43,864 research outputs found
A Multi-Language Comparison of Influences on Author Verification using Character N-Grams
No full textWe create a new multi-language corpus for author verification based on Wikipedia talkpages, and evaluate the influence that differences in topic and time have on character n-gram author profiles. Topic alignment between two texts is found to increase author verification precision, and an authors writing style is found to change over time, but not more significantly after 3 years than after 1 year.Information ArchitectureWISElectrical Engineering, Mathematics and Computer Scienc
Receptor Guanylyl Cyclase C Cross-talk With Tyrosine Kinases And The Adaptor Protein, Crk
No full textSignal transduction is a crucial event that enables cells to sense and respond to cues from their immediate environment. Guanylyl cyclase C (GC-C) is a member of the family of receptor guanylyl cyclases. GC-C is a single transmembrane protein that responds to its ligands by the production of the second messenger cGMP. The guanylin family of peptides, (including the bacterially produced heat-stable enterotoxin ST) is the ligand for GC-C, elevates intracellular cGMP levels and activates downstream pathways. GC-C regulates the cystic fibrosis transmembrane conductance regulator (CFTR) by inducing phosphorylation by protein kinase G, resulting in chloride ion and fluid efflux. GC-C also regulates cell cycle progression through cGMP-gated Ca2+ channels. These functions are seen in the intestinal epithelium, the primary site for GC-C expression.
GC-C as a molecule has been studied in detail, but its functioning in the context of other signaling pathways remains unknown. The aim of the present investigation was to understand the regulation of signal transduction by GC-C and its cross-talk with other signaling pathways operating in the cell. Molecular events that commonly connect components in a signaling pathway are protein phosphorylation and protein-protein interaction. These two aspects are explored in this thesis.
The possibility of tyrosine phosphorylation of GC-C has been explored earlier in our laboratory. In vitro studies indicated that the residue Tyr820 was a site for phosphorylation by the Src family of non-receptor tyrosine kinases and those studies also suggested that phosphorylated Tyr820 could bind to the SH2 domain of Src. We generated a nonphosphorylatable mutant of GC-C, GC-CY820F, and a phosphomimetic mutant GC-CY820E to study the effect of phosphorylation of Tyr820, on the functioning of GC-C. A stable cell line of HEK293:GC-CY820F cells was generated and compared with HEK293:GC-CWT. Dose response to ST in the two cell lines showed that cGMP accumulation by GC-CY820F was greater than that of GC-CWT, although the EC50 remained unchanged. The phosphomimetic GC-CY820E mutant receptor was non-responsive to ST. Further in HEK293 cells, phosphorylation of GC-CWT by constitutively active v-Src resulted in decreased ST stimulation and this effect of v-Src was reduced with GC-CY820F. Inhibition of ST stimulation brought about by v-Src required catalytically active Src, as the kinase inactive v-SrcK295R did not inhibit ST stimulation. These results were corroborated by in vitro studies by using the recombinant catalytic domain of GC-C expressed in insect cells and by phosphorylation using a purified kinase, Hck. Observations suggested that phosphorylation of Tyr820 in the catalytic domain of GC-C compromises the guanylyl cyclase activity of GC-C.
T84 and Caco-2 colon carcinoma cells endogenously express GC-C. The effect of tyrosine phosphorylation of GC-C was studied by using HgCl2, a known activator of Src kinases, and by the inhibition of protein tyrosine phosphatases using pervanadate, an irreversible inhibitor. Both these ways of achieving increased tyrosine phosphorylation resulted in decreased ST-stimulated cGMP production by GC-C, as suggested from v-Src transfection studies. This decrease was reversed by using a Src kinase specific inhibitor PP2, confirming the role of Src kinases in the inhibition of GC-C activity. Interestingly, in Caco-2 cells that differentiate in culture, the effect of pervanadate on the inhibition of ST-stimulated GC-C activation was dependent on the differentiation stage. Crypt-like cells showed higher inhibition with pervanadate. As they matured into villus-like cells, the effect of pervanadate on GC-C activation was gradually lost. This effect also correlated with a decrease in the expression of Lck, suggesting that in the context of the intestine there could be differential regulation of tyrosine phosphorylation of GC-C along the crypt-villus axis. Intestinal ligated loop assays in rats demonstrated that ST-induced fluid accumulation in the intestine was abrogated on pervanadate treatment. Reduction in this fluid accumulation by pervanadate was not observed with 8-Br-cGMP, a cell permeable analogue of cGMP. This indicated that tyrosine phosphorylation of proteins is important for ST-induced fluid accumulation, and perhaps pervanadate modulates this by phosphorylation of GC-C, thereby causing a reduction in fluid accumulation.
Earlier in vitro studies on Src-SH2 binding from the laboratory had suggested the possibility of activation of Src family kinases by GC-C. The activation status of Src kinases was monitored by using phosphorylation-state specific antibody, pSFK416. ST stimulation in T84 cells increased Tyr416 phosphorylation of Src kinases in a time dependent manner, indicating that Src kinases are activated downstream of GC-C. This activation of Src kinases was also seen with the endogenous ligand of GC-C, uroguanylin. Interestingly, 8-Br-cGMP a cell permeable analogue of cGMP that is known to mimic other cellular effects of GC-C, namely Cl-secretion and cell cycle progression, did not activate Src kinases, suggesting that the mechanism of Src kinase activation by GC-C could be independent of cGMP.
Binding affinities of Src, Lck, Fyn and Yes SH2 domains to Tyr820 phosphorylated GCC peptide were in the nM range, indicating a high affinity of interaction. In vitro GST-SH2 pull down experiments suggested that phosphorylation of Tyr820 in full length GC-C allows interaction of GC-C to the SH2 domain of Src. These studies suggest a dual cross-talk between Src kinases and GC-C; Src phosphorylation inhibits GC-C signaling and stimulation of GC-C by its ligands activates Src kinases.
Interaction of proteins containing SH2 and SH3 domains are commonly found in signaling molecules. In accordance with the observation that there are three PXXP motifs in GCC, many SH3 domains could interact with GC-C. GC-C appears to show a preference to bind the SH3 domains of Fyn, Hck, Abl tyrosine kinases, Grb2 and Crk adaptor proteins, the α-subunit of P85 PI3 kinase, PLC-γ and cortactin to various extents. The SH3 domains of spectrin and Nck did not show any detectable interaction with GC-C. In SH3 pull-down assays, the N-terminal SH3 domain of Crk, CrkSH3 (N), bound GC-C maximally, suggesting that Crk is a good candidate for interaction with GC-C.
By overlay analysis, the region of GC-C that binds CrkSH3 (N) was narrowed down to the catalytic domain of GC-C containing a ‘PGLP’ motif. Mutations were generated in GC-C at this site to generate GC-CP916Q and GC-CW918R. These mutations compromised the binding of full length receptor to CrkSH3 (N). In cells, CrkII and GC-C co-transfection inhibited the ST stimulation of GC-C. A CrkII mutant, that has compromised binding through its SH3 domain, did not inhibit the activity of GC-C. CrkII from T84 cells co-immunoprecipitated with GC-C and interestingly, the phosphorylated form of CrkII did not, indicating that GC-C - Crk interaction could be regulated by the phosphorylation of Crk.
In summary, this study places GC-C, in the context of tyrosine kinase signaling pathway and interaction with the adaptor protein Crk. These studies suggest that GC-C signal transduction can be altered by cross-talk with other signaling events in the cell. Reversible phosphorylation of tyrosine residues inhibits the activity of GC-C, and this is mediated by Src family kinases. Src kinases themselves are activated on stimulation of GC-C by its ligands, possibly because of SH2 domain interaction with GC-C. Association of Crk by its SH3 domain regulates GC-C functioning primarily by inhibiting ST-stimulated cGMP production. This opens up the possibility of GC-C signaling through a multimeric complex involving other binding partners of Crk, and these cross-talks involving GC-C with the two proto-oncogenes, Src and Crk, might have far reaching consequences in the regulation of cellular functions
Fast implementation of iterative adaptive approach for wideband unambiguous radar detection
No full textAccepted author manuscriptMicrowave Sensing, Signals & System
Radical Propagation Kinetics of N-Vinylpyrrolidone in Organic Solvents Studied by Pulsed-Laser Polymerization-Size-Exclusion Chromatography (PLP-SEC)
No full textPulsed-laser polymerization with subsequent analysis of the polymer molar mass distribution by size-exclusion chromatography, PLP-SEC, is used to measure the propagation rate coefficient, k(p), of N-vinylpyrrolidone (NVP) in a series of organic solvents, varying the NVP concentration from 5 to 100 wt% and varying the temperature between-5 and +80 degrees C. In contrast to the 20-fold increase observed in aqueous solution upon decreasing the NVP concentration from bulk to dilute conditions, the k(p) values of NVP in butyl acetate, iso-propyl acetate, N-ethylpyrrolidone, and N-ethylformamide stay within 20% of the bulk value and exhibit no significant dependence on monomer concentration. The k(p) behavior of NVP in methanol and n-butanol is intermediate between the one in water and in the other organic solvents, with k(p) increasing by about a factor of 2 upon lowering the monomer concentration from bulk to 5 wt% NVP. The activation energies for propagation in organic solvents agree within experimental uncertainty with the value reported for bulk NVP. The data demonstrate that hydrogen bonding is responsible for the increase in k(p) upon dilution, with this effect being much stronger in an aqueous environment than in a solution of alcohol
The importance of chain conformational mobility during 5-exo-cyclizations of C-, N- and O-centred radicals
No full textThe author thanks the EPSRC (grant EP/I003479/1) and EaStCHEM for funding.The reaction coordinates of an archetypical set of 5-exo cyclizations of C-, N- and O-centred radicals were investigated by computational methods. G4 theory, and DFT with the um062x functional, were able to rationalise counterintuitive factors such as the 'normal' order of rate constants being: N-centredPeer reviewe
Dissipative Range Scaling of Higher Order Structure Functions for Velocity and Passive Scalars
No full textDifferently to Kolmogorov's second similarity hypothesis, we find that the 2n-th order velocity and scalar structure functions scale with n-th order moment of the energy dissipation and the scalar dissipation, respectively. The origins of this scaling are analyzed by the transport equations of the fourth order velocity and scalar increment moments and by direct numerical simulations
The vanishing author in computer-generated works: a critical analysis of recent Australian case law
No full textAbstract
The use of software is ubiquitous in the creation of many copyright works, yet the requirement in copyright law that every work have a human author who engages in independent intellectual effort means that its use may prevent copyright subsistence. Several recent Australian cases have refocused attention on authorship as an essential criterion of copyright subsistence, and these cases suggest that much computer-produced output may be authorless and thus lack copyright protection. This article, the first in a two-part series, analyses how each case deals with the question of authorship of computer-produced works and why the use of software diminishes copyright protection for a significant number of computer-generated works. The article critiques the application of conventional notions of human authorship developed in the pre-computer age to modern productions and suggests alternative approaches to authorship that satisfy both the major objectives of copyright policy and the need to adapt to the computer age. The article argues that, without a broader judicial approach to authorship of computer-generated works, Parliament must remedy the lacuna in protection for these ‘authorless’ works. Possible solutions for reform are suggested. In a forthcoming article, the author comprehensively examines those reform proposals
Ratio of n-6/n-3 in the diets of beef cattle
No full textEffects of feeding heat-treated canola (C), soybean (S) and flax (F) or mixtures on growth and slaughter characteristics, taste and fatty acid (FA) composition of beef tissue were investigated using 128 crossbred steers to determine the potential of improving the nutritional quality of beef for humans. For Trial 1 (48 steers), dietary treatments were: roasted C, extruded C, roasted S, extruded S, roasted F and extruded F. For Trial 2 (80 steers), the dietary treatments were: S:F (1:1), S:C (1:1), C:F (1:1) and S:F:C (1:1:1), and the oilseeds were processed either by roasting or extruding before mixing. Soybean meal and soybean oil were used to give equivalent lipid and protein contents to each experimental diet. The basal diet consisted of grass silage, barley grain, vitamins and minerals. Steers were fed for a minimum of 100d then slaughtered at a uniform degree of finish. Growth and slaughter characteristics of the steers were only slightly affected by dietary treatment in that the soybean-fed steers consumed more feed and had a higher average daily gain than the canola or flax-fed animals in Trial 1. There was no difference in taste panel parameters for any of the treatments. Inclusion of flax in the diet increased the total n-3 content of meat. Similar results were found for canola and C18:1n-9 although this was not the case for soybean and the n-6 FA. For the n-6 FA in the PL and neutral lipid fractions of the meat samples, levels were correlated with high dietary levels of n-6 or n-9 with low levels of n-3 while for the n-3 FA, levels were correlated with high dietary n-3 levels and low n-6 levels. Oilseed processing method did not have an effect on any fatty acid levels. It is possible to modify the FA composition of beef meat toward a healthier profile by including heat-treated oilseeds in the diet to influence the degree of lipid metabolism in the rumen.ID: S0377840111004007; M3: Article; Accession Number: S0377840111004007; Author: M.A. McNiven (a, ⁎); Author: J.L. Duynisveld (b); Author: T. Turner (a); Author: A.W. Mitchell (a); Affiliation: Department of Health Management, Atlantic Veterinary College, University of PEI, Charlottetown, PEI, Canada C1A 4P3; Affiliation: Agriculture and Agri-Food Canada, Nappan, NS, Canada B0L 1C0; Keyword: Oilseeds; Keyword: Roasted; Keyword: Extruded; Keyword: Fatty acids; Keyword: Healthy fat; Number of Pages: 11; Language: English
Measurement of the ratio of prompt χ c to J / ψ production in pp collisions at √s = 7 TeV
No full textThe prompt production of charmonium χ c and J / ψ states is studied in proton-proton collisions at a centre-of-mass energy of √s = 7 TeV at the Large Hadron Collider. The χ c and J / ψ mesons are identified through their decays χ c → J / ψ γ and J / ψ → μ + μ - using 36 pb - 1 of data collected by the LHCb detector in 2010. The ratio of the prompt production cross-sections for χ c and J / ψ, σ (χ c → J / ψ γ) / σ (J / ψ), is determined as a function of the J / ψ transverse momentum in the range 2 < p T J / ψ < 15 GeV / c. The results are in excellent agreement with next-to-leading order non-relativistic expectations and show a significant discrepancy compared with the colour singlet model prediction at leading order, especially in the low p T J / ψ region
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