Phylogenetic island disequilibrium: evidence for ongoing long‐term population dynamics in two Mediterranean butterflies

Aim Our aims were to verify the existence of phylogenetic disequilibrium between butterfly lineages at the subcontinental scale for islands and the nearest mainland and to test the capacity of islands for hosting ancestral populations of butterflies and the significance of such relict populations. Location The western Mediterranean continental area of Europe and North Africa together with several large and small islands (Balearics, Tuscan Archipelago, Aeolian Archipelago, Capri, Sardinia, Sicily, Corsica). Methods Using geometric morphometrics, the shape of male genitalia was analysed in two common butterflies (Pyronia cecilia and Pyronia tithonus), whose spatial heterogeneity in the Mediterranean region has recently been described. Observed patterns in genital shapes were compared with shapes predicted for islands and fossil islands to assess the contribution of historical and current events in accounting for the transition from a refugial model to an equilibrium model. Measurements were taken for 473 specimens in 90 insular and mainland sites. Results The shape of the genitalia of populations of most islands differed substantially from that predicted by the equilibrium hypothesis while closely fitting the refugial hypothesis. The comparison between different models strongly suggests that islands maintain ancestral lineages similar to those living in Spain (P. cecilia) and France (P. tithonus). A high correlation between observed and predicted patterns on islands and fossil islands occurs during the first steps of modelled introgressive hybridization while the following steps exposed a successively lower fit, suggesting that the process from a refugial to an equilibrium situation is highly skewed towards an earlier non‐equilibrium. Main conclusions The observed non‐equilibrium pattern supports the refugial hypothesis, suggesting that an ancestral lineage was originally distributed from Spain to Italy, and also occupied offshore islands. This lineage, replaced in Italy, has persisted on the islands owing to their isolation. A comparison of the distribution patterns for genetic and morphometric markers in several species indicates that the situation highlighted for Pyronia may represent a common biogeographic feature for many Mediterranean butterflies.     

Glutamine cycling in isolated working rat heart

To what extent does glutamine turnover keep pace with oxidative metabolism in the rat heart? To address this question, the following groups of substrates were presented to the isolated, working rat heart: 1) glucose (5 mM), insulin (40 microU/ml), and [2-13C]acetate (5 mM; high workload, n = 5); 2) pyruvate (2.5 mM) and [2-13C]acetate (5 mM; normal workload, n = 5); or 3) propionate (1 mM) and [2-13C]acetate (2.5 mM; normal workload, n = 3). In a subset of these experiments, the exchange of glutamate and glutamine was quantified by separation with ion exchange chromatography and analysis by GC-MS. There was an apparent equilibration of mass isotopomers of glutamate and glutamine after 50 min of perfusion, although the extent of equilibration was not determined. The fractional enrichment in glutamine was 31% of the enrichment of glutamate with the three different perfusates. From high-resolution nuclear magnetic resonance spectra, we found a ratio of glutamine to glutamate content of 94.1, 53.4, and 96.9%, respectively, for each experimental group. In experiments for which l-[1-13C]glutamine (5 mM) was included in the perfusate of group 2, [1-13C]glutamine was detected in the heart, but transfer of 13C from glutamine to glutamate was not detected (n = 4). We conclude that, in the perfused working heart, production of glutamine by amidation of glutamate takes place and can be detected, whereas the reverse process, generation of glutamate from glutamine, remains undetected.     

DNA polymerase clamp shows little turnover at established replication sites but sequential de novo assembly at adjacent origin clusters

The spatial and temporal organization of DNA replication was investigated in living cells with a green fluorescent protein fusion to the DNA polymerase clamp PCNA. In situ extractions and photobleaching experiments revealed that PCNA, unlike RPA34, shows little if any turnover at replication sites, suggesting that it remains associated with the replication machinery through multiple rounds of Okazaki fragment synthesis. Photobleaching analyses further showed that the transition from earlier to later replicons occurs by disassembly into a nucleoplasmic pool of rapidly diffusing subcomponents and reassembly at newly activated sites. The fact that these replication sites were de novo assembled in close proximity to earlier ones suggests that activation of neighboring origins may occur by a domino effect possibly involving local changes in chromatin structure and accessibility.     

Isolation and purification of bacterial membrane proteins by the use of organic solvents: The lactose permease and the carbodiimide-reactive protein of the adenosinetriphosphatase complex of escherichia coli

Techniques for the solubilization and fractionation of integral membrane proteins have been developed in recent years. A small portion of membrane protein (about 2%, proteolipid fraction) will partition into chloroform or 1-butanol, and, in several cases, these proteins retain functional activity. A virtually complete solubilization can be achieved at neutral pH by use of aprotic solvents, like hexamethylphosphoric triamide or N-methylpyrrolidone. At relatively low concentrations (< 3 M) aprotic solvents inhibited β-D-galactoside transport by whole cells and the derivative membrane vesicles of Escherichia coli, but this inhibition could be largely reversed by a simple washing procedure. At higher concentrations of aprotic solvent (5–6 M), 50–80% of the total protein of lactose transport-positive membrane vesicles was solubilized. When these extracts were added to intact lactose transport-negative membrane vesicles, lactose transport was reconstituted, the required energy being provided by either respiration (e.g., addition of D-lactate) or by a K⁺ diffusion potential established with the aid of valinomycin. The dicyclohexylcarbodiimide (DCCD)-reactive subunit of the E. coli ATPase complex was found to partition into chloroform, and to be amenable to further purification in organic solvent. Ether precipitation and chromatography on DEAE-cellulose and hydroxypropyl-Sephadex G-50 yielded an homogeneous polypeptide of an apparent molecular weight of 9,000. The purified and unlabeled DCCD-reactive protein was incorporated into K⁺-loaded liposomes, and a membrane potential was generated by the addition of valinomycin. There are indications that the DCCD-reactive protein alone made the membrane specifically permeable for protons.     

Fine needle aspiration cytology and flow cytometry in the diagnosis and subclassification of non-Hodgkin's lymphoma based on the World Health Organization classification

OBJECTIVE: To determine the usefulness of fine needle aspiration cytology (FNAC) in combination with flow cytometry on the new World Health Organization (WHO) classification of malignant lymphoma. STUDY DESIGN: Smears and flow cytometry reports of patients who underwent both methods at the same time were independently examined. Both methods were classified according to the new WHO classification of malignant lymphoma. RESULTS: A group of 131 smears were examined. In 89 cases exact diagnosis was made by cytomorphology. Twenty-five cases were not classified exactly or were classified incorrectly, resulting in a sensitivity of 96.4% and a specificity of 85%. With flow cytometry, only 30 of 131 patients could be classified exactly, resulting in a sensitivity of 27% and specificity of 100%, respectively. The combination of methods showed a sensitivity of 85% and specificity of 100%. CONCLUSION: The combination of FNAC and flow cytometry obtained by FNAC can distinguish between benign and malignant lymphoid infiltrates and support a diagnosis of lymphoma.     

Mathematical analysis of multienzyme systems. II. Steady state and transient control.

The control theory of steady states, previously presented for linear enzymatic systems (Heinrich and Rapoport, 1974) is extended to nonlinear systems. On the basis of three theorems a new procedure for the calculation of the control strength and of the control matrix is developed. The theory is applied to the extended model of glycolysis of erythrocytes, which includes also ATP-consuming processes. Also in this model the glycolytic flux is mainly controlled by the hexokinase-phosphofructokinase-system. The control strengths of the pyruvate kinase and of the enzymes of the 2.3 P2G-bypass are negligibly small. The control strength of the ATPase is negative, i.e. an activation of this enzyme leads to a decrease of the flux. For transition states of multienzyme systems definitions are given for the mean time required for the transition of the metabolites and for the "transient control" of enzymes. Enzymes with a pronounced influence on the transition time are called time-limiting enzymes. Enzymes which excert strong control on the time-dependent processes may have little influence under steady state conditions and vice versa. The transition times of ATP have been calculated for transient states of glycolysis.     

Global classification of natural terrestrial ecosystems

Summary A global classification system of natural terrestrial ecosystems (including systematic notation), based on the climate zones of Walter, is presented. The basic units of the system are the ecological units biome and biogeocoene. The zonobiomes, which are climate zones corresponding to the largest vegetation units, are subdivided into subzonobiomes and these into individual biomes. The biomes are thus natural, geographical units within the climate zones. They are in turn subdivided into individual biogeocoenes and their constituent synusiae. In addition, the coordinate concepts of pedobiome and orobiome are introduced. These are distinguished from the zonobiomes as follows:1. the pedobiomes by extreme edaphic conditions which cause azonal vegetation.2. the orobiomes, as mountain ranges, by their vertical climate zonation and the altitudinal belts of vegetation.These relationships are explained, and two subseries of pedo-and oro-subunits are established. Transitional zones (zono-ecotones) between individual zonobiomes are also distinguished. The classification system is summarized in a schematic, and a world map of zonobiomes and zono-ecotones is included. More details are presented in Walter (1976).

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Zusammenfassung Ein globales Gliederungssystem der natürlichen terrestrischen Ökosysteme (einschließlich systematischen Bezeichnungen) wird in Beziehung zu den Walter'schen Klimazonen gesetzt. Grundeinheiten des Systems sind die ökologischen Einheiten Biom und Biogeozön. Die Zonobiome werden unterteilt in Subzonobiome und diese in Biome. Die Zonobiome sind Klimazonen und entsprechen den größten Vegetationseinheiten. Die Biome sind natürliche, geographische Einheiten innerhalb der Klimazonen. Sie werden bis zu einzelnen Biogeozönen und ihren Synusien (Teilsytemen) unterteilt. Parallel dazu werden die Begriffe Pedobiom und Orobiom eingeführt. Diese heben sich aus den Zonobiomen heraus: die Pedobiomen durch extreme Böden, die eine azonale Vegetation bedingen, die Orobiome als Gebirge durch die vertikale Klimagliederung und die Höhenstufen der Vegetation. Diese Beziehungen werden erklärt, und zwei Nebenreihen der Pedo- bzw. Orobiom-Untereinheiten werden aufgestellt. Zwischen den einzelnen Zonobiomen werden Übergangszonen (Zonoökotone) unterschieden. Das Gliederungssytem wird bereits in einem Schema zusammengefaßt, und eine Weltkarte der Zonobiome und Zonoökotone wird beigefügt. Ausführlich werden alle diese Fragen bei WALTER (1976) behandelt.

     

Novel reversible methionine aminopeptidase-2 (MetAP-2) inhibitors based on purine and related bicyclic templates

The natural product fumagillin 1 and derivatives like TNP-470 2 or beloranib 3 bind to methionine aminopeptidase 2 (MetAP-2) irreversibly. This enzyme is critical for protein maturation and plays a key role in angiogenesis. In this paper we describe the synthesis, MetAP-2 binding affinity and structural analysis of reversible MetAP-2 inhibitors. Optimization of enzymatic activity of screening hit 10 (IC₅₀: 1 μM) led to the most potent compound 27 (IC₅₀: 0.038 μM), with a concomitant improvement in LLE from 2.1 to 4.2. Structural analysis of these MetAP-2 inhibitors revealed an unprecedented conformation of the His339 side-chain imidazole ring being co-planar sandwiched between the imidazole of His331 and the aryl-ether moiety, which is bound to the purine scaffold. Systematic alteration and reduction of H-bonding capability of this metal binding moiety induced an unexpected 180° flip for the triazolo[1,5-a]pyrimdine bicyclic template.