Early development of the kelp fly,Coelopa frigida (Diptera). II. Morphology of cleavage and blastoderm formation

Cleavage and blastoderm formation in Coelopa frigida are extremely rapid developmental processes. In short (6–7 minutes) successive cell cycles, nuclei multiply and spread out through the egg. The movement seems to be aided by endoplasmic vesicles and cisternae which are in direct contact with the nuclear membrane. The first cells to separate from the egg plasmodium in early superficial cleavage stages are the pole cells. Precursor material from multivesicular bodies forms the pole cell membranes. The primary nuclei from the posterior pole region are removed from the blastoderm by the pole cell segregation. Blastoderm nuclei from the regions adjacent to the posterior pole migrate into the residual periplasm after pole cell segregation has been completed and constitute the blastoderm nuclei in that region of the egg. Nucleoli are not revealed during internal cleavage. They appear in pole cells shortly after their segregation. The generation time of the blastoderm nuclei increases after the twelfth cleavage. Concurrently, nucleoli form in the blastoderm nuclei and permanent cell membranes separate individual blastoderm cells. After blastoderm cells have been separated from each other, they remain in contact with the interior yolk sac by means of cytoplasmic canals. This contact is maintained at least during the early phases of blastokinesis. Observations on nuclear migration and rapid membrane formation are discussed as examples of protein assembly from subunits as an alternative to de novo protein synthesis in early stages of development.     

Lymphocyte function in patients with anti-D(Rh0) antibodies

The behaviour of humoral and cellular immunity as well as that of cells bearing Fc receptors was investigated in persons with anti-D antibodies. Peripheral lymphocytes could be identified in 6 of 26 examined test persons particularly sensitized against the stroma of Rh-positive erythrocytes. There was no relation to the anti-D titre. The parameters of humoral immunity showed no correlation to the anti-D titre. In 37 per cent of the persons with anti-D antibodies, which were produced "naturally" or artificially, an increased content of IgE could be proved in the serum. In each case, anti-D formation apparently leads to an increase of cells bearing Fc receptors, which can be recognized by the increased number of EA rosettes. A relation to the anti-D titre did not exist. None of the immunological test methods used is suitable for predicting the success of an artificial immunization for the purpose of gaining high titre anti-D antibodies.     

Wy-48,252 (1,1,1-trifluoro-N-[3-(2-quinolinylmethoxy)phenyl]methane sulfonamide) an orally active leukotriene D4 antagonist: pharmacological characterization in vitro and in vivo in the guinea pig

The following communicates the pharmacology of Wy-48,252 (1,1,1-trifluoro-N-[3-(2-quinolinylmethoxy)phenyl]methanesulfonamide) a chemically novel and orally potent leukotriene (LT) D4 receptor antagonist. In the isolated guinea-pig trachea pretreated with indomethacin (5 microM) and L-cysteine (10 mM), Wy-48,252 antagonized TD4-induced contraction with a pKB = 7.6. Against LTC4 on tissues pretreated with IND and glutathione (10 mM), Wy-48,252 had a pKB greater than 5. Wy-48,252 (10 microM) did not antagonize pilocarpine-, histamine- or PGF2 alpha-induced tracheal contraction. Further, in the presence of indomethacin and chlorpheniramine (1 microM), Wy-48,252 dose-dependently inhibited the antigen-induced contraction of guinea-pig trachea in a manner consistent with antagonism at the LTD4 receptor and inhibition of LT synthesis. In the Konzett-Rossler model of i.v. LTD4-induced bronchoconstriction in indomethacin treated guinea pigs, intragastric Wy-48,252 (2 hr) had an ID50 of 100 micrograms/kg and a functional half-life of 5 hr. Against i.v. antigen-induced bronchoconstriction in guinea pigs treated with indomethacin and chlorpheniramine, intragastric Wy-48,252 (2 hr) had an ID50 of 0.6 mg/kg and a 5 hr half life. Intragastric Wy-48,252 also selectively blocked the cutaneous wheal reaction to intradermal LTD4 but not histamine. We conclude that Wy-48,252 is distinguished from other selective LTD4 receptor antagonists by its oral potency and should be useful in ascertaining the role of LTD4 mediated processes in asthma, allergy and animal models.     

Sphingosine-1-phosphate: A Janus-faced mediator of fibrotic diseases

Sphingosine-1-phosphate (S1P) is a pleiotropic lipid mediator that acts either on G protein-coupled S1P receptors on the cell surface or via intracellular target sites. In addition to the well established effects of S1P in angiogenesis, carcinogenesis and immunity, evidence is now continuously accumulating which demonstrates that S1P is an important regulator of fibrosis. The contribution of S1P to fibrosis is of a Janus-faced nature as S1P exhibits both pro- and anti-fibrotic effects depending on its site of action. Extracellular S1P promotes fibrotic processes in a S1P receptor-dependent manner, whereas intracellular S1P has an opposite effect and dampens a fibrotic reaction by yet unidentified mechanisms. Fibrosis is a result of chronic irritation by various factors and is defined by an excess production of extracellular matrix leading to tissue scarring and organ dysfunction. In this review, we highlight the general effects of extracellular and intracellular S1P on the multistep cascade of pathological fibrogenesis including tissue injury, inflammation and the action of pro-fibrotic cytokines that stimulate ECM production and deposition. In a second part we summarize the current knowledge about the involvement of S1P signaling in the development of organ fibrosis of the lung, kidney, liver, heart and skin. Altogether, it is becoming clear that targeting the sphingosine kinase-1/S1P signaling pathway offers therapeutic potential in the treatment of various fibrotic processes. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.     

Development of joint application strategies for two microbial gene finders

MOTIVATION: As a starting point in annotation of bacterial genomes, gene finding programs are used for the prediction of functional elements in the DNA sequence. Due to the faster pace and increasing number of genome projects currently underway, it is becoming especially important to have performant methods for this task. RESULTS: This study describes the development of joint application strategies that combine the strengths of two microbial gene finders to improve the overall gene finding performance. Critica is very specific in the detection of similarity-supported genes as it uses a comparative sequence analysis-based approach. Glimmer employs a very sophisticated model of genomic sequence properties and is sensitive also in the detection of organism-specific genes. Based on a data set of 113 microbial genome sequences, we optimized a combined application approach using different parameters with relevance to the gene finding problem. This results in a significant improvement in specificity while there is similarity in sensitivity to Glimmer. The improvement is especially pronounced for GC rich genomes. The method is currently being applied for the annotation of several microbial genomes. AVAILABILITY: The methods described have been implemented within the gene prediction component of the GenDB genome annotation system.     

Comparing expression level-dependent features in codon usage with protein abundance: an analysis of 'predictive proteomics'

Synonymous codon usage is a commonly used means for estimating gene expression levels of Escherichia coli genes and has also been used for predicting highly expressed genes for a number of prokaryotic genomes. By comparison of expression level-dependent features in codon usage with protein abundance data from two proteome studies of exponentially growing E. coli and Bacillus subtilis cells, we try to evaluate whether the implicit assumption of this approach can be confirmed with experimental data. Log-odds ratio scores are used to model differences in codon usage between highly expressed genes and genomic average. Using these, the strength and significance of expression level-dependent features in codon usage were determined for the genes of the Escherichia coli, Bacillus subtilis and Haemophilus influenzae genomes. The comparison of codon usage features with protein abundance data confirmed a relationship between these to be present, although exceptions to this, possibly related to functional context, were found. For species with expression level-dependent features in their codon usage, the applied methodology could be used to improve in silico simulations of the outcome of two-dimensional gel electrophoretic experiments.     

PathFinder: reconstruction and dynamic visualization of metabolic pathways

MOTIVATION: Beyond methods for a gene-wise annotation and analysis of sequenced genomes new automated methods for functional analysis on a higher level are needed. The identification of realized metabolic pathways provides valuable information on gene expression and regulation. Detection of incomplete pathways helps to improve a constantly evolving genome annotation or discover alternative biochemical pathways. To utilize automated genome analysis on the level of metabolic pathways new methods for the dynamic representation and visualization of pathways are needed. RESULTS: PathFinder is a tool for the dynamic visualization of metabolic pathways based on annotation data. Pathways are represented as directed acyclic graphs, graph layout algorithms accomplish the dynamic drawing and visualization of the metabolic maps. A more detailed analysis of the input data on the level of biochemical pathways helps to identify genes and detect improper parts of annotations. As an Relational Database Management System (RDBMS) based internet application PathFinder reads a list of EC-numbers or a given annotation in EMBL- or Genbank-format and dynamically generates pathway graphs.     

Whole genome shotgun sequencing guided by bioinformatics pipelines--an optimized approach for an established technique

While the sequencing of bacterial genomes has become a routine procedure at major sequencing centers, there are still a number of genome projects at small- or medium-size facilities. For these facilities a maximum of control over sequencing, assembling and finishing is essential. At the same time, facilities have to be able to co-operate at minimum costs for the overall project. We have established a pipeline for the distributed sequencing of Alcanivorax borkumensis SK2, Azoarcus sp. BH72, Clavibacter michiganensis subsp. michiganensis NCPPB382, Sorangium cellulosum So ce56 and Xanthomonas campestris pv. vesicatoria 85-10. Our pipeline relies on standard tools (e.g. PHRED/PHRAP, CAP3 and Consed/Autofinish) wherever possible, supplementing them with new tools (BioMake and BACCardI) to achieve the aims described above.     

Construction and validation of a Sinorhizobium meliloti whole genome DNA microarray: genome-wide profiling of osmoadaptive gene expression

Based on the complete Sinorhizobium meliloti genome sequence we established DNA microarrays as a comprehensive tool for systematic genome-wide gene expression analysis in S. meliloti 1021. For these PCR fragment-based microarrays, called Sm6kPCR, a collection of probes for the 6207 predicted protein-coding genes consisting of 6046 gene-specific PCR fragments and 161 70 mer oligonucleotides was arrayed in high density on glass slides. To obtain these PCR fragments primer pairs were designed to amplify internal gene-specific DNA fragments of 80-350 bp. Additionally, these primers were characterized by a 5' extension that allowed for reamplification using standard primers after the first amplification employing the specific primers. In order to ascertain the quality of the Sm6kPCR microarrays and to validate gene expression studies in S. meliloti parallel hybridizations based on RNA samples obtained from cells cultured under identical conditions were performed. In addition, gene expression in S. meliloti in response to an osmotic upshift imposed by the addition of 0.38 M NaCl was monitored. 137 genes were identified showing significant changes in gene expression resulting from the osmotic upshift. From these genes 52 were induced and 85 genes were repressed. Among the genes displaying different RNA levels some functional groups could be identified that are particularly remarkable. Repression was observed for 8 genes related to motility and chemotaxis, 7 genes encoding amino acid biosynthesis enzymes and 15 genes involved in iron uptake whereas 14 genes involved in transport of small molecules and 4 genes related to polysaccharide biosynthesis were induced.