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Matthias Platzer

Publications and source records attributed to Matthias Platzer.

At least 19 recordsLinked to original sources
PubMed2007

Chicken ovalbumin upstream promoter transcription factor II regulates uncoupling protein 3 gene transcription in Phodopus sungorus.

BACKGROUND: Ucp3 is an integral protein of the inner mitochondrial membrane with a role in lipid metabolism preventing deleterious effects of fatty acids in states of high lipid oxidation. Ucp3 is expressed in brown adipose tissue and skeletal muscle and controlled by a transcription factor complex including PPARalpha, MyoD and the histone acetyltransferase p300. Several studies have demonstrated interaction of these factors with chicken ovalbumin upstream promoter transcription factor II (Coup-TFII). This nuclear receptor is involved in organogenesis and other developmental processes including skeletal muscle development, but also co-regulates a number of metabolic genes. In this study we in silico analyzed the upstream region of Ucp3 of the Djungarian hamster Phodopus sungorus and identified several putative response elements for Coup-TFII. We therefore investigated whether Coup-TFII is a further player in the transcriptional control of the Ucp3 gene in rodents. RESULTS: By quantitative PCR we demonstrated a positive correlation of Coup-TFII and Ucp3 mRNA expression in skeletal muscle and brown adipose tissue in response to food deprivation and cold exposure, respectively. In reporter gene assays Coup-TFII enhanced transactivation of the Ucp3 promoter conveyed by MyoD, PPARalpha, RXRalpha and/or p300. Using deletions and mutated constructs, we identified a Coup-TFII enhancer element 816-840 bp upstream of the transcriptional start site. Binding of Coup-TFII to this upstream enhancer was confirmed in electrophoretic mobility shift and supershift assays. CONCLUSION: Transcriptional regulation of the Coup-TFII gene in response to starvation and cold exposure seems to be the regulatory mechanism of Ucp3 mRNA expression in brown adipose and skeletal muscle tissue determining the final appropriate rate of transcript synthesis. These findings add a crucial component to the complex transcriptional machinery controlling expression of Ucp3. Given the substantial evidence for a function of Ucp3 in lipid metabolism, Coup-TFII may not only be a negative regulator of glucose responsive genes but also transactivate genes involved in lipid metabolism.

Animals↗
PubMed2006

TassDB: a database of alternative tandem splice sites.

Subtle alternative splice events at tandem splice sites are frequent in eukaryotes and substantially increase the complexity of transcriptomes and proteomes. We have developed a relational database, TassDB (TAndem Splice Site DataBase), which stores extensive data about alternative splice events at GYNGYN donors and NAGNAG acceptors. These splice events are of subtle nature since they mostly result in the insertion/deletion of a single amino acid or the substitution of one amino acid by two others. Currently, TassDB contains 114 554 tandem splice sites of eight species, 5209 of which have EST/mRNA evidence for alternative splicing. In addition, human SNPs that affect NAGNAG acceptors are annotated. The database provides a user-friendly interface to search for specific genes or for genes containing tandem splice sites with specific features as well as the possibility to download large datasets. This database should facilitate further experimental studies and large-scale bioinformatics analyses of tandem splice sites. The database is available at http://helios.informatik.uni-freiburg.de/TassDB/.

Alternative Splicing↗
PubMed2006

Comparative genome analysis: selection pressure on the Borrelia vls cassettes is essential for infectivity.

BACKGROUND: At least three species of Borrelia burgdorferi sensu lato (Bbsl) cause tick-borne Lyme disease. Previous work including the genome analysis of B. burgdorferi B31 and B. garinii PBi suggested a highly variable plasmid part. The frequent occurrence of duplicated sequence stretches, the observed plasmid redundancy, as well as the mainly unknown function and variability of plasmid encoded genes rendered the relationships between plasmids within and between species largely unresolvable. RESULTS: To gain further insight into Borreliae genome properties we completed the plasmid sequences of B. garinii PBi, added the genome of a further species, B. afzelii PKo, to our analysis, and compared for both species the genomes of pathogenic and apathogenic strains. The core of all Bbsl genomes consists of the chromosome and two plasmids collinear between all species. We also found additional groups of plasmids, which share large parts of their sequences. This makes it very likely that these plasmids are relatively stable and share common ancestors before the diversification of Borrelia species. The analysis of the differences between B. garinii PBi and B. afzelii PKo genomes of low and high passages revealed that the loss of infectivity is accompanied in both species by a loss of similar genetic material. Whereas B. garinii PBi suffered only from the break-off of a plasmid end, B. afzelii PKo lost more material, probably an entire plasmid. In both cases the vls gene locus encoding for variable surface proteins is affected. CONCLUSION: The complete genome sequences of a B. garinii and a B. afzelii strain facilitate further comparative studies within the genus Borrellia. Our study shows that loss of infectivity can be traced back to only one single event in B. garinii PBi: the loss of the vls cassettes possibly due to error prone gene conversion. Similar albeit extended losses in B. afzelii PKo support the hypothesis that infectivity of Borrelia species depends heavily on the evasion from the host response.

Borrelia↗
PubMed2006

tuple_plot: fast pairwise nucleotide sequence comparison with noise suppression.

SUMMARY: The program tuple_plot identifies and visualizes local similarities between two genomic sequences, typically 100 kb or longer, by applying the well-known dotplot principle. A dictionary of sequence words built from the input sequences serves to construct a task-specific expectancy model that is used to attribute significance values to pairwise word hits. The dictionary-based approach allows fast computation, the computation time scaling to O(N log N), depending on the size of the input sequences. The proposed scoring scheme appreciably increases the signal-to-noise ratio and may help to improve other word-based sequence comparison approaches. AVAILABILITY: tuple_plot is available at http://genome.fli-leibniz.de/software.html and may be used under GNU public license.

Algorithms↗
PubMed2006

Method for preparing single-stranded DNA templates for Pyrosequencing using vector ligation and universal biotinylated primers.

In Pyrosequencing, the addition of nucleotides to a primer-template hybrid is monitored by enzymatic conversion of chemical energy into detectable light. The technique yields both qualitative and quantitative sequence information because the chemical energy is released by a stoichiometric split off of pyrophosphates from incorporated deoxynucleotide triphosphates and a defined nucleotide dispensation order is given. Because Pyrosequencing works best if single-stranded DNA templates are used, template generation usually requires PCR with a target-specific biotinylated primer and a subsequent purification involving interaction of the biotin label with immobilized streptavidin. To circumvent the need for numerous and expensive template-specific biotinylated primers, we developed a method that uses the ligation of amplified DNA fragments into a plasmid vector, thereby facilitating subsequent PCR using a universal vector-specific biotinylated primer. This approach allows easy and straightforward isolation of single-stranded templates of any PCR product. As a proof of principle, we used the method for genotyping two single-nucleotide polymorphisms in the human genes CARD15 and A2M and for characterization of four multisite variations in the human DEFB104 gene.

Biotinylation↗
PubMed2006

A short isoform of NOD2/CARD15, NOD2-S, is an endogenous inhibitor of NOD2/receptor-interacting protein kinase 2-induced signaling pathways.

Alterations in splicing patterns of genes contribute to the regulation of gene function by generating endogenous inhibitor or activator molecules. Nucleotide-binding and oligomerization domain (NOD) 2 is an intracellular receptor for bacterial cell wall components and plays an important role in initiating immune responses against cytoinvasive pathogens. NOD2 overexpression sensitizes intestinal epithelial cells toward bacterial cell wall components, activates the proinflammatory transcription factor NF-kappaB, and induces the subsequent release of the chemotactic cytokine IL-8. Here, we have assessed the regulation and function of a transcript isoform of NOD2, NOD2-S, generated by the skipping of the third exon, which encodes for a protein that is truncated within the second caspase recruitment (CARD) domain. NOD2-S is preferentially expressed in the human colon and is up-regulated by the antiinflammatory cytokine IL-10. Overexpression of NOD2-S down-regulates NOD2-induced NF-kappaB activation and IL-8 release. Moreover, NOD2-S also interferes with the maturation and secretion of pro-IL-1beta downstream of NOD2 and its adaptor molecule receptor-interacting protein kinase 2. We provide a molecular basis for these effects, as we show that NOD2-S interacts with both, NOD2 and receptor-interacting protein kinase 2 and inhibits the "nodosome" assembly by interfering with the oligomerization of NOD2. These data unveil another level of complexicity in the regulation of intracellular innate immunity and may have important implications for the molecular understanding of NOD/NALP protein-driven disease pathophysiology.

Cell Line↗
PubMed2006

Prevalence, spectrum, and functional characterization of melanocortin-4 receptor gene mutations in a representative population-based sample and obese adults from Germany.

CONTEXT: Autosomal dominant inheritance of mutations in the melanocortin-4 receptor gene (MC4R) is currently regarded as the most relevant genetic cause for extreme obesity and affects 2-4% of extremely obese individuals. OBJECTIVE: Our objective was to assess the relevance of MC4R mutations in a German population-based sample. DESIGN AND SETTING: We conducted a mutation screen of the MC4R gene by capillary electrophoresis-based single-strand conformation polymorphism analysis and denaturing HPLC. PARTICIPANTS: Subjects included 4068 individuals of a German population-based study group [Kooperative Gesundheitsforschung im Raum Augsburg, Survey 4 (KORA-S4); i.e. Cooperative Health Research in the Region of Augsburg] and 1003 German obese adults (body mass index >or= 30 kg/m(2)). MAIN OUTCOME MEASURES: Samples with aberrant capillary electrophoresis-based single-strand conformation polymorphism analysis/denaturing HPLC patterns were resequenced. Functional studies including agonistic receptor stimulation (Nle-D-Phe-alpha-, alpha-, and beta-MSH) and cell surface expression assays were performed. RESULTS: Sixteen (six novel) coding nonsynonymous mutations were detected in 27 heterozygous individuals of KORA-S4. Four of the mutation alleles led to impaired receptor function in vitro; however, none of these six heterozygous mutation carriers was obese (body mass index >or= 30 kg/m(2)). In the obese adults, six coding nonsynonymous and a nonsense mutation were detected in 13 individuals. Only the nonsense mutation allele entailed impaired receptor function. CONCLUSIONS: Our study depicts prevalence, spectrum, and functional characterization of MC4R mutations in the German population-based sample KORA-S4. In this epidemiological study group, individuals heterozygous for nonsynonymous MC4R mutation alleles entailing impaired function were not obese. Furthermore, nonsynonymous MC4R mutations causing impaired receptor function were rare in German obese adults (two in 1003 = 0.2%).

Adult↗
PubMed2006

DNA sequence and analysis of human chromosome 8.

The International Human Genome Sequencing Consortium (IHGSC) recently completed a sequence of the human genome. As part of this project, we have focused on chromosome 8. Although some chromosomes exhibit extreme characteristics in terms of length, gene content, repeat content and fraction segmentally duplicated, chromosome 8 is distinctly typical in character, being very close to the genome median in each of these aspects. This work describes a finished sequence and gene catalogue for the chromosome, which represents just over 5% of the euchromatic human genome. A unique feature of the chromosome is a vast region of approximately 15 megabases on distal 8p that appears to have a strikingly high mutation rate, which has accelerated in the hominids relative to other sequenced mammals. This fast-evolving region contains a number of genes related to innate immunity and the nervous system, including loci that appear to be under positive selection--these include the major defensin (DEF) gene cluster and MCPH1, a gene that may have contributed to the evolution of expanded brain size in the great apes. The data from chromosome 8 should allow a better understanding of both normal and disease biology and genome evolution.

Animals↗
PubMed2006

Should the draft chimpanzee sequence be finished?

Owing to the availability of genome working drafts (WDs), current comparative-sequence studies are frequently performed on a genome-wide scale. In this article, we appraise the utility of WD sequences in the detection of genomic differences in closely related species. We compared human DNA sequences with draft and high-quality versions of the corresponding chimpanzee loci to reveal the overall high quality of the chimp WD sequence. Nevertheless, a significant proportion of the differences between WD and high-quality sequences we observed can be attributed to sequencing errors in the draft. Although we suggest methods to reduce the number of such false positives efficiently, our study emphasizes the benefit expected from finishing the chimpanzee genome sequence.

Animals↗
PubMed2006

SNPSplicer: systematic analysis of SNP-dependent splicing in genotyped cDNAs.

Functional annotation of SNPs (as generated by HapMap (http://www.hapmap.org) for instance) is a major challenge. SNPs that lead to single amino acid substitutions, stop codons, or frameshift mutations can be readily interpreted, but these represent only a fraction of known SNPs. Many SNPs are located in sequences of splicing relevance-the canonical splice site consensus sequences, exonic and intronic splice enhancers or silencers (exonic splice enhancer [ESE], intronic splice enhancer [ISE], exonic splicing silencer [ESS], and intronic splicing silencer [ISS]), and others. We propose using sets of matching DNA and complementary DNA (cDNA) as a screening method to investigate the potential splice effects of SNPs in RT-PCR experiments with tissue material from genotyped sources. We have developed a software solution (SNPSplicer; http://www.ikmb.uni-kiel.de/snpsplicer) that aids in the rapid interpretation of such screening experiments. The utility of the approach is illustrated for SNPs affecting the donor splice sites (rs2076530:A>G, rs3816989:G>A) leading to the use of a cryptic splice site and exon skipping, respectively, and an exonic splice enhancer SNP (rs2274987:C/T), leading to inclusion of a new exon. We anticipate that this methodology may help in the functional annotation of SNPs in a more high-throughput fashion.

Alternative Splicing↗
PubMed2006

The analysis of cell division and cell wall synthesis genes reveals mutationally inactivated ftsQ and mraY in a protoplast-type L-form of Escherichia coli.

Cell division and cell wall synthesis are tightly linked cellular processes for bacterial growth. A protoplast-type L-form Escherichia coli, strain LW1655F+, indicated that bacteria can divide without assembling a cell wall. However, the molecular basis of its phenotype remained unknown. To establish a first phenotype-genotype correlation, we analyzed its dcw locus, and other genes involved in division of E. coli. The analysis revealed defective ftsQ and mraY genes, truncated by a nonsense and a frame-shift mutation, respectively. Missense mutations were determined in the ftsA and ftsW products yielding amino-acid replacements at conserved positions. FtsQ and MraY, obviously nonfunctional in the L-form, are essential for cell division and cell wall synthesis, respectively, in all bacteria with a peptidoglycan-based cell wall. LW1655F+ is able to survive their loss-of-functions. This points to compensatory mechanisms for cell division in the absence of murein sacculus formation. Hence, this L-form represents an interesting model to investigate the plasticity of cell division in E. coli, and to demonstrate how concepts fundamental for bacterial life can be bypassed.

Amino Acid Sequence↗
PubMed2006

Phylogenetically widespread alternative splicing at unusual GYNGYN donors.

BACKGROUND: Splice donor sites have a highly conserved GT or GC dinucleotide and an extended intronic consensus sequence GTRAGT that reflects the sequence complementarity to the U1 snRNA. Here, we focus on unusual donor sites with the motif GYNGYN (Y stands for C or T; N stands for A, C, G, or T). RESULTS: While only one GY functions as a splice donor for the majority of these splice sites in human, we provide computational and experimental evidence that 110 (1.3%) allow alternative splicing at both GY donors. The resulting splice forms differ in only three nucleotides, which results mostly in the insertion/deletion of one amino acid. However, we also report the insertion of a stop codon in four cases. Investigating what distinguishes alternatively from not alternatively spliced GYNGYN donors, we found differences in the binding to U1 snRNA, a strong correlation between U1 snRNA binding strength and the preferred donor, over-represented sequence motifs in the adjacent introns, and a higher conservation of the exonic and intronic flanks between human and mouse. Extending our genome-wide analysis to seven other eukaryotic species, we found alternatively spliced GYNGYN donors in all species from mouse to Caenorhabditis elegans and even in Arabidopsis thaliana. Experimental verification of a conserved GTAGTT donor of the STAT3 gene in human and mouse reveals a remarkably similar ratio of alternatively spliced transcripts in both species. CONCLUSION: In contrast to alternative splicing in general, GYNGYN donors in addition to NAGNAG acceptors enable subtle protein variations.

Alternative Splicing↗
PubMed2005

Single-nucleotide polymorphisms in NAGNAG acceptors are highly predictive for variations of alternative splicing.

Aberrant or modified splicing patterns of genes are causative for many human diseases. Therefore, the identification of genetic variations that cause changes in the splicing pattern of a gene is important. Elsewhere, we described the widespread occurrence of alternative splicing at NAGNAG acceptors. Here, we report a genomewide screen for single-nucleotide polymorphisms (SNPs) that affect such tandem acceptors. From 121 SNPs identified, we extracted 64 SNPs that most likely affect alternative NAGNAG splicing. We demonstrate that the NAGNAG motif is necessary and sufficient for this type of alternative splicing. The evolutionarily young NAGNAG alleles, as determined by the comparison with the chimpanzee genome, exhibit the same biases toward intron phase 1 and single-amino acid insertion/deletions that were already observed for all human NAGNAG acceptors. Since 28% of the NAGNAG SNPs occur in known disease genes, they represent preferable candidates for a more-detailed functional analysis, especially since the splice relevance for some of the coding SNPs is overlooked. Against the background of a general lack of methods for identifying splice-relevant SNPs, the presented approach is highly effective in the prediction of polymorphisms that are causal for variations in alternative splicing.

Alleles↗
PubMed2005

Validation of mRNA/EST-based gene predictions in human Xp11.4 revealed differences to the organization of the orthologous mouse locus.

Careful manual annotation of the human reference sequence provides a solid basis for the identification of disease-associated genes. Toward this end, we focused on a medically relevant 2.6-Mb region of the human chromosome Xp11.4 between markers DXS9851 and DXS9751 and identified 16 transcription units according to the Vertebrate Genome Annotation (Vega) rules. In order to validate these annotations, we performed a comprehensive RT-PCR expression analysis and a human-mouse comparison. This revealed, despite the high overall genomic conservation of the region, remarkable differences of the gene content between human and mouse. Whereas 12 of 16 annotations were confirmed by RT-PCR in human tissues, for only seven genes mouse orthologs could be identified and found to be expressed. This indicates that a comprehensive and experimentally supported annotation effort of the human genome simultaneously highlights regions with striking differences in gene organization to other species and may indicate evolutionary events specific to the human lineage demanding further functional analyses.

Animals↗