-cauda sperm. The terminal stage of sperm maturity (proximal to distal cauda) was associated with a markedly reduced level of theophylline-dependent motility activation (approximately 50%). The data are consistent with the view that PDE plays an important role in the initiation of motility during epididymal sperm maturation.Key words: epididymal sperm, cyclic AMP, cyclic AMP phosphodiesterase, flagellar motility, theophylline
Transfer of cholesterol between high density lipoproteins and cultured rat Sertoli cells
Authors: Fofana Mohamed , Maboundou Jean-Claude , Bocquet Jean , Goff Dominique Le
Published: December 2009

In the testes, the Sertoli cells are separated from the blood capillaries by the basement membrane, thereby excluding the passage of low density lipoproteins (LDLs) but allowing the passage of high density lipoproteins (HDLs). The present study examines first the capacity of Sertoli cells to uptake cholesterol from HDL and secondly the role of apolipoproteins (apo) A-I and E in cholesterol flux between HDL and cultured rat Sertoli cells. In the presence of HDL in cultured medium, rat Sertoli cells accumulated few amounts of esterified cholesterol. Incubation of [14C]cholesterol–labelled Sertoli cells with [3H]cholesterol–labelled HDL showed that the amount of cholesterol influx slightly exceeded its efflux, thus resulting in a net uptake of cholesterol from HDL to rat Sertoli cells. The amount of HDL–cholesterol converted to steroids by Sertoli cells was about 32% of influx. Uptake of cholesterol by Sertoli cells was three times higher with phospholipid – apo A-I vesicles and seven times higher with phospholipid – apo E vesicles than that with phospholipid vesicles without apolipoprotein. Phospholipid – apo A-I vesicles promoted cholesterol efflux at the same rate as native HDL and twice as efficiently as phospholipid – apo E vesicles. Thus, this study shows that rat Sertoli cells have the capacity to take up HDL–cholesterol for membrane renewal and steroid production mainly by apo E dependent pathways.Key words: apolipoproteins, cholesterol flux, phospholipid vesicle, steroid, testis.
The roles of adhesion molecules and proteinases in lymphocyte transendothelial migration
Authors: Madri Joseph A. , Graesser Donnasue , Haas Tara
Published: December 2009

T cell extravasation into perivascular tissue during inflammation involves transmigration through the endothelial cell (EC) layer and basement membrane. We have demonstrated that matrix metalloproteinase-2 (MMP-2) is induced in T cells upon adhesion to endothelial cells and that the induction of MMP-2 is mediated by binding of T cell VLA-4 to VCAM-1. Cloned murine Th1 cells antigenic to myelin basic protein, either expressing VLA-4 on their cell surface and causing experimental autoimmune encephalomyelitis (EAE) or not expressing VLA-4 and not causing EAE, were used. VLA-4 positive (+) T cells that adhered to VCAM-1 positive (+) endothelial cells exhibited an induction in MMP-2 mRNA, protein, and activity, whereas MMP-2 was not induced in the T cells that adhered to the VCAM-1 negative (−) endothelial cells or VLA-4 negative (−) T cells that adhered to VCAM-1+ endothelial cells. Incubating T cells with rVCAM-1-coated dishes showed that VLA-4+ T cells adhered to the molecule and that adhesion to rVCAM-1 was sufficient to induce MMP-2. VLA-4+ T cells that had transmigrated through a VCAM-1+ endothelial cell monolayer exhibited MMP-2 activity. TTMP-2 was shown to reduce T cell transmigration in vitro. Transmigrated T cells exhibited downregulation of VLA-4 and LFA-1 integrin surface expression and decreased binding to rVCAM-1 and rICAM-1 and increased binding to collagens I and IV, fibronectin, and laminin. Brain sections of mice demonstrated that as T cells migrated farther into the tissue, VLA-4 expression was lost, although CD4 expression remained unchanged. These results demonstrate that binding to VCAM-1 on endothelial cells induces MMP-2 in T cells, which, in turn, may facilitate T cell migration into perivascular tissue. The significance of these findings in the modulation of the inflammatory response is discussed.Key words: T lymphocyte, endothelial cell, matrix metalloproteinase, inflammation, transendothelial migration, integrins, cell adhesion molecules.
Ligand binding and affinity modulation of integrins
Authors: Tozer Eileen Collins , Hughes Paul E. , Loftus Joseph C.
Published: December 2009

Integrins are cell adhesion receptors that mediate cell–cell and cell–extracellular matrix interactions. The extracellular domains of these receptors possess binding sites for a diverse range of protein ligands. Ligand binding is divalent cation dependent and involves well-defined motifs in the ligand. Integrins can dynamically regulate their affinity for ligands (inside-out signaling). This ability to rapidly modulate their affinity state is key to their involvement in such processes as cell migration and platelet aggregation. This review will focus on two aspects of integrin function: first, on the molecular basis of ligand–integrin interactions and, second, on the underlying mechanisms controlling the affinity state of integrins for their ligands.Key words: integrins, ligand binding, affinity modulation.
The integrin α6β4 and the biology of carcinoma
Authors: Rabinovitz Isaac , Mercurio Arthur M.
Published: December 2009

The integrin family of adhesion receptors plays a major role in epithelial organization and function. Moreover, the altered expression and function of specific integrins most likely contributes significantly to carcinoma progression. The integrin α6β4, the focus of this review, is a receptor for several members of the laminin family and is preferentially expressed at the basal surface of most epithelia, where it contributes to basement membrane interactions. Mounting evidence suggests that the α6β4 integrin plays a key role in carcinoma cell biology. Several histopathological studies have established a correlation between α6β4 integrin expression and tumor progression. The importance of α6β4 expression in tumors is underscored by the findings that invading fronts of several carcinomas are enriched in the expression of α6β4 integrin ligands, such as laminin-1 and laminin-5. The participation of the α6β4 integrin in invasion is supported further by in vitro functional studies using carcinoma cells that have been transfected with the β4 cDNA. The mechanisms by which α6β4 contributes to tumor progression are probably related to its mechanical and signaling properties and are currently under intense study.Key words: integrins, laminin receptors, basement membrane, extracellular matrix, metastasis.
Characteristics and densities of Williamson’s Sapsucker nest trees in British Columbia
Authors: Gyug L. W. , Steeger C. , Ohanjanian I.
Published: December 2009

We characterized Williamson’s Sapsucker ( Sphyrapicus thyroideus (Cassin, 1852)) nest trees in southern British Columbia from 1995 to 2008 to determine the nesting requirements of this endangered species in Canada. In the East Kootenay (n = 32) and the Okanagan (n = 157) regions where western larch ( Larix occidentalis Nutt.) occurred, 81% and 75% of the nest trees were western larch, respectively. In regions west of the Okanagan Valley where western larch did not occur (n = 73), 77% of the nest trees were trembling aspen ( Populus tremuloides Michx.). Conifer nest trees were larger (72 cm DBH), on average, than broad-leaved nest trees (35 cm DBH), and this pattern was consistent across the breeding range of Williamson’s Sapsucker. Live western larch >67 cm DBH with either broken or dead tops were the trees with the highest frequency (20%) among the trees estimated to be suitable for nesting, primarily because these larch tended to be infected with heart rots. Based on observed densities, targets should be 1.36 suitable nest trees/ha in managed stands, although in many instances such as aspen patches, this target will only be met in portions of the nesting territories. The recommended stand structure for long-term maintenance of high nest-tree density in western larch stands is for ≥20 trees/ha with DBHs >57 cm and 60–150 trees/ha with DBHs >22 cm.
Temperate forest herbs are adapted to high air humidity — evidence from climate chamber and humidity manipulation experiments in the field
Authors: Lendzion Jasmin , Leuschner Christoph
Published: December 2009

How growth and morphology of wild plants are controlled by the water vapor saturation deficit of the air (vapor pressure deficit, VPD) is not sufficiently understood. We tested the hypothesis that VPD acts on temperate woodland herbs independently from soil moisture by exposing two species ( Mercurialis perennis L. and Stachys sylvatica Torr.) to variable VPD levels in climate chambers and in open-top chambers on the forest floor. A decrease in air humidity from 85% to 40% in the climate chamber experiment, which simulates a microclimate change after canopy gap creation, resulted in a 40% decrease in biomass production of both species when grown in hydroponic culture. This result is supported by the more realistic open-top chamber experiment, which showed a productivity decrease by approximately 25% when plants were continuously exposed to an atmosphere with 15% lower air humidity than ambient. Elevated VPD levels reduced biomass production through either a reduced leaf expansion rate or a lower number of formed leaf buds. We conclude that many woodland herbs require sufficiently high air humidity for optimal growth; permanently decreased air humidity, as may occur in a future drier climate, after gap creation, or after clear-cutting of the forest, may threaten the vitality and survival of woodland herbs.
Patterns of late-season photosynthate movement in sugar maple saplings
Authors: Horowitz Michael E. , Fahey Timothy J. , Yavitt Joseph B. , Feldpausch Ted R. , Sherman Ruth E.
Published: December 2009

A more detailed understanding of seasonal dynamics of carbon allocation between roots and shoots of temperate zone trees is needed. We labeled sapling stands of sugar maple ( Acer saccharum Marsh) with 13CO2 in four 3 m diameter chambers during a 3 week interval in September 2006 and traced the movement of this 13C pulse through trees and soil during autumn and spring. High enrichment of 13C in foliage was achieved (δ13C = 387‰ ± 16‰). Late-season photosynthate was strongly mobilized during leaf senescence and stored in twigs, wood, and roots. Very high 13C enrichment of soil CO2 emissions at this time (δ13C = 766‰ ± 82‰) indicated the role of late-season photosynthesis in supplying root metabolism after the growing season. Rhizosphere soil was weakly enriched in 13C during fall, and increased significantly over the winter. Earthworms were highly enriched in fall, indicating that they consumed roots. In spring, 13C was strongly mobilized to growing shoots but not to growing fine roots; apparently, fine root growth in spring was not supplied by late-season stored photosynthate. These results provide insights into the seasonal dynamics of temperate tree carbon allocation with implications for disruption of these dynamics by global warming.
Diversity and fruiting patterns of ectomycorrhizal and saprobic fungi as indicators of land-use severity in managed woodlands dominated by Quercus suber — a case study from southern Portugal
Authors: Azul Anabela Marisa , Castro Paula , Sousa José Paulo , Freitas Helena
Published: December 2009

We assessed the impacts of current management practices used to control shrub strata in Portuguese oak woodlands dominated by Quercus suber L. (montado) on fruiting diversity and abundance of ectomycorrhizal-forming fungi (ECMF) and saprobic fungi. Fruit bodies were collected over four fruiting seasons in 16 plots (20 m × 20 m) selected in a montado landscape with extensive silvopastoral exploitation. A total of 9484 fruit bodies were found in 171 taxa (74 ECMF, 96 saprobic, and 1 parasitic). Our results show that shrub density control by permanent grazing or by cutting practices followed by soil tillage leads to lower fruiting production and greater changes in taxa composition, particularly for ECMF fruit bodies, than cutting practices without soil tillage. Principal response curve analysis showed that ECMF reacted more sensitively to these practices, in particular Laccaria laccata, Hebeloma cistophilum, Russula cyanoxantha, Cortinarius trivialis, and Lactarius volemus. We also observed that shrub cutting without soil tillage allowed ECMF fruiting to recover to predisturbance levels after 3 years. Our data imply that fruit bodies were useful indicators for assessing the severity of the effects of different land-use practices applied in montado areas on soil fungal populations.
Evaluating relationships among tree growth rate, shade tolerance, and browse tolerance following disturbance in an eastern deciduous forest
Authors: Krueger Lisa M. , Peterson Chris J. , Royo Alejandro , Carson Walter P.
Published: December 2009

Interspecific differences in shade tolerance among woody species are considered a primary driving force underlying forest succession. However, variation in shade tolerance may be only one of many interspecific differences that cause species turnover. For example, tree species may differ in their sensitivity to herbivory. Nonetheless, existing conceptual models of forest dynamics rarely explicitly consider the impact of herbivores. We examined whether browsing by white-tailed deer ( Odocoileus virginianus Zimmermann) alters the relationship between light availability and plant performance. We monitored growth and survival for seedlings of six woody species over 2 years within six windthrow gaps and the nearby intact forest in the presence and absence of deer. Browsing decreased seedling growth for all species except beech ( Fagus grandifolia Ehrh.). More importantly, browsing altered growth rankings among species. Increased light availability enhanced growth for three species when excluded from deer, but browsing obscured these relationships. Browsing also reduced survival for three species; however, survival rankings did not significantly differ between herbivory treatments. Our results indicated that browsing and light availability operated simultaneously to influence plant growth within these forests. Thus, existing models of forest dynamics may make inaccurate predictions of the timing and composition of species reaching the canopy, unless they can account for how plant performance varies as a result of a variety of environmental factors, including herbivory.
Present-day expansion of American beech in northeastern hardwood forests: Does soil base status matter?
Authors: Duchesne Louis , Ouimet Rock
Published: December 2009

Recently, sugar maple ( Acer saccharum Marsh.) decline in northeastern North America has been regarded as a major factor structuring hardwood forests by favouring American beech ( Fagus grandifolia Ehrh.) in the understory of maple-dominated stands. To determine whether soil fertility differences associated with sugar maple decline may have promoted the expansion of American beech, we explored the relationships between the soil base status and the sapling and tree strata density and composition, using data from 426 permanent sample plots distributed throughout Quebec. Our results indicate that American beech is currently expanding in the sugar maple range of Quebec. The abundance and proportion of American beech in the sapling stratum are mainly associated with the proportion of American beech in the tree stratum, the relative basal area of dead sugar maple trees, and the base status of soils. In accordance with the many studies reporting on the high sensitivity of sugar maple to the acid–base status of soils and the decline of the sugar maple population, this study supports the hypothesis that soil base cation depletion, caused in part by atmospheric acid deposition, is among the main factors involved in the present-day expansion of American beech over a large area in Quebec.
CRYSTALLIZATION BEHAVIOR OF MAGNESIUM SALTS: A SUMMARY OF SOME EXPERIMENTAL OBSERVATIONS
Authors: XUE DONGFENG , ZOU LONGJIANG , WANG LEI , YAN XIAOXING
Published: December 2009

Some efficient methods have been developed to chemically prepare some magnesium salt products with multiscale morphologies such as MgCO 3·3 H 2 O and 5 Mg ( OH )2· MgSO 4·2 H 2 O whiskers, Mg 5( CO 3)4( OH )2·4 H 2 O microplates and aggregates, and Mg ( OH )2 plates and spheres. The crystallization processes of the as-prepared products were studied by designing different reaction routes and corresponding growth morphologies. In these solution chemical systems, Mg 2+ cation was found to be easily coordinated by various ligands, and can be easily transformed into various compounds with variable compositions. The current crystallization observations of some magnesium salts can be readily simulated by the chemical bonding theory of single crystal growth. These results can guide the further studies of various magnesium salts with controllable morphologies.
GROWTH OF ONE-DIMENSIONAL MnO2 NANOSTRUCTURE
Authors: LU PAI , XUE DONGFENG
Published: December 2009

Large scale MnO 2 nanorods were controllably synthesized from the inexpensive precursors (e.g., manganese acetate, ammonium persulfate) via a facile one-step low temperature hydrothermal strategy. The crystal phase and microscopic morphology of the as-prepared MnO 2 nanorods were characterized by X-ray powder diffraction (XRD) and scanning electron microscope (SEM). Through investigating the morphology evolution of MnO 2 products in the whole synthesis process, a novel growth mechanism of these MnO 2 nanorods was proposed, which may be efficiently extended to other material systems as a general approach towards one-dimensional nanostructures. The obtained MnO 2 nanorods may have potential applications in Li -ion batteries and supercapacitors.