Mapping and Use of a Sequence that Targets DNA Ligase I to Sites of DNA Replication In Vivo

The mammalian nucleus is highly organized, and nuclear processes such as DNA replication occur in discrete nuclear foci, a phenomenon often termed “functional organization” of the nucleus. We describe the identification and characterization of a bipartite targeting sequence (amino acids 1–28 and 111–179) that is necessary and sufficient to direct DNA ligase I to nuclear replication foci during S phase. This targeting sequence is located within the regulatory, NH₂-terminal domain of the protein and is dispensable for enzyme activity in vitro but is required in vivo. The targeting domain functions position independently at either the NH₂ or the COOH termini of heterologous proteins.

We used the targeting sequence of DNA ligase I to visualize replication foci in vivo. Chimeric proteins with DNA ligase I and the green fluorescent protein localized at replication foci in living mammalian cells and thus show that these subnuclear functional domains, previously observed in fixed cells, exist in vivo. The characteristic redistribution of these chimeric proteins makes them unique markers for cell cycle studies to directly monitor entry into S phase in living cells.

     

Measurement of gap junctional communication by fluorescence activated cell sorting

Summary Cell-to-cell communication via gap junctions has played a fundamental role in the orderly development of multicellular organisms. Current methods for measuring this function apply mostly to homotypic cell populations. The newly introduced Fluorescence Activated Cell Sorting (FACS) method, albeit with some limitations, is simple, reliable, and quantitative in measuring the dye transfer via gap junctions in both homotypic and heterotypic cell populations. In the homotypic setting, the result in dye transfer from the FACS method is comparable to the scrape-loading and microinjection methods. Using this FACS method, we observed a decline of cell-to-cell communication in transformed and cancer cells. We also observed a differential degree of communication between two heterotypic cell populations depending on the direction of dye transfer.     

Initial mass of avian eggs: Comparison between measured and calculated values

Summary It had previously been demonstrated that by weighing an egg after replacing the air cell with water the initial egg mass at the time of laying is obtained during any stage of incubation. The average egg mass ofSchönwetter (1960–84) compared with those obtained by weighing fresh eggs of the same species after replacement of the air cell with water gives an excellent agreement, as shown in the regression curve, with a slope of 1.0.

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Zusammenfassung Das Wägen eines Vogeleies, dessen Luftkammer mit Wasser gefüllt worden ist, gibt sein Frischgewicht zum Zeitpunkt des Legens wieder. So kann auch nachträglich auf einfache Weise das Frischgewicht eines Vogeleies bestimmt werden. Die durchschnittlichen Frischvollgewichte vonSchönwetter (1960–84) stimmen auffallend gut mit unseren Wägungen überein, wie die berechnete Regressionsgerade (Steigung=1.0) beweist. Letztere stützt sich einerseits auf 97 an 1164 Eiern ermittelten Durchschnittsgewichte von 68 Arten (9 Ordnungen) der verschiedensten Faunengebiete und anderseits auf 7616Schönwetters Berechnungen zugrundeliegende Eiern derselben Arten.

     

Partial characterization of cotton plants expressing two toxin proteins from Bacillus thuringiensis: relative toxin contribution, toxin interaction, and resistance management

Abstract:  Laboratory studies were performed to characterize the lepidopteran toxicity of cotton plants expressing two different toxin proteins from Bacillus thuringiensis (Bt), in order to assess insect resistance management implications of a commercial, two-toxin transgenic cotton. An independent and additive interactive effect of two Bt δ -endotoxins expressed by the transgenic cotton variety 15985 was demonstrated by examining the responses of Heliothis virescens (F.), Helicoverpa zea (Boddie), and Spodoptera frugiperda (J.E. Smith) larvae to field- or greenhouse-grown tissue from genetic near-isolines, which expressed Cry1A only, Cry2Ab only, or both toxins. In all cases, the Cry2Ab component was the larger contributor to total toxicity in the two-toxin isoline. Toxin-specific, quantitative enzyme-linked immunosorbent assay (ELISA) tests confirmed that the levels of each toxin in tissues of the two-toxin isoline were not statistically different (P > 0.05) from the levels found in the corresponding tissues of the respective single-toxin isoline. Resistance management considerations were discussed. Considering the additive interaction of toxins, a relatively simple insect resistance-monitoring procedure was proposed for the monitoring of commercial cotton varieties expressing both toxins.     

Characterization of plasmids in a sucrose-fermenting strain of Escherichia coli.

A multiply drug-resistant strain of Escherichia coli isolated from a patient in Bangladesh was shown to carry four types of plasmids based on size differences. One type carries a gene or genes for sucrose fermentation.     

Thiazolidinedione response in familial lipodystrophy patients with LMNA mutations: a case series

Type 2 familial partial lipodystrophy (FPLD2) patients show impaired glucose and lipid metabolism resulting from lipodystrophic 'lipid pressure' and an intrinsic defect in skeletal muscle metabolism. Since mutated lamin A may interfere with peroxisome proliferator activator gamma (PPARγ) expression, we hypothesized that PPARγ stimulation improves fat distribution and metabolic abnormalities in these patients. 5 nondiabetic FPLD2 patients were treated with rosiglitazone over 12 months. We assessed body composition, body fat distribution, and skinfold thickness/subcutaneous tissue thickness. We also determined venous glucose, insulin, and free fatty acid (FFA) concentrations, and respiratory quotient (RQ) before and during oral glucose tolerance testing. Adipose tissue and muscle fasting and postprandial metabolism were studied by microdialysis. Within 12 months treatment, hip circumference increased from 93.6±2.78 cm to 96.2±2.3 cm (p<0.05). Rosiglitazone reduced fasting glucose levels and liver transaminases. Baseline and postprandial FFA concentrations were significantly lower after 12 months treatment. RQ and muscle interstitial pyruvate and lactate did not respond to treatment. We conclude that PPARγ stimulation with rosiglitazone modestly improves glucose metabolism in FPLD2 patients presumably through proximal adipose tissue expansion. The intrinsic muscular metabolic defect does not respond to rosiglitazone.     

Colonization of the satellite cell niche by skeletal muscle progenitor cells depends on notch signals

Skeletal muscle growth and regeneration rely on myogenic progenitor and satellite cells, the stem cells of postnatal muscle. Elimination of Notch signals during mouse development results in premature differentiation of myogenic progenitors and formation of very small muscle groups. Here we show that this drastic effect is rescued by mutation of the muscle differentiation factor MyoD. However, rescued myogenic progenitors do not assume?a satellite cell position and contribute poorly to myofiber growth. The disrupted homing is due to a deficit in basal lamina assembly around emerging satellite cells and to their impaired adhesion to myofibers. On a molecular level, emerging satellite cells deregulate the expression of basal lamina components and adhesion molecules like integrin α7, collagen XVIIIα1, Megf10, and Mcam. We conclude that Notch signals control homing of satellite cells, stimulating them to?contribute to their own microenvironment and to adhere to myofibers.