Gene-Ontology-based clustering of gene expression data

The expected correlation between genetic co-regulation and affiliation to a common biological process is not necessarily the case when numerical cluster algorithms are applied to gene expression data. GO-Cluster uses the tree structure of the Gene Ontology database as a framework for numerical clustering, and thus allowing a simple visualization of gene expression data at various levels of the ontology tree. AVAILABILITY: The 32-bit Windows application is freely available at http://www.mpibpc.mpg.de/go-cluster/     

PathoPlant: a database on plant-pathogen interactions

Pathogen recognition and signal transduction during plant pathogenesis is essential for the activation of plant defense mechanisms. To facilitate easy access to published data and to permit comparative studies of different pathogen response pathways, a database is indispensable to give a broad overview of the components and reactions so far known. PathoPlant has been developed as a relational database to display relevant components and reactions involved in signal transduction related to plant-pathogen interactions. On the organism level, the tables 'plant', 'pathogen' and 'interaction' are used to describe incompatible interactions between plants and pathogens or diseases. On the molecular level, plant pathogenesis related information is organized in PathoPlant's main tables 'molecule', 'reaction' and 'location'. Signal transduction pathways are modeled as consecutive sequences of known molecules and corresponding reactions. PathoPlant entries are linked to associated internal records as well as to entries in external databases such as SWISS-PROT, GenBank, PubMed, and TRANSFAC. PathoPlant is available as a web-based service at http://www.pathoplant.de.     

Is telomere erosion a mechanism of species extinction?

According to the fossil record, 99.9% of all species that have ever lived on Earth have disappeared. However, only about 4% died out during the five mass extinction events, whereas it seems that the majority of species vanished without any signs of significant earthbound or extraterrestrial physical threats. Clearly, biological extinction mechanisms are by far the most important, but they are subject to serious limitations concerning the worldwide disappearance of species. In view of that, species-inherent mechanisms, which could lead to the worldwide destabilization of a population, might be worth reconsideration. Telomeres, the protective caps of chromosome ends, and the enzyme telomerase have been well preserved in plants and animals during evolution. In the absence of telomerase activity, telomeric DNA has been shown to shorten every time a cell divides. The concept of a mitotic clock based on the gradual erosion of telomeres is now generally accepted and has been confirmed in numerous plants and animals. Chromosomal rearrangements are the hallmarks of two completely different biological phenomena, cancer and speciation. In premalignant cells, gradual telomere erosion beyond a critical threshold is a well-known inducer of chromosomal instability. The species clock hypothesis, as presented here, is based on the idea of a tiny loss of mean telomere length per generation. This mechanism would not rapidly endanger the survival of a particular species. Yet, after many thousands of generations, critically short telomeres could lead to the weakening and even the extinction of old species and would simultaneously create the unstable chromosomal environment that might result in the origination of new species.     

Synthesis of novel 4- and 5-substituted benzyl ether derivatives of vesamicol and in vitro evaluation of their binding properties to the vesicular acetylcholine transporter site

Detection of the central cholinergic deficits, a consistent feature of Alzheimer's disease, is essential to allow preventive measures and/or symptomatic treatment already at a very early stage of the disease. The vesicular acetylcholine transporter (VAChT) represents an appropriate target to establish PET radiotracer that are adequate for brain imaging the loss of cholinergic terminals. Here we describe the synthesis and binding characteristics of novel derivatives of vesamicol, known to represent a specific antagonist of VAChT sites. Novel benzyl ether derivatives of vesamicol either 4- or 5-substituted at the cyclohexylring have been synthesized by different regioselective ring opening reactions of a same epoxide precursor. The affinity and selectivity of the novel compounds to VAChT sites were analyzed by competitive radioligand binding studies in rat brain and liver membrane preparations using tritium labeled radioligands. The 4-substituted fluorobenzylether of vesamicol 10b was shown to exhibit a high affinity to VAChT sites (K(i)-value(10b)=10.7+/-1.7 nM), but demonstrated also binding capacities to sigma receptors (K(i-)value(10b)=18.5+/-6.9 nM, [(3)H]DTG; K(i)-value(10b)=30.6+/-9.6 nM, [(3)H]haloperidol). The data suggest the potential of vesamicol derivatives to design appropriate radiotracer for PET imaging of central cholinergic deficits.     

A novel, high-affinity, fluorescent progesterone receptor antagonist. Synthesis and in vitro studies

The present paper describes the chemical synthesis and in vitro characterization of a novel, high-affinity, fluorescent progesterone receptor (PR) antagonist. The three-step synthesis was carried out starting from mifepristone. After demethylation with calcium oxide, the methylamino group was alkylated with 6-bromohexanol, and the resulting compound was reacted with fluorescein 5-isothiocyanate, yielding the fluorescein-mifepristone conjugate. Interaction of the conjugate as well as of its precursors with PR was determined in cell culture (alkaline phosphatase assay and transactivation assay). Antiprogestagenic activity of the intermediates were comparable to that of the parent compound. Even after attachment of the bulky fluorescein moiety, considerable antiprogestagenic activity was maintained. Microscopic studies revealed that fluorescence of the conjugate was almost confined to the nuclei of steroid hormone receptor-positive cells, whereas the nuclei of steroid hormone receptor-negative cells remained unstained. To our knowledge, this is the first report on a fluorescent ligand for PR suitable for studies in living cells. It is proposed that the present fluorescent PR antagonist might serve as a lead compound for the development of contrast agents for PR imaging, e.g., by near-infrared optical imaging.     

Epidermolysis bullosa simplex-type mutations alter the dynamics of the keratin cytoskeleton and reveal a contribution of actin to the transport of keratin subunits

Dominant keratin mutations cause epidermolysis bullosa simplex by transforming keratin (K) filaments into aggregates. As a first step toward understanding the properties of mutant keratins in vivo, we stably transfected epithelial cells with an enhanced yellow fluorescent protein-tagged K14R125C mutant. K14R125C became localized as aggregates in the cell periphery and incorporated into perinuclear keratin filaments. Unexpectedly, keratin aggregates were in dynamic equilibrium with soluble subunits at a half-life time of <15 min, whereas filaments were extremely static. Therefore, this dominant-negative mutation acts by altering cytoskeletal dynamics and solubility. Unlike previously postulated, the dominance of mutations is limited and strictly depends on the ratio of mutant to wild-type protein. In support, K14R125C-specific RNA interference experiments resulted in a rapid disintegration of aggregates and restored normal filaments. Most importantly, live cell inhibitor studies revealed that the granules are transported from the cell periphery inwards in an actin-, but not microtubule-based manner. The peripheral granule zone may define a region in which keratin precursors are incorporated into existing filaments. Collectively, our data have uncovered the transient nature of keratin aggregates in cells and offer a rationale for the treatment of epidermolysis bullosa simplex by using short interfering RNAs.     

Helifulvanolsäkure—ein neues diterpen mit anomalem kohlenstoffgerüst aus Helichrysum fulvum

The aerial parts of Helichrysum fulvum afforded, in addition to beyerenic acid and ent-kaurenic acid, two new diterpenic acids with the hitherto unknown carbon skeleton of an isotrachylobane type. The structures of these acids, isolated as their methyl esters, were elucidated by extensive NMR studies, some chemical transformations and by X-ray structural analysis of the corresponding acetate. The related alcohol on reaction with pyridinochlorochromate afforded a homoconjugated diene probably formed by fragmentation of a cyclopropyl carbinol. The possible biogenesis of the new carbon skeleton is discussed briefly.     

Deletion of mdmB impairs mitochondrial distribution and morphology in Aspergillus nidulans

Mitochondria form a dynamic network of interconnected tubes in the cells of Saccharomyces cerevisiae or filamentous fungi such as Aspergillus nidulans, Neurospora crassa, or Podospora anserina. The dynamics depends on the separation of mitochondrial fragments, their movement throughout the cell, and their subsequent fusion with the other parts of the organelle. Interestingly, the microtubule network is required for the distribution in N. crassa and S. pombe, while S. cerevisiae and A. nidulans appear to use the actin cytoskeleton. We studied a homologue of S. cerevisiae Mdm10 in A. nidulans, and named it MdmB. The open reading frame is disrupted by two introns, one of which is conserved in mdm10 of P. anserina. The MdmB protein consists of 428 amino acids with a predicted molecular mass of 46.5 kDa. MdmB shares 26% identical amino acids to Mdm10 from S. cerevisiae, 35% to N. crassa, and 32% to the P. anserina homologue. A MdmB-GFP fusion protein co-localized evenly distributed along mitochondria. Extraction of the protein was only possible after treatment with a non-ionic and an ionic detergent (1% Triton X-100; 0.5% SDS) suggesting that MdmB was tightly bound to the mitochondrial membrane fraction. Deletion of the gene in A. nidulans affected mitochondrial morphology and distribution at 20 degrees C but not at 37 degrees C. mdmB deletion cells contained two populations of mitochondria at lower temperature, the normal tubular network plus some giant, non-motile mitochondria.     

Backbone dynamics of the human MIA protein studied by (15)N NMR relaxation: implications for extended interactions of SH3 domains

The melanoma inhibitory activity (MIA) protein is a clinically valuable marker in patients with malignant melanoma as enhanced values diagnose metastatic melanoma stages III and IV. Here, we report the backbone dynamics of human MIA studied by (15)N NMR relaxation experiments. The folded core of human MIA is found to be rigid, but several loops connecting beta-sheets, such as the RT-loop for example, display increased mobility on picosecond to nanosecond time scales. One of the most important dynamic features is the pronounced flexibility of the distal loop, comprising residues Asp 68 to Ala 75, where motions on time scales up to milliseconds occur. Further, significant exchange contributions are observed for residues of the canonical binding site of SH3 domains including the RT-loop, the n-Src loop, for the loop comprising residues 13 to 19, which we refer to as the"disulfide loop", in part for the distal loop, and the carboxyl terminus of human MIA. The functional importance of this dynamic behavior is discussed with respect to the biological activity of several point mutations of human MIA. The results of this study suggest that the MIA protein and the recently identified highly homologous fibrocyte-derived protein (FDP)/MIA-like (MIAL) constitute a new family of secreted proteins that adopt an SH3 domain-like fold in solution with expanded ligand interactions.