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  • 1
    Keywords: brain ; CELLS ; EXPRESSION ; Germany ; GENE ; GENES ; GENOME ; PROTEIN ; PROTEINS ; transcription ; TISSUE ; MECHANISM ; mechanisms ; PROMOTER ; NUMBER ; DATABASE ; LOCALIZATION ; B-CELLS ; INVOLVEMENT ; TESTIS ; representational difference analysis ; RE ; VARIANT ; genomics ; regulation ; TRANSLATION ; GENE-REGULATION ; gene regulation ; NUCLEAR-PORE COMPLEX ; OVERLAPPING READING FRAMES ; SIGNAL PEPTIDES
    Abstract: Background: Given the complexity of higher organisms, the number of genes encoded by their genomes is surprisingly small. Tissue specific regulation of expression and splicing are major factors enhancing the number of the encoded products. Commonly these mechanisms are intragenic and affect only one gene. Results: Here we provide evidence that the IL4I1 gene is specifically transcribed from the apparent promoter of the upstream NUP62 gene, and that the first two exons of NUP62 are also contained in the novel IL4I1_2 variant. While expression of IL4I1 driven from its previously described promoter is found mostly in B cells, the expression driven by the NUP62 promoter is restricted to cells in testis (Sertoli cells) and in the brain (e.g., Purkinje cells). Since NUP62 is itself ubiquitously expressed, the IL4I1_2 variant likely derives from cell type specific alternative pre-mRNA processing. Conclusion: Comparative genomics suggest that the promoter upstream of the NUP62 gene originally belonged to the IL4I1 gene and was later acquired by NUP62 via insertion of a retroposon. Since both genes are apparently essential, the promoter had to serve two genes afterwards. Expression of the IL4I1 gene from the "NUP62" promoter and the tissue specific involvement of the pre-mRNA processing machinery to regulate expression of two unrelated proteins indicate a novel mechanism of gene regulation
    Type of Publication: Journal article published
    PubMed ID: 16029492
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  • 2
    Keywords: EXPRESSION ; proliferation ; CELL-PROLIFERATION ; Germany ; INFORMATION ; screening ; GENE ; GENE-EXPRESSION ; GENES ; GENOME ; PROTEIN ; PROTEINS ; gene expression ; ASSAY ; DATABASE ; bioinformatics ; INTERFACE ; PROJECT ; INTEGRATION ; FEATURES ; RE ; cell proliferation ; FULL-LENGTH HUMAN ; HUMAN CDNAS ; ASSAYS ; genomic ; NORTHERN
    Abstract: LIFEdb (http://www.LIFEdb.de) integrates data from large-scale functional genomics assays and manual cDNA annotation with bioinformatics gene expression and protein analysis. New features of LIFEdb include (i) an updated user interface with enhanced query capabilities, (ii) a configurable output table and the option to download search results in XML, (iii) the integration of data from cell-based screening assays addressing the influence of protein-overexpression on cell proliferation and (iv) the display of the relative expression ('Electronic Northern') of the genes under investigation using curated gene expression ontology information. LIFEdb enables researchers to systematically select and characterize genes and proteins of interest, and presents data and information via its user-friendly web-based interface
    Type of Publication: Journal article published
    PubMed ID: 16381901
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  • 3
    Keywords: PEPTIDE ; EXPRESSION ; Germany ; ALGORITHM ; DISTINCT ; GENOME ; PROTEIN ; ACCURACY ; COMPLEX ; COMPLEXES ; SEQUENCE ; SEQUENCES ; MOUSE ; IDENTIFICATION ; PATTERNS ; Drosophila ; NUMBER ; HUMAN GENOME ; LOCALIZATION ; PROTEOMICS ; PROGRAM ; RE ; INCREASE ; REQUIREMENT ; NUCLEOTIDE-SEQUENCES ; FULL-LENGTH HUMAN ; HUMAN CDNAS ; FIDELITY ; Internet
    Abstract: Background: The identification of patterns in biological sequences is a key challenge in genome analysis and in proteomics. Frequently such patterns are complex and highly variable, especially in protein sequences. They are frequently described using terms of regular expressions (RegEx) because of the user-friendly terminology. Limitations arise for queries with the increasing complexity of patterns and are accompanied by requirements for enhanced capabilities. This is especially true for patterns containing ambiguous characters and positions and/or length ambiguities. Results: We have implemented the 3of5 web application in order to enable complex pattern matching in protein sequences. 3of5 is named after a special use of its main feature, the novel n-of-m pattern type. This feature allows for an extensive specification of variable patterns where the individual elements may vary in their position, order, and content within a defined stretch of sequence. The number of distinct elements can be constrained by operators, and individual characters may be excluded. The n-of-m pattern type can be combined with common regular expression terms and thus also allows for a comprehensive description of complex patterns. 3of5 increases the fidelity of pattern matching and finds ALL possible solutions in protein sequences in cases of length-ambiguous patterns instead of simply reporting the longest or shortest hits. Grouping and combined search for patterns provides a hierarchical arrangement of larger patterns sets. The algorithm is implemented as internet application and freely accessible. The application is available at http://dkfz.de/mga2/3of5/3of5.html. Conclusion: The 3of5 application offers an extended vocabulary for the definition of search patterns and thus allows the user to comprehensively specify and identify peptide patterns with variable elements. The n-of-m pattern type offers an improved accuracy for pattern matching in combination with the ability to find all solutions, without compromising the user friendliness of regular expression terms
    Type of Publication: Journal article published
    PubMed ID: 16542452
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  • 4
    Keywords: CELLS ; human ; DISTINCT ; GENE ; GENES ; PROTEIN ; PROTEINS ; COMPLEX ; DOMAIN ; SEQUENCE ; SEQUENCES ; VARIANTS ; MOUSE ; IDENTIFICATION ; PATTERNS ; PROMOTERS ; HUMAN GENOME ; LOCALIZATION ; KAPPA-B ; DOMAINS ; SUBCELLULAR-LOCALIZATION ; RE ; VARIANT ; LOCUS ; EVENTS ; OPEN READING FRAMES ; function ; SPLICING VARIANTS ; transcriptome ; MAMMALIAN GENOMES ; PRE-MESSENGER-RNA
    Abstract: We report the first genome-wide identification and characterization of alternative splicing in human gene transcripts based on analysis of the full-length cDNAs. Applying both manual and computational analyses for 56 419 completely sequenced and precisely annotated full-length cDNAs selected for the H-Invitational human transcriptome annotation meetings, we identified 6877 alternative splicing genes with 18 297 different alternative splicing variants. A total of 37 670 exons were involved in these alternative splicing events. The encoded protein sequences were affected in 6005 of the 6877 genes. Notably, alternative splicing affected protein motifs in 3015 genes, subcellular localizations in 2982 genes and transmembrane domains in 1348 genes. We also identified interesting patterns of alternative splicing, in which two distinct genes seemed to be bridged, nested or having overlapping protein coding sequences (CDSs) of different reading frames (multiple CDS). In these cases, completely unrelated proteins are encoded by a single locus. Genome-wide annotations of alternative splicing, relying on full-length cDNAs, should lay firm groundwork for exploring in detail the diversification of protein function, which is mediated by the fast expanding universe of alternative splicing variants
    Type of Publication: Journal article published
    PubMed ID: 16914452
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  • 5
    Keywords: EXPRESSION ; Germany ; PATHWAY ; CLASSIFICATION ; GENE ; GENE-EXPRESSION ; GENES ; GENOME ; ACCURACY ; BIOLOGY ; TARGET ; gene expression ; MICROARRAY DATA ; DATABASE ; RE ; databases ; analysis ; methods ; HIGH-THROUGHPUT ; technique ; CANDIDATE ; microbiology ; ENGLAND ; biotechnology ; ONTOLOGIES
    Abstract: Background: High-throughput technologies like functional screens and gene expression analysis produce extended lists of candidate genes. Gene-Set Enrichment Analysis is a commonly used and well established technique to test for the statistically significant over-representation of particular pathways. A shortcoming of this method is however, that most genes that are investigated in the experiments have very sparse functional or pathway annotation and therefore cannot be the target of such an analysis. The approach presented here aims to assign lists of genes with limited annotation to previously described functional gene collections or pathways. This works by comparing InterPro domain signatures of the candidate gene lists with domain signatures of gene sets derived from known classifications, e. g. KEGG pathways. Results: In order to validate our approach, we designed a simulation study. Based on all pathways available in the KEGG database, we create test gene lists by randomly selecting pathway genes, removing these genes from the known pathways and adding variable amounts of noise in the form of genes not annotated to the pathway. We show that we can recover pathway memberships based on the simulated gene lists with high accuracy. We further demonstrate the applicability of our approach on a biological example. Conclusion: Results based on simulation and data analysis show that domain based pathway enrichment analysis is a very sensitive method to test for enrichment of pathways in sparsely annotated lists of genes. An R based software package domainsignatures, to routinely perform this analysis on the results of high-throughput screening, is available via Bioconductor
    Type of Publication: Journal article published
    PubMed ID: 18177498
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  • 6
    Keywords: CANCER ; GROWTH ; INHIBITOR ; proliferation ; SURVIVAL ; tumor ; CELL-PROLIFERATION ; Germany ; KINASE ; INFORMATION ; TOOL ; DISEASE ; GENE ; GENES ; GENOME ; microarray ; PROTEIN ; PROTEINS ; transcription ; TUMORS ; RESOLUTION ; ACTIVATION ; DNA ; BIOLOGY ; cell cycle ; CELL-CYCLE ; CYCLE ; ASSOCIATION ; MOUSE ; IDENTIFICATION ; PROGRESSION ; ASSAY ; microarrays ; PROSTATE-CANCER ; STRATEGIES ; DNA-REPLICATION ; REPLICATION ; signaling ; RE ; TUMORIGENICITY ; genomics ; TRANSITION ; DNA replication ; C-ELEGANS ; cell proliferation ; PROTEIN-ANALYSIS ; development ; ASSAYS ; DIFFERENTIALLY EXPRESSED GENES ; high throughput ; HIGH-THROUGHPUT ; LONG ; PRIME ; PRINCIPLES ; REPRESSOR ; ROLES
    Abstract: Cancer transcription microarray studies commonly deliver long lists of "candidate" genes that are putatively associated with the respective disease. For many of these genes, no functional information, even less their relevance in pathologic conditions, is established as they were identified in large-scale genomics approaches. Strategies and tools are thus needed to distinguish genes and proteins with mere tumor association from those causally related to cancer. Here, we describe a functional profiling approach, where we analyzed 103 previously uncharacterized genes in cancer relevant assays that probed their effects on DNA replication (cell proliferation). The genes had previously been identified as differentially expressed in genome-wide microarray studies of tumors. Using an automated high-throughput assay with single-cell resolution, we discovered seven activators and nine repressors of DNA replication. These were further characterized for effects on extracellular signal-regulated kinase 1/2 (ERK1/2) signaling (G(1)-S transition) and anchorage-independent growth (tumorigenicity). One activator and one inhibitor protein of ERK1/2 activation and three repressors of anchorage-independent growth were identified. Data from tumor and functional profiling make these proteins novel prime candidates for further in-depth study of their roles in cancer development and progression. We have established a novel functional profiling strategy that links genomics to cell biology and showed its potential for discerning cancer relevant modulators of the cell cycle in the candidate lists from microarray studies
    Type of Publication: Journal article published
    PubMed ID: 16140941
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  • 7
    Keywords: Germany ; CLASSIFICATION ; INFORMATION ; TOOL ; SITE ; CLONING ; GENOME ; PROTEIN ; PROTEINS ; TIME ; SEQUENCE ; SIGNAL ; VARIANTS ; ASSAY ; DATABASE ; LOCALIZATION ; PREDICTION ; SELECTION ; REJECTION ; SEQUENCE-ANALYSIS ; HUMAN GENES ; FUNCTIONAL GENOMICS ; CDNAS ; FEATURES ; PROGRAM ; RE ; VARIANT ; assembly ; databases ; ANNOTATION ; CPG ISLANDS ; FULL-LENGTH HUMAN ; ASSAYS ; HIGH-THROUGHPUT ; TESTS ; GENOMIC DNA ; genomic ; SIGNALS ; E ; SET ; transcriptome ; POLYADENYLATION
    Abstract: Background: The German cDNA Consortium has been cloning full length cDNAs and continued with their exploitation in protein localization experiments and cellular assays. However, the efficient use of large cDNA resources requires the development of strategies that are capable of a speedy selection of truly useful cDNAs from biological and experimental noise. To this end we have developed a new high-throughput analysis tool, CAFTAN, which simplifies these efforts and thus fills the gap between large-scale cDNA collections and their systematic annotation and application in functional genomics. Results: CAFTAN is built around the mapping of cDNAs to the genome assembly, and the subsequent analysis of their genomic context. It uses sequence features like the presence and type of PolyA signals, inner and flanking repeats, the GC-content, splice site types, etc. All these features are evaluated in individual tests and classify cDNAs according to their sequence quality and likelihood to have been generated from fully processed mRNAs. Additionally, CAFTAN compares the coordinates of mapped cDNAs with the genomic coordinates of reference sets from public available resources ( e. g., VEGA, ENSEMBL). This provides detailed information about overlapping exons and the structural classification of cDNAs with respect to the reference set of splice variants. The evaluation of CAFTAN showed that is able to correctly classify more than 85% of 5950 selected "known protein-coding" VEGA cDNAs as high quality multi-or single-exon. It identified as good 80.6% of the single exon cDNAs and 85% of the multiple exon cDNAs. The program is written in Perl and in a modular way, allowing the adoption of this strategy to other tasks like EST-annotation, or to extend it by adding new classification rules and new organism databases as they become available. We think that it is a very useful program for the annotation and research of unfinished genomes. Conclusion: CAFTAN is a high-throughput sequence analysis tool, which performs a fast and reliable quality prediction of cDNAs. Several thousands of cDNAs can be analyzed in a short time, giving the curator/scientist a first quick overview about the quality and the already existing annotation of a set of cDNAs. It supports the rejection of low quality cDNAs and helps in the selection of likely novel splice variants, and/or completely novel transcripts for new experiments
    Type of Publication: Journal article published
    PubMed ID: 17064411
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  • 8
    Keywords: CELLS ; CELL ; Germany ; PATHWAYS ; QUANTIFICATION ; SYSTEM ; SYSTEMS ; microarray ; PROTEIN ; PROTEINS ; SAMPLES ; DRUG ; BIOLOGY ; SIGNAL ; ASSAY ; microarrays ; EFFICIENT ; ELECTROPHORESIS ; BIOPSY ; PROTEOMICS ; signaling ; RECOMBINANT ; RE ; CAPACITY ; ARRAY ; GELS ; analysis ; methods ; BIOPSIES ; signaling networks ; protein arrays ; protein quantification ; reverse phase protein microarray
    Abstract: The advancement of efficient technologies to comply with the needs of systems biology and drug discovery has so far not received adequate attention. A substantial bottleneck for the time-resolved quantitative description of signaling networks is the limited throughput and the inadequate sensitivity of currently established methods. Here, we present an improved protein microarray-based approach towards the sensitive detection of proteins in the fg-range which is based on signal detection in the near-infrared range. The high sensitivity of the assay permits the specific quantification of proteins derived from as little as only 20 000 cells with an error rate of only 5%. The capacity is limited to the analysis of up to 500 different samples per microarray. Protein abundance is determined qualitatively, and quantitatively, if recombinant protein is available. This novel approach was called IPAQ (infrared-based protein arrays with quantitative readout). IPAQ offers a highly sensitive experimental approach superior to the established standard protein quantification technologies, and is suitable for quantitative proteomics. Employing the IPAQ approach, a detailed analysis of activated signaling networks in biopsy samples and of crosstalk between signaling modules as required in drug discovery strategies can easily be performed
    Type of Publication: Journal article published
    PubMed ID: 17309101
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  • 9
    Keywords: CANCER ; CANCER CELLS ; CELLS ; EXPRESSION ; INVASION ; tumor ; TUMOR-CELLS ; carcinoma ; CELL ; Germany ; CLASSIFICATION ; GENE-EXPRESSION ; GENOME ; PROTEIN ; PROTEINS ; TUMORS ; TIME ; kidney ; primary ; FLOW ; BIOLOGY ; CELL-CYCLE ; MOLECULAR-BIOLOGY ; BREAST ; breast cancer ; BREAST-CANCER ; PROGRESSION ; PATTERNS ; MEMBRANE ; METASTASIS ; genetics ; metastases ; CANCER-CELLS ; ONCOGENE ; heredity ; molecular biology ; molecular ; E-cadherin ; ONCOLOGY ; RE ; INCREASE ; LEVEL ; LOSSES ; REDUCED EXPRESSION ; ENGLAND ; INCREASES ; detachment ; cell junctions ; initial cell-cell contact
    Abstract: Vacuole membrane protein 1 (Vmp1) is described as a cancer-relevant cell cycle modulator, but the function of this protein and its mode of action in tumor progression are still unknown. In this study, we show that the VMP1 mRNA level is significantly reduced in kidney cancer metastases as compared to primary tumors. Further, VMP1 expression is also decreased in the invasive breast cancer cell lines HCC1954 and MDA-MB-231 as compared to the non-invasive cell lines MCF-12A, T-47D and MCF-7. We show for the first time that Vmp1 is a plasma membrane protein and an essential component of initial cell-cell contacts and tight junction formation. It interacts with the tight junction protein Zonula Occludens-1 and colocalizes in spots between neighboring HEK293 cells. Downregulation of VMP1 by RNAi results in loss of cell adherence, and increases the invasion capacity of the non-invasive kidney cancer cell line Caki-2. In conclusion, our findings establish Vmp1 to be a novel cell-cell adhesion protein and that its expression level determines the invasion and metastatic potential of cancer cells
    Type of Publication: Journal article published
    PubMed ID: 17724469
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  • 10
    Keywords: EXPRESSION ; Germany ; human ; DISEASE ; GENE ; GENE-EXPRESSION ; GENES ; TISSUE ; TISSUES ; LINKAGE ; MOUSE ; IDENTIFICATION ; IN-SITU ; gene expression ; NUMBER ; DATABASE ; REGION ; REGIONS ; LOCALIZATION ; ORGANIZATION ; RE ; EXPRESSION PATTERNS ; MAP ; MENTAL-RETARDATION ; SUBUNIT PROTEIN ; CPG-BINDING PROTEIN-2
    Abstract: Background: Well known for its gene density and the large number of mapped diseases, the human sub-chromosomal region Xq28 has long been a focus of genome research. Over 40 of approximately 300 X-linked diseases map to this region, and systematic mapping, transcript identification, and mutation analysis has led to the identification of causative genes for 26 of these diseases, leaving another 17 diseases mapped to Xq28, where the causative gene is still unknown. To expedite disease gene identification, we have initiated the functional characterisation of all known Xq28 genes. Results: By using a systematic approach, we describe the Xq28 genes by RNA in situ hybridisation and Northern blotting of the mouse orthologs, as well as subcellular localisation and data mining of the human genes. We have developed a relational web-accessible database with comprehensive query options integrating all experimental data. Using this database, we matched gene expression patterns with affected tissues for 16 of the 17 remaining Xq28 linked diseases, where the causative gene is unknown. Conclusion: By using this systematic approach, we have prioritised genes in linkage regions of Xq28-mapped diseases to an amenable number for mutational screens. Our database can be queried by any researcher performing highly specified searches including diseases not listed in OMIM or diseases that might be linked to Xq28 in the future
    Type of Publication: Journal article published
    PubMed ID: 16503986
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