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61.
Giamas G Hirner H Shoshiashvili L Grothey A Gessert S Kühl M Henne-Bruns D Vorgias CE Knippschild U 《The Biochemical journal》2007,406(3):389-398
The involvement of CK1 (casein kinase 1) delta in the regulation of multiple cellular processes implies a tight regulation of its activity on many different levels. At the protein level, reversible phosphorylation plays an important role in modulating the activity of CK1delta. In the present study, we show that PKA (cAMP-dependent protein kinase), Akt (protein kinase B), CLK2 (CDC-like kinase 2) and PKC (protein kinase C) alpha all phosphorylate CK1delta. PKA was identified as the major cellular CK1deltaCK (CK1delta C-terminal-targeted protein kinase) for the phosphorylation of CK1delta in vitro and in vivo. This was implied by the following evidence: PKA was detectable in the CK1deltaCK peak fraction of fractionated MiaPaCa-2 cell extracts, PKA shared nearly identical kinetic properties with those of CK1deltaCK, and both PKA and CK1deltaCK phosphorylated CK1delta at Ser370 in vitro. Furthermore, phosphorylation of CK1delta by PKA decreased substrate phosphorylation of CK1delta in vitro. Mutation of Ser370 to alanine increased the phosphorylation affinity of CK1delta for beta-casein and the GST (gluthatione S-transferase)-p53 1-64 fusion protein in vitro and enhanced the formation of an ectopic dorsal axis during Xenopus laevis development. Anchoring of PKA and CK1delta to centrosomes was mediated by AKAP (A-kinase-anchoring protein) 450. Interestingly, pre-incubation of MiaPaCa-2 cells with the synthetic peptide St-Ht31, which prevents binding between AKAP450 and the regulatory subunit RII of PKA, resulted in a 6-fold increase in the activity of CK1delta. In summary, we conclude that PKA phosphorylates CK1delta, predominantly at Ser370 in vitro and in vivo, and that site-specific phosphorylation of CK1delta by PKA plays an important role in modulating CK1delta-dependent processes. 相似文献
62.
Although the extensive use of Aspergillus-active antifungals has been recently associated with an increase in zygomycosis in several cancer centers, the frequency of this opportunistic mycosis began to rise earlier, since the mid 1990s. The reasons for that emergence are unclear. Recent evidence suggests that endosymbiotic bacteria of Rhizopus species produce toxins that enhance fungal pathogenicity. We postulate that, although Zygomycetes appear equally ubiquitous and virulent to Aspergillus, zygomycosis was rare in the past in immunosuppressed patients specifically because of the widespread use of antibacterials in this patient population. Such use may have resulted in inhibition of endosymbiotic, toxin-producing bacteria and led indirectly in attenuation of Zygomycetes virulence. Thus, the growing rates of antimicrobial resistance over the past decade selected for multidrug-resistant endosymbiotic bacteria of Zygomycetes, which could facilitate the emergence of zygomycosis. This hypothesis, if true, will be the first paradigm of modulation of virulence of opportunistic fungi by antibacterials. 相似文献
63.
Lepori MB Lovicu M Dessi V Zappu A Incollu S Zancan L Giacchino R Iorio R Vajro P Maggiore G Marcellini M Barbera C Pellecchia MT Simonetti R Kostic V Farci AM Solinas A De Virgiliis S Cao A Loudianos G 《Genetic testing》2007,11(3):328-332
Herein we report the results of mutation analysis of the ATP7B gene in a group of 134 Wilson disease (WD) families (268 chromosomes) prevalently of Italian origin. Using the SSCP and sequencing methods we identified 71 disease-causing mutations. Twenty-four were novel, while 19 more mutations already described, were identified in new populations in this study. A known mutation G591D showed a regional distribution, since it was only detected in 38.5% of the analyzed chromosomes in WD patients originating from Apulia, a region of South Italy. Detection of new mutations in the ATP7B gene increases our capability of molecular analysis that is essential for early diagnosis and treatment of WD. 相似文献
64.
Dimou M Venieraki A Liakopoulos G Kouri ED Tampakaki A Katinakis P 《Journal of molecular microbiology and biotechnology》2011,20(3):176-190
The soil nitrogen-fixing bacterium Azotobacter vinelandii possesses two cyclophilins, comprising putative cytoplasmic and periplasmic isoforms, designated as AvPPIB and AvPPIA, respectively. Both recombinant cyclophilins have been purified and their peptidyl-prolyl cis/trans isomerase activity against Suc-Ala-Xaa-Pro-Phe-pNA synthetic peptides has been characterized. The substrate specificity of both cyclophilins is typical for bacterial cyclophilins, with Suc-Ala-Ala-Pro-Phe-pNA being the most rapidly catalyzed substrate. The cytoplasmic cyclophilin also displays a chaperone function in the citrate synthase thermal aggregation assay. Using real-time quantitative RT-PCR, we demonstrate that AvppiB is expressed under various physiological and growth conditions, mainly upregulated by acetate and downregulated by the stationary growth state, while AvppiA shows a tendency for downregulation under the tested conditions. Further, we identified chaperone protein dnaK and UDP-2, 3-diacylglucosamine hydrolase lpxH as probable interacting partners of AvPPIB and we demonstrate their physical interaction by coexpression studies. An increase in AvPPIB PPIase activity in the presence of AvdnaK and a decrease in the presence of AvlpxH further confirms each interaction. However, the PPIase activity does not seem to be essential for those interactions since AvPPIB active site mutants still interact with dnaK and lpxH, while their minor PPIase activity cannot be modulated by the interaction. 相似文献
65.
Fookes M Schroeder GN Langridge GC Blondel CJ Mammina C Connor TR Seth-Smith H Vernikos GS Robinson KS Sanders M Petty NK Kingsley RA Bäumler AJ Nuccio SP Contreras I Santiviago CA Maskell D Barrow P Humphrey T Nastasi A Roberts M Frankel G Parkhill J Dougan G Thomson NR 《PLoS pathogens》2011,7(8):e1002191
The genus Salmonella contains two species, S. bongori and S. enterica. Compared to the well-studied S. enterica there is a marked lack of information regarding the genetic makeup and diversity of S. bongori. S. bongori has been found predominantly associated with cold-blooded animals, but it can infect humans. To define the phylogeny of this species, and compare it to S. enterica, we have sequenced 28 isolates representing most of the known diversity of S. bongori. This cross-species analysis allowed us to confidently differentiate ancestral functions from those acquired following speciation, which include both metabolic and virulence-associated capacities. We show that, although S. bongori inherited a basic set of Salmonella common virulence functions, it has subsequently elaborated on this in a different direction to S. enterica. It is an established feature of S. enterica evolution that the acquisition of the type III secretion systems (T3SS-1 and T3SS-2) has been followed by the sequential acquisition of genes encoding secreted targets, termed effectors proteins. We show that this is also true of S. bongori, which has acquired an array of novel effector proteins (sboA-L). All but two of these effectors have no significant S. enterica homologues and instead are highly similar to those found in enteropathogenic Escherichia coli (EPEC). Remarkably, SboH is found to be a chimeric effector protein, encoded by a fusion of the T3SS-1 effector gene sopA and a gene highly similar to the EPEC effector nleH from enteropathogenic E. coli. We demonstrate that representatives of these new effectors are translocated and that SboH, similarly to NleH, blocks intrinsic apoptotic pathways while being targeted to the mitochondria by the SopA part of the fusion. This work suggests that S. bongori has inherited the ancestral Salmonella virulence gene set, but has adapted by incorporating virulence determinants that resemble those employed by EPEC. 相似文献
66.
Spagou K Theodoridis G Wilson I Raikos N Greaves P Edwards R Nolan B Klapa MI 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2011,879(17-18):1467-1475
Metabolic profiling of biofluids, based on the quantitative analysis of the concentration profile of their free low molecular mass metabolites, has been playing increasing role employed as a means to gain understanding of the progression of metabolic disorders, including obesity. Chromatographic methods coupled with mass spectrometry have been established as a strategy for metabolic profiling. Among these, GC-MS, targeting mainly the primary metabolism intermediates, offers high sensitivity, good peak resolution and extensive databases. However, the derivatization step required for many involatile metabolites necessitates specific data validation, normalization and analysis protocols to ensure accurate and reproducible performance. In this study, the GC-MS metabolic profiles of plasma samples from mice maintained on 12- or 15-month long low (10 kcal%) or high (60 kcal%) fat diets were obtained. The profiles of the trimethylsilyl(TMS)-methoxime(MeOx) derivatives of the free polar metabolites were acquired through GC-(ion trap)MS, using [U-(13)C]-glucose as the internal standard. After the application of a recently developed data correction and normalization/filtering protocol for GC-MS metabolomic datasets, the profiles of 48 out of the 77 detected metabolites were used in multivariate statistical analysis. Data mining suggested a decrease in the activity of the energy metabolism with age. In addition, the metabolic profiles indicated the presence of subpopulations with different physiology within the high- and low-fat diet mice, which correlated well with the difference in body weight among the animals and current knowledge about hyperglycemic conditions. 相似文献
67.
Marcella A. Calfon Amir Rosenthal Georgios Mallas Adam Mauskapf R. Nika Nudelman Vasilis Ntziachristos Farouc A. Jaffer 《Journal of visualized experiments : JoVE》2011,(54)
The vascular response to injury is a well-orchestrated inflammatory response triggered by the accumulation of macrophages within the vessel wall leading to an accumulation of lipid-laden intra-luminal plaque, smooth muscle cell proliferation and progressive narrowing of the vessel lumen. The formation of such vulnerable plaques prone to rupture underlies the majority of cases of acute myocardial infarction. The complex molecular and cellular inflammatory cascade is orchestrated by the recruitment of T lymphocytes and macrophages and their paracrine effects on endothelial and smooth muscle cells.1Molecular imaging in atherosclerosis has evolved into an important clinical and research tool that allows in vivo visualization of inflammation and other biological processes. Several recent examples demonstrate the ability to detect high-risk plaques in patients, and assess the effects of pharmacotherapeutics in atherosclerosis.4 While a number of molecular imaging approaches (in particular MRI and PET) can image biological aspects of large vessels such as the carotid arteries, scant options exist for imaging of coronary arteries.2 The advent of high-resolution optical imaging strategies, in particular near-infrared fluorescence (NIRF), coupled with activatable fluorescent probes, have enhanced sensitivity and led to the development of new intravascular strategies to improve biological imaging of human coronary atherosclerosis.Near infrared fluorescence (NIRF) molecular imaging utilizes excitation light with a defined band width (650-900 nm) as a source of photons that, when delivered to an optical contrast agent or fluorescent probe, emits fluorescence in the NIR window that can be detected using an appropriate emission filter and a high sensitivity charge-coupled camera. As opposed to visible light, NIR light penetrates deeply into tissue, is markedly less attenuated by endogenous photon absorbers such as hemoglobin, lipid and water, and enables high target-to-background ratios due to reduced autofluorescence in the NIR window. Imaging within the NIR ''window'' can substantially improve the potential for in vivo imaging.2,5Inflammatory cysteine proteases have been well studied using activatable NIRF probes10, and play important roles in atherogenesis. Via degradation of the extracellular matrix, cysteine proteases contribute importantly to the progression and complications of atherosclerosis8. In particular, the cysteine protease, cathepsin B, is highly expressed and colocalizes with macrophages in experimental murine, rabbit, and human atheromata.3,6,7 In addition, cathepsin B activity in plaques can be sensed in vivo utilizing a previously described 1-D intravascular near-infrared fluorescence technology6, in conjunction with an injectable nanosensor agent that consists of a poly-lysine polymer backbone derivatized with multiple NIR fluorochromes (VM110/Prosense750, ex/em 750/780nm, VisEn Medical, Woburn, MA) that results in strong intramolecular quenching at baseline.10 Following targeted enzymatic cleavage by cysteine proteases such as cathepsin B (known to colocalize with plaque macrophages), the fluorochromes separate, resulting in substantial amplification of the NIRF signal. Intravascular detection of NIR fluorescence signal by the utilized novel 2D intravascular NIRF catheter now enables high-resolution, geometrically accurate in vivo detection of cathepsin B activity in inflamed plaque.
In vivo molecular imaging of atherosclerosis using catheter-based 2D NIRF imaging, as opposed to a prior 1-D spectroscopic approach,6 is a novel and promising tool that utilizes augmented protease activity in macrophage-rich plaque to detect vascular inflammation.11,12 The following research protocol describes the use of an intravascular 2-dimensional NIRF catheter to image and characterize plaque structure utilizing key aspects of plaque biology. It is a translatable platform that when integrated with existing clinical imaging technologies including angiography and intravascular ultrasound (IVUS), offers a unique and novel integrated multimodal molecular imaging technique that distinguishes inflammatory atheromata, and allows detection of intravascular NIRF signals in human-sized coronary arteries.Download video file.(61M, mov) 相似文献
68.
Athanasiadis G Buil A Souto JC Borrell M López S Martinez-Perez A Lathrop M Fontcuberta J Almasy L Soria JM 《PloS one》2011,6(12):e29168
The Protein C anticoagulant pathway regulates blood coagulation by preventing the inadequate formation of thrombi. It has two main plasma components: protein C and protein S. Individuals with protein C or protein S deficiency present a dramatically increased incidence of thromboembolic disorders. Here, we present the results of a genome-wide association study (GWAS) for protein C and protein S plasma levels in a set of extended pedigrees from the Genetic Analysis of Idiopathic Thrombophilia (GAIT) Project. A total number of 397 individuals from 21 families were typed for 307,984 SNPs using the Infinium® 317 k Beadchip (Illumina). Protein C and protein S (free, functional and total) plasma levels were determined with biochemical assays for all participants. Association with phenotypes was investigated through variance component analysis. After correcting for multiple testing, two SNPs for protein C plasma levels (rs867186 and rs8119351) and another two for free protein S plasma levels (rs1413885 and rs1570868) remained significant on a genome-wide level, located in and around the PROCR and the DNAJC6 genomic regions respectively. No SNPs were significantly associated with functional or total protein S plasma levels, although rs1413885 from DNAJC6 showed suggestive association with the functional protein S phenotype, possibly indicating that this locus plays an important role in protein S metabolism. Our results provide evidence that PROCR and DNAJC6 might play a role in protein C and free protein S plasma levels in the population studied, warranting further investigation on the role of these loci in the etiology of venous thromboembolism and other thrombotic diseases. 相似文献
69.
Hoehndorf R Dumontier M Oellrich A Rebholz-Schuhmann D Schofield PN Gkoutos GV 《PloS one》2011,6(7):e22006
Researchers design ontologies as a means to accurately annotate and integrate experimental data across heterogeneous and disparate data- and knowledge bases. Formal ontologies make the semantics of terms and relations explicit such that automated reasoning can be used to verify the consistency of knowledge. However, many biomedical ontologies do not sufficiently formalize the semantics of their relations and are therefore limited with respect to automated reasoning for large scale data integration and knowledge discovery. We describe a method to improve automated reasoning over biomedical ontologies and identify several thousand contradictory class definitions. Our approach aligns terms in biomedical ontologies with foundational classes in a top-level ontology and formalizes composite relations as class expressions. We describe the semi-automated repair of contradictions and demonstrate expressive queries over interoperable ontologies. Our work forms an important cornerstone for data integration, automatic inference and knowledge discovery based on formal representations of knowledge. Our results and analysis software are available at http://bioonto.de/pmwiki.php/Main/ReasonableOntologies. 相似文献
70.
Stax MJ Naarding MA Tanck MW Lindquist S Hernell O Lyle R Brandtzaeg P Eggesbø M Pollakis G Paxton WA 《PloS one》2011,6(2):e17316