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  • DKFZ Publication Database  (1,990)
  • PROTEIN  (1,990)
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  • Articles  (2)
  • DKFZ Publication Database  (1,990)
Keywords
  • 1
    Keywords: PROTEIN ; MICE ; ACTIVATION ; PROGRESSION ; inactivation ; NEPHROPATHY ; 2 PARTS ; MANNOSE-BINDING LECTIN ; VASCULAR COMPLICATIONS ; CD59
    Abstract: Coagulation and complement regulators belong to two interactive systems constituting emerging mechanisms of diabetic nephropathy. Thrombomodulin (TM) regulates both coagulation and complement activation, in part through discrete domains. TM's lectin like domain dampens complement activation, while its EGF-like domains independently enhance activation of the anti-coagulant and cytoprotective serine protease protein C (PC). A protective effect of activated PC in diabetic nephropathy is established. We hypothesised that TM controls diabetic nephropathy independent of PC through its lectin-like domain by regulating complement. Diabetic nephropathy was analysed in mice lacking TM's lectin-like domain (TMLeD/LeD) and controls (TMwt/wt). Albuminuria (290 mug/mg vs. 166 mug/mg, p=0.03) and other indices of experimental diabetic nephropathy were aggravated in diabetic TMLeD/LeD mice. Complement deposition (C3 and C5b-9) was markedly increased in glomeruli of diabetic TMLeD/LeD mice. Complement inhibition with enoxaparin ameliorated diabetic nephropathy in TMLeD/LeD mice (e.g. albuminuria 85 mug/mg vs. 290 mug/mg, p〈0.001). In vitro TM's lectin-like domain cell-autonomously prevented glucose-induced complement activation on endothelial cells and - notably - on podocytes. Podocyte injury, which was enhanced in diabetic TMLeD/LeD mice, was reduced following complement inhibition with enoxaparin. The current study identifies a novel mechanism regulating complement activation in diabetic nephropathy. TM's lectin-like domain constrains glucose-induced complement activation on endothelial cells and podocytes and ameliorates albuminuria and glomerular damage in mice.
    Type of Publication: Journal article published
    PubMed ID: 23014597
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  • 2
    Keywords: DEGRADATION ; BETA-HEXOSAMINIDASE-A ; carbohydrates ; GANGLIOSIDE GM2 ; GM2-ACTIVATOR PROTEIN ; LYSOSOMAL LIPIDS ; MEMBRANES ; MULTIVESICULAR BODIES ; RECOGNITION ; STIMULATION ; ESCHERICHIA-COLI ; SYSTEM ; SYSTEMS ; PROTEIN ; Germany ; EXPRESSION
    Type of Publication: Book chapter
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  • 3
    Keywords: CONJUGATE ; MRP2 ; PROTEIN ; PROTEINS
    Type of Publication: Book chapter
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  • 4
    Keywords: microarrays ; microarray ; PROTEIN ; SUPPORT
    Type of Publication: Book chapter
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  • 5
    Keywords: review ; molecular ; PHASE-II ; HUMAN HEPATOCYTES ; ORGANIC ANION TRANSPORTER ; multidrug resistance ; MOLECULAR-MECHANISMS ; hepatobiliary transport ; TRANSPORTER ; EFFLUX PUMPS ; SUBFAMILY ; multidrug resistance protein ; phase II ; SALT EXPORT PUMP ; MUTANT RATS ; TRANSCELLULAR TRANSPORT ; VECTORIAL TRANSPORT ; uptake ; enzymology ; PHASE ; ENZYME ; methods ; MEMBERS ; CELL-LINES ; mechanisms ; hepatocytes ; resistance ; GLUTATHIONE ; TRANSPORT ; MRP2 ; SUBSTRATE-SPECIFICITY ; ATP-dependent transport ; MULTIDRUG-RESISTANCE ; cell lines ; CELL-LINE ; MEMBRANE ; LINE ; HUMAN LIVER ; MULTIDRUG-RESISTANCE PROTEIN ; LOCALIZATION ; DNA ; MECHANISM ; PROTEIN ; CLONING ; LINES ; TOOL ; SYSTEMS ; SYSTEM ; ENZYMES ; human ; VITRO ; IN-VITRO ; EXPRESSION ; BLOOD ; Germany
    Abstract: Conjugates of endogenous substances and of xenobiotics, formed extrahepatically or inside hepatocytes, undergo vectorial transport into bile. Substances conjugated with glucuronate, sulfate, or glutathione are substrates for organic anion uptake transporters in the basolateral (sinusoidal) membrane as well as substrates for the unidirectional ATP-driven conjugate efflux pump in the apical (canalicular) membrane, termed multidrug resistance protein 2 (MRP2; systematic name ABCC2). Localization of the efflux pumps ABCC3 and ABCC4 to the basolateral membrane of human hepatocytes has provided insight into the molecular mechanisms of conjugate efflux from hepatocytes into blood, as exemplified by the efflux of bilirubin glucuronosides mediated by ABCC3. The cloning and stable expression of the complementary DNAs encoding the organic anion transporters in the basolateral membrane of human hepatocytes and of members of the ABCC subfamily of efflux pumps in the apical as well as in the basolateral membrane have improved our understanding of hepatobiliary elimination and of the substrate specificity with respect to anionic conjugates. The stable expression of human hepatocyte uptake and efflux transporters in polarized cell lines, as described in this chapter, provides valuable tools for the in vitro analysis of human hepatobiliary transport in general and specifically for uptake and efflux of the anionic conjugates formed in various phase 2 reactions
    Type of Publication: Book chapter
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  • 6
    Keywords: PROTEIN ; MASS-SPECTROMETRY ; tandem mass spectrometry ; ELECTROSPRAY ; mass spectrometry ; MASSES ; analysis ; MASS
    Type of Publication: Book chapter
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  • 7
    Keywords: RESOLUTION ; PROTEIN ; PHOSPHORYLATION ; HIGH-RESOLUTION
    Type of Publication: Book chapter
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  • 8
    Keywords: METASTASIS ; PROTEIN ; CANCER ; EXPRESSION ; cancer research ; BONE ; protein expression
    Type of Publication: Book chapter
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  • 9
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    ELSEVIER ACADEMIC PRESS INC
    Keywords: BIOLOGY ; BINDING ; MOBILITY ; SPECTROSCOPY ; DOMAINS ; sensitivity ; MOTION ; LIVING CELLS ; FLUORESCENCE ; PROTEIN-PROTEIN INTERACTIONS ; Jun ; DNA ; PROTEINS ; PROTEIN ; PATHWAY ; CELLS ; CELL ; Germany ; interactions ; CELL BIOLOGY ; FCS ; interaction ; CROSS-CORRELATION SPECTROSCOPY ; CORRELATION SPECTROSCOPY ; fluorescence correlation spectroscopy ; DIFFUSION ; molecular ; review ; RE ; correlation ; USA ; technique ; FOS ; methods
    Abstract: Fluorescence correlation spectroscopy (FCS) is an emerging technique where the interaction between biomolecules is detected through their correlated motion. It offers the advantage of high (single-molecule) sensitivity; independence of molecular orientation or distance; and simultaneous measurement of molecular interactions, concentrations, and mobilities. Here we introduce the principle of the technique and review some recent examples from the literature where FCS has been used with autofluorescent proteins for measuring protein-protein interactions and mobilities in living cells
    Type of Publication: Book chapter
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  • 10
    Keywords: BIOLOGY ; ARCHITECTURE ; cytoskeleton ; PROTEIN ; CELL ; MICROSCOPY ; intermediate filament ; AMINO-ACID-SEQUENCE ; CROSS-LINKING ; review ; DESMIN ; ROD DOMAIN ; HUMAN VIMENTIN ; KERATIN FILAMENTS ; RAY SOLUTION SCATTERING ; SWITZERLAND ; COILED-COIL
    Type of Publication: Book chapter
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  • 11
    Keywords: FLUORESCENT ; FLUORESCENT PROTEIN ; methods ; HUMAN-CELLS ; FLUORESCENCE MICROSCOPY ; fluorescent proteins ; FLUORESCENCE ; SERIES ; PHENOTYPE ; BIOLOGY ; CELLS ; CELL ; MICROSCOPY ; human ; IMAGES ; CLASSIFICATION ; PROTEIN ; PROTEINS
    Type of Publication: Book chapter
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  • 12
    Keywords: Rb ; PKB/AKT ; lipid ; mechanism of action ; erufosine ; synergistic combinations ; pAkt ; BCR-ABL ; CML ; caspases ; KINASE INHIBITOR ; LEVEL ; INCREASE ; SUBSTRATE ; AGENT ; signaling ; CASPASE ; INHIBITORS ; protein expression ; MAMMALIAN-CELLS ; PROTEIN-KINASE-C ; leukemia ; FUSION ; LINE ; MEMBRANE ; SIGNAL-TRANSDUCTION ; COMPONENT ; ALKYLPHOSPHOCHOLINES ; ANTILEUKEMIC EFFICACY ; CELL-LINE ; chemotherapy ; cell lines ; p27 ; ENTRY ; HEALTH ; ASSAY ; PROGRESSION ; SIGNAL ; treatment ; TYROSINE KINASE INHIBITOR ; CYCLE ; CELL-LINES ; CELL-CYCLE ; cell cycle ; PROTEIN-KINASE ; signal transduction ; INDUCTION ; REDUCTION ; TYROSINE KINASE ; COMBINATION ; KINASE ; PATHWAY ; THERAPY ; PATHWAYS ; CELL ; INHIBITOR ; APOPTOSIS ; EXPRESSION ; CELLS ; DISEASE ; NEW-YORK ; LINES ; cell line ; DRUG ; PROTEINS ; PROTEIN ; TRANSDUCTION ; cell signaling ; MECHANISM ; PATIENT
    Abstract: The ether lipid analog erufosine (erucylphospho-N,N,N,- trimethylpropylammonium, ErPC3) has high activity against leukemic cells without affecting the normal hematopoiesis. It belongs to the group of alkylphosphocholines (APC) that are inhibitors of protein kinase C and phospholipase C. However, the mechanism of action of erufosine remains rather unclear. We focused on combination effects with the tyrosine kinase inhibitor imatinib mesylate (gleevec, former STI-571 or CGP-57148) against two chronic myeloid leukemia (CML)-derived cell lines (K-562 and BV-173). The influence of erufosine on proteins involved in the phosphatidylinositol-3-phosphate pathway and on expression of the retinoblastoma protein Rb was studied, the latter being a key component for cell cycle entry and progression in mammalian cells. The consecutive treatment of K-562 and BV-173 cells with erufosine (2.5, 5, 15, 30 mu M) and imatinib mesylate (0.05, 0.1 mu M) led to synergism as measured by the MTT-dye reduction assay and this is reason to hypothesize that such combinations could be beneficial for relapsed patients with drug-resistant disease. Whole cell lysates from K-562 and BV-173 were investigated for the expression of Rb, PKB/Akt, pAkt, and p27 by Western blot. Erufosine caused decreases of pAkt and CML fusion protein p210 (BCR-ABL) protein expression, but induced the Rb protein expression in K-562 cells. A parallel increase in p27 level was observed after 24 and 48 h treatment. These alterations in signal transduction could be an explanation for the drug interaction found. Furthermore, Rb is a substrate of caspases and is cleaved during apoptosis as already evidenced for BV-173 cells. Our experimental findings suggest that erufosine acts through induction of changes in protein signaling and especially through Rb induction. This unique mode of action makes it an attractive partner for combination therapies, for example, in combination with imatinib mesylate for treatment of CML
    Type of Publication: Book chapter
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  • 13
    Keywords: BEHAVIOR ; PHENOTYPE ; BIOLOGY ; IDENTIFICATION ; NUCLEI ; RECOGNITION ; culture ; PHENOTYPES ; PATTERNS ; IMAGES ; CLASSIFICATION ; CELLS ; CELL ; Germany ; evaluation ; human ; ACCURACY ; PROTEIN ; PROTEINS ; GENE ; TOOL ; GENOME-WIDE ; MICROSCOPE IMAGES ; SUBCELLULAR STRUCTURES ; TEXTURE ; Cell nuclei ; CELL BIOLOGY ; RE ; review ; HIGH-THROUGHPUT ; SCREEN ; analysis ; methods ; CELL-NUCLEI ; USA ; HUMAN-CELLS ; TOOLS
    Abstract: High-throughput screens of the gene function provide rapidly increasing amounts of data. In particular, the analysis of image data acquired in genome-wide cell phenotype screens constitutes a substantial bottleneck in the evaluation process and motivates the development of automated image analysis tools for large-scale experiments. In this chapter, we present a computational scheme to process multicell time-lapse images as they are produced in high-throughput screens. We describe an approach to automatically segment and classify cell nuclei into different mitotic phenotypes. This enables automated identification of cell cultures that show an abnormal mitotic behavior. Our scheme proves high classification accuracy, suggesting a promising future for automating the evaluation of high-throughput experiments
    Type of Publication: Book chapter
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  • 14
    Keywords: PATTERN ; signaling ; MEMBRANE-PROTEIN ; ROLES ; PATTERNS ; MEMBRANE ; PROTEINS ; PROTEIN
    Type of Publication: Book chapter
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  • 15
    Keywords: genomic ; protein profiling ; PROTEOMICS ; protein microarray ; PROTEIN MICROARRAYS ; TOOL ; microarray ; PROTEIN ; IDENTIFICATION ; microarrays
    Type of Publication: Book chapter
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  • 16
    Keywords: analysis ; CAPILLARIES ; PROTEIN ; mass spectrometry ; MASS-SPECTROMETRY ; PHOSPHORYLATION
    Type of Publication: Book chapter
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  • 17
    Keywords: function ; TRAFFICKING ; PROTEIN
    Type of Publication: Book chapter
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  • 18
    Keywords: INHIBITORS ; KINASES ; pharmacology ; PROTEIN ; KINASE ; INHIBITOR ; protein kinase ; PROTEIN-KINASE ; PKA ; PROTEIN-KINASES
    Type of Publication: Book chapter
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  • 19
    Keywords: gene expression ; TRANSCRIPTION FACTORS ; EXPRESSION ; THERAPY ; GENE ; GENE-EXPRESSION ; PROTEIN ; PROTEINS ; transcription ; TRANSCRIPTION FACTOR
    Type of Publication: Book chapter
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  • 20
    Keywords: KINASE INHIBITOR ; PROTEIN-KINASE ; PROTEIN ; COMPLEX ; COMPLEXES ; INHIBITOR ; KINASE
    Type of Publication: Miscellaneous publication
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  • 21
    Keywords: NS1 PROTEIN ; PROTEIN ; INDUCTION
    Type of Publication: Patent
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  • 22
    Keywords: CELLS ; EXPRESSION ; CELL ; Germany ; PATHWAY ; PATHWAYS ; ACETATE-NONUTILIZING MUTANTS ; CDNA ; CDNA CLONES ; CLONES ; CLONING ; CRASSA ; DISTINCT ; DNA-microarray analysis,transcriptional profiling,correspondence analysis,acetate metabolism,Neurosp ; ENZYMES ; FILAMENTOUS FUNGUS ; GENE ; GENE-EXPRESSION ; GENES ; GENOME ; GENOME SEQUENCE ; HYBRIDIZATION ; microarray ; NMT1 ; PROTEIN ; PROTEINS ; RNA ; SACCHAROMYCES-CEREVISIAE ; SAMPLE ; SAMPLES ; transcription
    Abstract: Nutrient-dependent variations in transcript levels of the filamentous fungus Neurospora crassa were studied on a microarray containing some 4700 cDNAs. Cells were grown in minimal and acetate medium. The isolated RNA was analyzed in comparison to the results obtained upon the hybridization of samples prepared from the RNA of cells grown in full medium. Altogether, 160 cDNA clones exhibited significant variations, falling into five distinct subgroups of very similar transcription profiles. This is indicative of the occurrence of a high degree of co-regulation of genes in N. crassa. Especially the regulation of the expression of proteins involved in metabolic pathways was found to be strongly regulated at the RNA level. (C) 2003 Elsevier Inc. All rights reserved
    Type of Publication: Journal article published
    PubMed ID: 14599890
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  • 23
    Keywords: CELLS ; tumor ; CELL ; Germany ; MICROSCOPY ; DIAGNOSIS ; NEW-YORK ; PROTEIN ; PROTEINS ; ADHESION MOLECULES ; cell line ; COMPONENTS ; CONSTITUTIVE TRANSMEMBRANE GLYCOPROTEIN ; CULTURED-CELLS ; DESMOCOLLIN ; desmoplakin ; desmosome ; DIFFERENTIATION ; EPITHELIA ; HUMAN PLAKOGLOBIN ; INTERMEDIATE-SIZED FILAMENTS ; meninges,meningioma,desmosome,desmocollin 3 ; meningioma ; MOLECULAR CHARACTERIZATION ; MOLECULES ; MONOCLONAL-ANTIBODY ; MR-165000 DESMOGLEIN ; NON-EPIDERMAL DESMOSOMES ; PLAQUE PROTEIN ; SUBDURAL SPACE ; TISSUE ; TUMORS
    Abstract: Intercellular junctions morphologically identical to epithelial desmosomes are known structures in meningiomas and arachnoidal tissue. Desmoplakin as one of the desmosomal plaque components has proven to be a reliable marker for diagnosis of meningeal tumors. Here we demonstrate by immunofluorescence microscopy, immunoblot and reverse transcription-PCR reactions that cells of arachnoidal tissue, of diverse meningioma subtypes and of a meningioma-derived cell line contain the full complement of the typical desmosomal proteins desmoplakin (DP), plakophilin 2 (PP2), desmocollin 2 (Dsc2) and desmoglein 2 (Dsg2). Consequently, all these molecules are suitable for diagnostic applications of meningioma tumors. In addition to these constitutive desmosomal components, representative for single-layered (simple) epithelia, the dural border cells of the arachnoid and about 60% of the meningiomas tested were positive for desmocollin 3 (Dsc3), a protein in epithelia taken as an indicator for differentiation
    Type of Publication: Journal article published
    PubMed ID: 12845453
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  • 24
    Keywords: MODEL ; PROTEIN ; FAMILY ; PROTEIN FAMILIES ; PROTEIN FAMILY ; DOMAIN ; PAIRS ; recombination
    Abstract: There is a limited repertoire of domain families in nature that are duplicated and combined in different ways to form the set of proteins in a genome. Most proteins in both prokaryote and eukaryote genomes consist of two or more domains, and we show that the family size distribution of multi-domain protein families follows a power law like that of individual families. Most domain pairs occur in four to six different domain architectures: in isolation and in combinations with different partners. We showed previously that within the set of all pairwise domain combinations, most small and medium-sized families are observed in combination with one or two other families, while a few large families are very versatile and combine with many different partners. Though this may appear to be a stochastic pattern, in which large families have more combination partners by virtue of their size, we establish here that all the domain families with more than three members in genomes are duplicated more frequently than would be expected by chance considering their number of neighbouring domains. This duplication of domain pairs is statistically significant for between one and three quarters of all families with seven or more members. For the majority of pairwise domain combinations, there is no known three-dimensional structure of the two domains together, and we term these novel combinations. Novel domain combinations are interesting and important targets for structural elucidation, as the geometry and interaction between the domains will help understand the function and evolution of multi-domain proteins. Of particular interest are those combinations that occur in the largest number of multi-domain proteins, and several of these frequent novel combinations contain DNA-binding domains.
    Type of Publication: Journal article published
    PubMed ID: 14649290
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  • 25
    Keywords: EXPRESSION ; INHIBITOR ; Germany ; KINASE ; GENE ; GENES ; PROTEIN ; SACCHAROMYCES-CEREVISIAE ; transcription ; COMPONENTS ; COMPLEX ; COMPLEXES ; cell cycle ; CELL-CYCLE ; CYCLE ; protein kinase ; PROTEIN-KINASE ; IDENTIFICATION ; gene expression ; protein kinase CK2 ; Saccharomyces cerevisiae ; SUBUNIT ; YEAST ; BUDDING YEAST ; DISRUPTION ; EXPRESSION ANALYSIS ; HOLOENZYME ; PHO pathway
    Abstract: The budding yeast Saccharomyces cerevisiae encounters phosphate starvation by the transcription-regulated PHO pathway. We find that genetic perturbation of protein kinase CK2, a conserved tetrameric Ser/Thr phosphotransferase with links to cell cycle and transcription, affects expression of PHO pathway genes in a subunit- and isoform-specific manner. Remarkably, the genes encoding phosphate supplying phosphatases and transporters are significantly repressed, while the genes encoding components of the central pathway regulator complex, a cyclin-dependent kinase (CDK), a cyclin, and a CDK inhibitor, remain unaltered. Thus, perturbation of CK2 uncouples the executive part of the PHO pathway from its cyclin-CDK control complex. (C) 2003 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies
    Type of Publication: Journal article published
    PubMed ID: 12606059
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  • 26
    Keywords: RECEPTOR ; EXPRESSION ; GROWTH ; IN-VIVO ; PATHWAYS ; PROTEIN ; SACCHAROMYCES-CEREVISIAE ; COMPLEX ; COMPLEXES ; BINDING ; IDENTIFICATION ; Saccharomyces cerevisiae ; YEAST ; lifestyle ; BETA ; CONSERVATION ; EGG EXTRACTS ; GTPASE RAN ; IMPORTIN-ALPHA ; LOCALIZATION ; MESSENGER-RNA EXPORT ; NUCLEAR EXPORT RECEPTOR ; NUCLEUS ; TRANSPORT FACTOR ; XENOPUS
    Abstract: The small Ras-like GTPase Ran plays an essential role in the transport of macromolecules in and out of the nucleus and has been implicated in spindle (1, 2) and nuclear envelope formation (3, 4) during mitosis in higher eukaryotes. We identified Saccharomyces cerevisiae open reading frame YGL164c encoding a novel RanGTP-binding protein, termed Yrb30p. The protein competes with yeast RanBP1 (Yrb1p) for binding to the GTP-bound form of yeast Ran (Gsp1p) and is, like Yrb1p, able to form trimeric complexes with RanGTP and some of the karyopherins. In contrast to Yrb1p, Yrb30p does not coactivate but inhibits RanGAP1(Rna1p)-mediated GTP hydrolysis on Ran, like the karyopherins. At steady state, Yrb30p localizes exclusively to the cytoplasm, but the presence of a functional nuclear export signal and the localization of truncated forms of Yrb30p suggest that the protein shuttles between nucleus and cytoplasm and is exported via two alternative pathways, dependent on the nuclear export receptor Xpo1p/Crm1p and on RanGTP binding. Whereas overproduction of the full-length protein and complete deletion of the open reading frame reveal no obvious phenotype, overproduction of C-terminally truncated forms of the protein inhibits yeast vegetative growth. Based on these results and the exclusive conservation of the protein in the fungal kingdom, we hypothesize that Yrb30p represents a novel modulator of the Ran GTPase switch related to fungal lifestyle
    Type of Publication: Journal article published
    PubMed ID: 12578832
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  • 27
    Keywords: CANCER ; CELLS ; INHIBITOR ; INVASION ; tumor ; CELL ; COMBINATION ; Germany ; INHIBITION ; KINASE ; PATHWAY ; THERAPY ; DISEASE ; DISEASES ; SITE ; PROTEIN ; COMPLEX ; COMPLEXES ; MECHANISM ; DOMAIN ; BINDING ; PHOSPHORYLATION ; protein kinase ; PROTEIN-KINASE ; treatment ; CATALYTIC SUBUNIT ; inactivation ; FIBER ; ADHESION ; ATP ; CELL-ADHESION ; crystal structure ; CRYSTAL-STRUCTURE ; MUSCLE ; PKA ; PROTEIN-KINASES
    Abstract: Protein kinases require strict inactivation to prevent spurious cellular signaling; overactivity can cause cancer or other diseases and necessitates selective inhibition for therapy. Rho-kinase is involved in such processes as tumor invasion, cell adhesion, smooth muscle contraction, and formation of focal adhesion fibers, as revealed using inhibitor Y-27632. Another Rho-kinase inhibitor, HA-1077 or Fasudil, is currently used in the treatment of cerebral vasospasm; the related nanomolar inhibitor H-1152P improves on its selectivity and potency. We have determined the crystal structures of HA-1077, H-1152P, and Y-27632 in complexes with protein kinase A (PKA) as a surrogate kinase to analyze Rho-kinase inhibitor binding properties. Features conserved between PKA and Rho-kinase are involved in the key binding interactions, while a combination of residues at the ATP binding pocket that are unique to Rho-kinase may explain the inhibitors' Rho-kinase selectivity. Further, a second H-1152P binding site potentially points toward PKA regulatory domain interaction modulators
    Type of Publication: Journal article published
    PubMed ID: 14656443
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  • 28
    Keywords: PEPTIDE ; RECEPTOR ; CELLS ; EXPRESSION ; Germany ; PROTEIN ; DRUG ; MOLECULES ; LINES ; MICE ; COMPLEX ; murine ; primary ; ANTIGEN ; ANTIGENS ; BIOSYNTHESIS ; LYMPHOCYTES ; antigen presentation ; B-CELLS ; CLASS-II MOLECULES ; EXCHANGE ; H2-O ; HLA-DM ; HLA-DO ; IA MOLECULES ; INVARIANT CHAIN ; MHC MOLECULES ; MONOCLONAL- ANTIBODY ; PEPTIDE REPERTOIRE ; PEPTIDES ; T- CELLS
    Abstract: Peptide loading onto MHC class II molecules takes place in endosomal compartments along the endocytic pathway. There, loading is facilitated by the catalytic function of the accessory molecule H2-M, which helps to exchange the invariant chain-derived CLIP peptide in the groove of class II molecules for antigenic peptide. H2-O is another accessory molecule specific to the class II pathway, which is found tightly associated with H2-M and selectively expressed in B cells. Using stable H2-O ribozyme-antisense transfectants, H2-O overexpressing murine B cell lines, and H2-O-transgenic mice, we investigated the effects of H2-O on antigen presentation. The results show that presentation of a variety of exogenous protein antigens to a panel of T cell hybridomas depended on the levels of H2-O in the antigenpresenting B cells. Thus, increased H2-O expression downmodulated, whereas reduced H2-O levels, enhanced presentation. Presentation of endogenous antigen was also diminished by H2-O. Despite the pronounced effects on antigen presentation, the mass spectrometric profiles of peptides eluted from A(b) molecules were very similar in cells expressing different H2-O levels. The intracellular location of H2-O inhibitory activity was investigated with the drug chloroquine, which prevents acidification of the endocytic pathway. The observations indicate that H2-O predominantly inhibits antigen presentation in early endosomal compartments. Thus, H2-O appears to skew peptide loading to late endosomal/lysosomal compartments. This may favor presentation of antigens taken up by the B cell receptor
    Type of Publication: Journal article published
    PubMed ID: 12645938
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  • 29
    Keywords: EXPRESSION ; tumor ; carcinoma ; CELL ; Germany ; TYROSINE KINASE ; screening ; SITE ; SITES ; DISTINCT ; microarray ; PROTEIN ; TISSUE ; TUMORS ; primary ; GROWTH-FACTOR RECEPTOR ; FREQUENCY ; FREQUENCIES ; STAGE ; PROGRESSION ; immunohistochemistry ; ABERRATIONS ; HEAD ; ONCOPROTEIN ; CARCINOMAS ; NECK ; squamous cell carcinoma ; GREECE ; gene amplification ; head and neck ; laryngeal carcinoma ; OROPHARYNGEAL ; C-MYC ; CANCER PATIENTS ; CYCLIN D1 OVEREXPRESSION ; cytogenetic aberration ; head and neck squamous cell carcinoma (HNSCC) ; immunohistochemistry (IHC) ; MICROARRAY ANALYSIS ; oncoprotein overexpression ; OVEREXPRESSION ; POOR-PROGNOSIS ; tissue microarray (TMA) ; tumor classification
    Abstract: Background: Tissue microarray (TMA) analysis is a high-throughput approach that allows the screening of large tumor collectives for cytogenetic aberrations. In this study, a TMA of a large collection of clinically well-defined primary squamous cell carcinomas of the head and neck (HNSCC) was used to determine the expression of several oncoproteins. Materials and Methods: A TMA containing 547 primary HNSCC was used for the analysis of cyclinD1, c-myc, erbb1 and erbb2 expression by immunohistochemistry (IHC). Results: CyclinD1 and c-myc were overexpressed at higher frequencies in primary pharyngeal and laryngeal carcinomas compared with primary oral carcinomas (p 〈 0.001 and p 〈 0.001), while erbb1 and erbb2 overexpression was associated with oral site (p 〈 0.001 and p = 0.04, respectively). Furthermore, cyclinD1 overexpression correlated with stage IV primary carcinomas (p = 0.04). Conclusion: HNSCC is a heterogenous group of tumors, which, depending on anatomic sites and clinical stage, shows variable expressions of the oncoproteins described. This indicates a specific pathogenic role of these oncoproteins in different subtypes of HNSCC and may have therapeutic implications
    Type of Publication: Journal article published
    PubMed ID: 14666705
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  • 30
    Keywords: OPTIMIZATION ; PEPTIDE ; SPECTRA ; CELLS ; EXPRESSION ; INHIBITOR ; Germany ; PROTEIN ; PROTEINS ; MOLECULES ; MICE ; ACTIVATION ; COMPLEX ; COMPLEXES ; murine ; MEMBER ; MHC ; LYMPHOCYTES ; antigen presentation ; PEPTIDES ; STABILITY ; MHC class I ; DEGRADATION ; SUBUNITS ; TRANSLOCATION ; antigen processing ; CELL-LINE .220 ; HISTOCOMPATIBILITY COMPLEX ; LOADING COMPLEX ; MUTANT MICE ; NEWLY SYNTHESIZED PROTEINS
    Abstract: Tapasin is a member of the MHC class I loading complex where it bridges the TAP peptide transporter to class I molecules. The main role of tapasin is assumed to be the facilitation of peptide loading and optimization of the peptide cargo. Here, we describe another important function for tapasin. In tapasin- deficient (Tpn(-/-)) mice the absence of tapasin was found to have a dramatic effect on the stability of the TAP1/TAP2 heterodimeric peptide transporter. Steady-state expression of TAP protein was reduced more than 100-fold from about 3 x 10(4) TAP molecules per wild-type splenocyte to about 1 x 10(2) TAP per Tpn(-/-) splenocyte. Thus, a major function of murine tapasin appears to be the stabilization of TAR The low amount of TAP molecules in Tpn(-/-) lymphocytes is likely to contribute to the severe impairment of MHC class I expression. Surprisingly, activation of Tpn(-/-) lymphocytes yielded strongly enhanced class I expression comparable to wild-type levels, although TAP expression remained low and in the magnitude of several hundred molecules per cell. The high level of class I on activated Tpn(-/-) cells depended on peptides generated by the proteasome as indicated by blockade with the proteasome-specific inhibitor lactacystin. Lymphocyte activation induced an increase in ubiquitinated proteins that are cleaved into peptides by the proteasome. These findings suggest that in the presence of a large peptide pool in the cytosol, a small number of TAP transporters is sufficient to translocate enough peptides for high class I expression. However, these class I molecules were less stable than those of wild-type cells, indicating that tapasin is not only required for stabilization of TAP but also for optimization of the spectrum of bound peptides
    Type of Publication: Journal article published
    PubMed ID: 12594855
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  • 31
    Keywords: CELLS ; EXPRESSION ; CELL ; Germany ; human ; SYSTEM ; DISTINCT ; PROTEIN ; EPITHELIA ; MOLECULES ; TISSUE ; TISSUES ; SKIN ; GLYCOPROTEIN ; ELEMENTS ; SURFACE ; LOCALIZATION ; GLANDS ; SEGMENTS ; calnexin ; ESTABLISHMENT ; MUCINS ; salivary gland ; sebaceous gland ; SEBACEOUS GLANDS
    Abstract: Calnexin (Cnx) has been characterized as a membrane-bound protein that transiently interacts in a unique chaperone system with newly synthesized glycoproteins in order to allow the establishment of their proper tertiary and, in most cases, quarternary structures. The aim of the study was to identify and to locate the expression of Cnx in the three major salivary glands of humans by different methods. Strong expression of Cnx protein and mRNA were generally found in serous salivary secretory units. With regard to mucous secretory units, expression of Cnx was only detectable at a low level in mucous acinar cells of sublingual glands, but not of submandibular glands. Expression of Cnx was always preserved in the surface epithelium of intralobar and interlobular duct segments. In addition, expression of Cnx was detected in sebaceous glands of parotid tissues, with a distribution pattern resembling that seen in sebaceous glands of the normal skin. In conclusion, production of saliva is associated with the expression of Cnx. Synthesis of molecules in mucous secretory units is not necessarily associated with a strong Cnx expression, whereas synthesis in serous secretory units apparently is. The tissue- specific Cnx expression is also paralleled by the observation that the secretions produced by the major salivary glands differ in their composition and amount
    Type of Publication: Journal article published
    PubMed ID: 12507291
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    Keywords: CELLS ; EXPRESSION ; tumor ; BLOOD ; carcinoma ; human ; MICROSCOPY ; liver ; PROTEIN ; PROTEINS ; TUMORS ; FAMILY ; RAT ; hepatocytes ; MEMBER ; MEMBERS ; antibodies ; MOUSE ; IDENTIFICATION ; RAT-LIVER ; MEMBRANE ; metastases ; CONJUGATE ; LOCALIZATION ; EPITHELIAL-CELLS ; HEPATOCYTE CANALICULAR ISOFORM ; METASTATIC CARCINOMAS ; MULTIDRUG-RESISTANCE PROTEIN ; POLYPEPTIDE OATP2
    Abstract: Transport proteins mediating the selective uptake of organic anions into human hepatocytes include the organic anion transporters SLC21A6 (also termed OATP2, OATP-C, or LST-1) and SLC21A8 (OATP8). Both transporters are localized to the basolateral membrane of human hepatocytes. Because of the importance of these transporters for hepatobiliary elimination, including the removal of bilirubin and its conjugates from the blood circulation, we have generated monoclonal antibodies for studies on the expression and localization of these transport proteins. We describe two antibodies, designated monoclonal antibody MDQ (mMDQ) and monoclonal antibody ESL (mESL), directed against the amino terminus and the carboxyl terminus of human SLC21A6, respectively. Both antibodies have been characterized by immunoblot analysis, immunoprecipitation, and immunofluorescence microscopy. While mESL reacted specifically with SLC21A6, mMDQ detects both SLC21A6 and SLC21A8. Neither of the two antibodies reacted with other human, or with dog, rat, or mouse liver SLC21A family members. Antibody mMDQ may be used for the simultaneous detection of SLC21A6 and SLC21A8 in immunoblotting because of its immunoreactivity with both molecules and because of the different molecular masses of both glycosylated proteins in human hepatocytes. This is exemplified in hepatocellular carcinomas where SLC21A6 and SLC21A8 were differentially synthesized and showed an irregular staining pattern. Both transport proteins have not been detected in human hepatoma HepG2 cells. In routine paraffin sections, 10 of 12 hepatocellular carcinomas were focally positive with antibody mMDQ. In contrast, cholangiocarcinomas and liver metastases of colorectal and pancreatic adenocarcinoma were negative without exception. This suggests the usefulness of SLC21A6/SLC21A8 within a panel of tumor markers for hepatocellular carcinomas. Moreover, both antibodies should be useful in studies on the expression and localization of two important uptake transporters of human hepatocytes under physiologic and pathophysiologic conditions
    Type of Publication: Journal article published
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    Keywords: CANCER ; EXPRESSION ; carcinoma ; CELL ; Germany ; human ; LUNG ; lung cancer ; LUNG-CANCER ; EXPOSURE ; RISK ; GENE ; PROTEIN ; METABOLISM ; TISSUE ; PATIENT ; RISK-FACTORS ; FREQUENCY ; polymorphism ; SUSCEPTIBILITY ; PROMOTER ; OVARIAN-CANCER ; WOMEN ; MEN ; risk factors ; smoking ; PROSTATE-CANCER ; cancer risk ; RISK FACTOR ; CYP3A4 ; LINKAGE DISEQUILIBRIUM ; CANCER-PATIENTS ; CARCINOMAS ; POLYMERASE-CHAIN-REACTION ; adenocarcinoma ; ADENOCARCINOMAS ; CARRIERS ; case-control studies ; CLINICAL PRESENTATION ; CYP3A,genetic polymorphism,lung cancer susceptibility,small cell lung cancer,LightCycler ; EXPRESSED HUMAN CYTOCHROME-P450S ; GENETIC VARIANT ; HUMAN LIVER-MICROSOMES ; PROSTATE TUMORS ; PROTEIN LEVELS ; squamous cell carcinoma ; TOBACCO
    Abstract: CYP3A isozymes are involved in tobacco carcinogen- and steroid-metabolism, and are expressed in human lung tissue showing interindividual variation in expression and activity. The CYP3A4* 1 B allele has been associated with a two-fold higher promoter activity and with high-grade prostate cancers. The very frequent intron 3 polymorphism in the CYP3A5 gene (CYP3A5*3) results in decreased CYP3A5 protein levels. A case-control study was conducted in 801 Caucasian lung cancer patients that included 330 adenocarcinomas, 260 squamous cell carcinomas, 171 small cell lung cancers (SCLC) and 432 Caucasian hospital-based controls. CYP3A-genotyping was performed by capillary polymerase chain reaction followed by fluorescence-based melting curve analysis. A significantly increased SCLC risk for CYP3A4* 1B allele carriers [odds ratio (OR) 2.25, 95% confidence interval (CI) 1.11-4.55, P = 0.02] was found. After dividing cases and controls by gender, an increased lung cancer risk for CYP3A4* 1B carriers (OR 3.04, 95% CI 0.94-9.90, P= 0.06) for women but not for men (OR 1.00, 95% CI 0.56-1.81) was revealed. Heavier smoking men (greater than or equal to 20 pack-years) with the CYP3A4* 1 B allele had a significant OR for lung cancer of 3.42 (95% CI 1.65-7.14, P= 0.001) compared to * 1A/1* 1A carriers with lower tobacco exposure (〈 20 pack-years). For women, the respective OR was 8.00 (95% CI 2.12-30.30, P = 0.005). Genotype frequencies were generally in Hardy-Weinberg equilibrium, except for CYP3A5 where a greater than expected number of CYP3A5* 1 homozygotes was observed among cases (P = 0.006). In addition, we observed linkage disequilibrium of CYP3A4 and CYP3A5 (P 〈 0.00001), but a nonsignificantly increased lung cancer risk was only found for homozygous CYP3A5* 1 allele carriers (OR 5.24,95% CI 0.85-102.28, P = 0.14) but not for heterozygotes. To confirm our observation that the CYP3A4* 1B allele increases SCLC risk and modifies the smoking-related lung cancer risk in a gender-specific manner, further studies, including CYP3A haplotype analysis, will be necessary. Pharmacogenetics 13:607-618 (C) 2003 Lippincott Williams Wilkins
    Type of Publication: Journal article published
    PubMed ID: 14515059
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    Keywords: PEPTIDE ; human ; DISEASE ; SITE ; SITES ; PROTEIN ; SAMPLE ; SAMPLES ; SERA ; INDUCTION ; BINDING ; treatment ; ACID ; IDENTIFICATION ; SUBUNIT ; DIFFERENCE ; MOBILITY ; GAS ; ACUTE LYMPHOBLASTIC-LEUKEMIA ; IONIZATION ; ACETYLATED SIALIC ACIDS ; sialic acid ; VISCERAL LEISHMANIASIS ; C-REACTIVE PROTEIN ; acute-phase protein ; BINDING CHARACTERISTICS ; CATLA-CATLA ; CHROMATOGRAPHY ; ELECTROPHORESIS ; FRAGMENTS ; GLUCOSE ; glycosylation ; INDIVIDUALS ; LABEO-ROHITA ; LECTIN ; lectin binding ; LIQUID-CHROMATOGRAPHY ; MAJOR CARP ; matrix-assisted laser-desorption ionization analysis (MALDI analysis) ; molecular modelling ; protein-protein interaction ; PROTEIN-PROTEIN INTERACTIONS ; SIALIC-ACID ; SUBUNITS
    Abstract: As an acute-phase protein, human C-reactive protein (CRP) is clinically important. CRPs were purified from several samples in six different pathological conditions, where their levels ranged from 22 to 342 mug/ml. Small, but significant, variations in electrophoretic mobilities on native PAGE suggested differences in molecular mass, charge and/or shape. Following separation by 9 SDS/PAGE, the), showed single subunits with some differences in their molecular masses ranging between 27 and 30.5 kDa, but for a particular disease, the mobility was the same for CRPs purified from multiple individuals or pooled sera. Isoelectric focusing (IEF) also indicated that the purified CRPs differed from each other. Glycosylation was demonstrated in these purified CRPs by Digoxigenin kits, neuraminidase treatment and binding with lectins. The presence of N-linked sugar moiety was confirmed by N-glycosidase F digestion. The presence of sialic acid, glucose, galactose and mannose has been demonstrated by gas liquid chromatography, mass spectroscopic and fluorimetric analysis. Matrix-assisted laser-desorption ionization analysis of the tryptic digests of three CRPs showed systematic absence of two peptide fragments, one at the N-terminus and the other near the C-terminus. Model-building suggested that the loss of these fragments exposed two potential glycosylation sites on a cleft floor keeping the protein-protein interactions in pentraxins and calcium-dependent phosphorylcholine-binding qualitatively unaffected. Thus we have convincingly demonstrated that human CRP is glycosylated in some pathological conditions
    Type of Publication: Journal article published
    PubMed ID: 12693993
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    Keywords: CELLS ; EXPRESSION ; THERAPY ; GENE ; PROTEIN ; gene therapy ; TRANSDUCTION ; GENE-TRANSFER ; DNA ; IDENTIFICATION ; VECTORS ; AAV SEROTYPES ; adeno-associated virus ; ADENOASSOCIATED VIRUS ; HIGH-LEVEL EXPRESSION ; REP ; serotypes ; TYPE-2 ; vector production
    Abstract: We present a simple and safe strategy for producing high-titer adeno-associated virus (AAV) vectors derived from six different AAV serotypes (AAV-1 to AAV-6). The method, referred to as "HOT," is helper virus free, optically controllable, and based on transfection of only two plasmids, i.e., an AAV vector construct and one of six novel AAV helper plasmids. The latter were engineered to carry AAV serotype rep and cap genes together with adenoviral helper functions, as well as unique fluorescent protein expression cassettes, allowing confirmation of successful transfection and identification of the transfected plasmid. Cross-packaging of vector DNA derived from AAV-2, -3, or -6 was up to 10-fold more efficient using our novel plasmids, compared to a conservative adenovirus-dependent method. We also identified a variety of useful antibodies, allowing detection of Rep or VP proteins, or assembled capsids, of all six AAV serotypes. Finally, we describe unique cell tropisms and kinetics of transgene expression for AAV serotype vectors in primary or transformed cells from four different species. In sum, the HOT strategy and the antibodies presented here, together with the reported findings, should facilitate and support the further development of AAV serotype vectors as powerful new tools for human gene therapy
    Type of Publication: Journal article published
    PubMed ID: 12788658
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    Keywords: CANCER ; BLOOD ; DISEASE ; RISK ; GENE ; PROTEIN ; SAMPLES ; PATIENT ; DNA ; FAMILY ; FREQUENCY ; BREAST ; BREAST-CANCER ; family history ; OVARIAN-CANCER ; WOMEN ; MUTATION ; MUTATIONS ; PREVALENCE ; BRCA1/2 ; BRCA2 MUTATIONS ; early-onset breast cancer ; German population ; germline mutations ; POPULATION-BASED SAMPLE
    Abstract: This study was undertaken to investigate the prevalence of BRCA1 and BRCA2 germline mutations in 91 German patients unselected for family history, who were diagnosed with breast cancer before the age of 41 years. Clinical information and blood samples were obtained from all patients. A comprehensive BRCA1 and BRCA2 mutational analysis was performed using the protein truncation assay and single-strand conformational polymorphism analysis followed by DNA sequencing of variant signals detected by these assays. Five different deleterious germline mutations including four frameshift mutations and one missense mutation were identified, three in BRCA1 (3.3%) and two mutations (2.2%) in BRCA2. Both BRCA2 mutations are novel and might be specific for the German population. An additional BRCA1 missense mutation previously described and classified as an unknown variant was found. This mutation was also detected in two breast cancer patients of family P 328 and not in 140 healthy controls suggesting that it is disease associated. In addition, one common polymorphism and five novel intronic sequence variants with unknown significance were found. Our findings show that mutations in BRCA1 and BRCA2 may contribute similarly to early-onset breast cancer in Germany. Given current constraints on health-care resources, these results support the notion that BRCA1 and BRCA2 mutation screening may have the strongest impact on health-care when targeted to high- risk populations
    Type of Publication: Journal article published
    PubMed ID: 12774040
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    Keywords: RECEPTOR ; IN-VIVO ; GENE ; PROTEIN ; animals ; PROMOTERS ; MAMMALIAN-CELLS ; COMPLEMENTATION ; LUCIFERASE ; REPORTER GENE-EXPRESSION
    Abstract: Genomic research is expected to generate new types of complex observational data, changing the types of experiments as well as our understanding of biological processes. The investigation and definition of relationships among proteins is essential for understanding the function of each gene and the mechanisms of biological processes that specific genes are involved in. Recently, a study by Paulmurugan et al. demonstrated a tool for in vivo noninvasive imaging of protein-protein interactions and intracellular networks
    Type of Publication: Journal article published
    PubMed ID: 12788540
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    Keywords: CELLS ; EXPRESSION ; IN-VITRO ; CELL ; human ; IN-VIVO ; VITRO ; VIVO ; NETWORK ; GENE ; PROTEIN ; PROTEINS ; TIME ; INFECTION ; RAT ; CONTRAST ; STAGE ; DISRUPTION ; MUTATION ; MUTATIONS ; MUSCLE ; ASSEMBLY PROPERTIES ; SERIES ; GREEN FLUORESCENT PROTEIN ; ORGANIZATION ; ADENOVIRUS ; ALPHA-B-CRYSTALLIN ; DILATED CARDIOMYOPATHY ; EPIDERMOLYSIS-BULLOSA SIMPLEX ; INTERMEDIATE-FILAMENT PROTEINS ; MICE LACKING DESMIN ; MUSCULAR-DYSTROPHY ; SKELETAL MYOPATHY ; SMOOTH-MUSCLE ; Z-DISCS
    Abstract: Mutations in desmin have been associated with a subset of human myopathies. Symptoms typically appear in the second to third decades of life, but in the most severe cases can manifest themselves earlier. How desmin mutations lead to aberrant muscle function, however, remains poorly defined. We created a series of four mutations in rat desmin and tested their in vitro filament assembly properties. RDM-G, a chimera between desmin and green fluorescent protein, formed protofilament-like structures in vitro. RDM-1 and RDM-2 blocked in vitro assembly at the unit-length filament stage, while RDM-3 had more subtle effects on assembly. When expressed in cultured rat neonatal cardiac myocytes via adenovirus infection, these mutant proteins disrupted the endogenous desmin filament to an extent that correlated with their defects in in vitro assembly properties. Disruption of the desmin network by RDM-1 was also associated with disruption of plectin, myosin, and a-actinin organization in a significant percentage of infected cells. In contrast, expression of RDM-2, which is similar to previously characterized human mutant desmins, took longer to disrupt desmin and plectin organization and had no significant effect on myosin or alpha-actinin organization over the 5-day time course of our studies. RDM-3 had the mildest effect on in vitro assembly and no discernable effect on either desmin, plectin, myosin, or a-actinin organization in vivo. These results indicate that mutations in desmin have both direct and indirect effects on the cytoarchitecture of cardiac myocytes
    Type of Publication: Journal article published
    PubMed ID: 12529857
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    Keywords: IN-VITRO ; IONIZING-RADIATION ; IRRADIATION ; CELL ; Germany ; human ; IN-VIVO ; KINASE ; PATHWAY ; VITRO ; VIVO ; SITE ; PROTEIN ; radiation ; ACTIVATION ; COMPLEX ; COMPLEXES ; DNA ; CARCINOGENESIS ; cell cycle ; CELL-CYCLE ; CYCLE ; PHOSPHORYLATION ; MUTANT ; LESIONS ; PROGRESSION ; CYCLE PROGRESSION ; DAMAGE ; DNA-DAMAGE ; DEGRADATION ; ATR ; CHK1 ; CYCLE CONTROL ; G(1)/S TRANSITION ; PROTEASOME ; S-PHASE ; serine
    Abstract: The human Cdc25A phosphatase plays a pivotal role at the G(1)/S transition by activating cyclin E and A/Cdk2 complexes through dephosphorylation. In response to ionizing radiation, Cdc25A is phosphorylated by both Chk1 and Chk2 on Ser-123. This in turn leads to ubiquitylation and rapid degradation of Cdc25A by the proteasome resulting in cell cycle arrest. We found that in response to UV irradiation, Cdc25A is phosphorylated at a different serine residue, Ser-75. Significantly, Cdc25A mutants carrying alanine instead of either Ser-75 or Ser-123 demonstrate that only Ser-75 mediates protein stabilization in response to UV-induced DNA damage. As a consequence, cyclin E/Cdk2 kinase activity was high. Furthermore, we find that Cdc25A was phosphorylated by Chk1 on Ser-75 in vitro and that the same site was also phosphorylated in vivo. Taken together, these data strongly suggest that phosphorylation of Cdc25A on Ser-75 by Chk1 and its subsequent degradation is required to delay cell cycle progression in response to UV-induced DNA lesions
    Type of Publication: Journal article published
    PubMed ID: 12759351
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    Keywords: GROWTH ; tumor ; Germany ; MORTALITY ; NEW-YORK ; RISK ; PROTEIN ; PATIENT ; TUMOR-NECROSIS-FACTOR ; INTERVENTION ; treatment ; PLASMA ; DECREASE ; AGE ; MUSCLE ; AMINO-ACIDS ; OXIDATIVE STRESS ; ANTIOXIDANT ; aging-related wasting ; ALPHA-LIPOIC ACID ; antioxidants and aging ; cysteine ; ELDERLY HUMANS ; INJURIOUS FALLS ; MUSCLE PROTEIN-SYNTHESIS ; muscular aging ; P70 S6 KINASE ; PLASMA REDOX STATE ; RAT SKELETAL-MUSCLE ; RESISTANCE EXERCISE ; role in aging ; tumor necrosis factor in aging
    Abstract: Aging-related loss of muscle function is a predictor of mortality and a surrogate parameter of the aging process. Its consequences include a high risk for falls, hip fractures, and loss of autonomy. Aging is associated with changes in the oxidant/antioxidant balance including a decrease in plasma thiol (cysteine) concentration. To assess the importance of cysteine, we determined in a double-blind study the effects of N-acetylcysteine on the functional capacity of frail geriatric patients and their response to physical exercise. The subjects on placebo showed only a relatively weak response, and 31% showed even a decrease in more than one parameter during the observation period. Low plasma arginine levels were correlated with a weak overall performance before exercise and a poor response to exercise. N-Acetyl-cysteine strongly enhanced the increase in knee extensor strength and significantly increased the sum of all strength parameters if adjusted for baseline arginine level as a confounding parameter. N-acetylcysteine had no significant effect on growth hormone and IGF-1 levels but caused a significant decrease in plasma TNF-alpha. These findings may provide a basis for therapeutic intervention and suggest that the loss of function involves limitations in cysteine and one or more other amino acids which may compromise muscular protein synthesis
    Type of Publication: Journal article published
    PubMed ID: 12601528
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    Keywords: GROWTH ; PROTEIN ; DIFFERENTIATION ; DNA ; DATABASE ; INTERFACE ; W2H
    Abstract: In high throughput sequence analysis, it is often necessary to combine the results of contemporary bioinformatics tools, because no individual tool alone computes all the requested information. ESTAnnotator is a tool for the high throughput annotation of expressed sequence tags (ESTs) by automatically running a collection of bioinformatics applications. In the first step, a quality check is performed and repeats, vector parts and low quality sequences are masked. Then successive steps of database searching and EST clustering are performed. Already known transcripts present within mRNA and genomic DNA reference databases are identified. Subsequently, tools for the clustering of anonymous ESTs, and for further database searches at the protein level, are applied. Finally, the outputs of each individual tool are gathered and the relevant results presented in a descriptive summary. ESTAnnotator was already successfully applied for the systematic identification and characterisation of novel human genes involved in cartilage/bone formation, growth, differentiation and homeostasis. ESTAnnotator is available at http://genome.dkfz-heidelberg.de, contact: genome@dkfz.de
    Type of Publication: Journal article published
    PubMed ID: 12824401
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