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At least 19 recordsLinked to original sources
PubMed1996

Clusters of DNA damage induced by ionizing radiation: formation of short DNA fragments. II. Experimental detection.

The basic 30-nm chromatin fiber in the mammalian cell consists of an unknown (possibly helical) arrangement of nucleosomes, with about 1.2 kb of DNA per 10-nm length of fiber. Track-structure considerations suggest that interactions of single delta rays or high-LET particles with the chromatin fiber might result in the formation of multiple lesions spread over a few kilobases of DNA (see the accompanying paper: W.R. Holley and A. Chatterjee, Radiat. Res. 145, 188-199, 1996). In particular, multiple DNA double-strand breaks and single-strand breaks may form. To test this experimentally, primary human fibroblasts were labeled with [3H]thymidine and exposed at 0 degrees C to X rays or accelerated nitrogen or iron ions in the LET range of 97-440 keV/microns. DNA was isolated inside agarose plugs and subjected to agarose gel electrophoresis under conditions that allowed good separation of 0.1-2 kb size DNA. The bulk of DNA remained in the well or migrated only a small distance into the gel. It was found that DNA fragments in the expected size range were formed linearly with dose with an efficiency that increased with LET. A comparison of the yield of such fragments with the yield of total DNA double-strand breaks suggests that for the high-LET ions a substantial proportion (20-90%) of DNA double-strand breaks are accompanied within 0.1-2 kb by at least one additional DNA double-strand break. It is shown that these results are in good agreement with theoretical calculations based on treating the 30-nm chromatin fiber as the target for ionizing particles. Theoretical considerations also predict that the clusters will contain numerous single-strand breaks and base damages. It is proposed that such clusters be designated "regionally multiply damaged sites." Postirradiation incubation at 37 degrees C resulted in a decline in the number of short DNA fragments, suggesting a repair activity. The biological significance of regionally multiply damaged sites is presently unknown.

Cells, Cultured↗
PubMed1996

A mutation in protein phosphatase 2A regulatory subunit A affects auxin transport in Arabidopsis.

The phytohormone auxin controls processes such as cell elongation, root hair development and root branching. Tropisms, growth curvatures triggered by gravity, light and touch, are also auxin-mediated responses. Auxin is synthesized in the shoot apex and transported through the stem, but the molecular mechanism of auxin transport is not well understood. Naphthylphthalamic acid (NPA) and other inhibitors of auxin transport block tropic curvature responses and inhibit root and shoot elongation. We have isolated a novel Arabidopsis thaliana mutant designated roots curl in NPA (rcn1). Mutant seedlings exhibit altered responses to NPA in root curling and hypocotyl elongation. Auxin efflux in mutant seedlings displays increased sensitivity to NPA. The rcn1 mutation was transferred-DNA (T-DNA) tagged and sequences flanking the T-DNA insert were cloned. Analysis of the RCN1 cDNA reveals that the T-DNA insertion disrupts a gene for the regulatory A subunit of protein phosphatase 2A (PP2A-A). The RCN1 gene rescues the rcn1 mutant phenotype and also complements the temperature-sensitive phenotype of the Saccharomyces cerevisiae PP2A-A mutation, tpd3-1. These data implicate protein phosphatase 2A in the regulation of auxin transport in Arabidopsis.

Amino Acid Sequence↗
PubMed1993

Structure and function of the homeotic gene complex (HOM-C) in the beetle, Tribolium castaneum.

The powerful combination of genetic, developmental and molecular approaches possible with the fruit fly, Drosophila melanogaster, has led to a profound understanding of the genetic control of early developmental events. However, Drosophila is a highly specialized long germ insect, and the mechanisms controlling its early development may not be typical of insects or Arthropods in general. The beetle, Tribolium castaneum, offers a similar opportunity to integrate high resolution genetic analysis with the developmental/molecular approaches currently used in other organisms. Early results document significant differences between insect orders in the functions of genes responsible for establishing developmental commitments.

Animals↗
PubMed1997

The effect of space flight on monoclonal antibody synthesis in a hybridoma mouse cell line.

The hybridoma cell line, 3G10G5, producing a monoclonal antibody to the major capsid protein VP1 from the avian polyomavirus budgerigar fledgling disease virus, was produced from a Balb/C mouse. This cell line was used to test the effects of microgravity on cellular processes, specifically protein synthesis. A time course study utilizing incorporation of [35S]methionine into newly synthesized monoclonal antibody was performed on STS-77. After 5.5 days, it was observed that cell counts for the samples exposed to microgravity were lower than those of ground-based samples. However, radiolabel incorporation of the synthesized monoclonal antibody was similar in both orbiter and ground control samples. Overall, microgravity does not seem to have an effect on this cell line's ability to synthesize IgG protein.

Animals↗
PubMed1995

Oligonucleotides as probes for studying polymerization reactions in dilute aqueous solution: II. Polycondensations.

We have prepared a [32P]-labeled oligonucleotide probe carrying a ureido (-NH-CO-NH2) function at its 3'-terminus. This labeled oligomer was used to study polycondensations of urea and formaldehyde and of various phenols and formaldehyde in aqueous solution. The formation of formaldehyde copolymers attached to the amido-function of the probe was monitored by gel electrophoresis. Our results are generally in agreement with those obtained using conventional techniques. Our method is suitable for monitoring potentially prebiotic polycondensation reactions involving formaldehyde.

Electrophoresis↗
PubMed1993

Prebiotic syntheses of vitamin coenzymes: I. Cysteamine and 2-mercaptoethanesulfonic acid (coenzyme M).

The reaction of NH3 and SO3(2-) with ethylene sulfide is shown to be a prebiotic synthesis of cysteamine and 2-mercaptoethanesulfonic acid (coenzyme M). A similar reaction with ethylene imine would give cysteamine and taurine. Ethylene oxide would react with NH3 and N(CH3)3 to give the phospholipid components ethanolamine and choline. The prebiotic sources of ethylene sulfide, ethylene imine and ethylene oxide are discussed. Cysteamine itself is not a suitable thioester for metabolic processes because of acyl transfer to the amino group, but this can be prevented by using an amide of cysteamine. The use of cysteamine in coenzyme A may have been due to its prebiotic abundance. The facile prebiotic synthesis of both cysteamine and coenzyme M suggests that they were involved in very early metabolic pathways.

Aziridines↗
PubMed1993

Prebiotic syntheses of vitamin coenzymes: II. Pantoic acid, pantothenic acid, and the composition of coenzyme A.

Pantoic acid can by synthesized in good prebiotic yield from isobutyraldehyde or alpha-ketoisovaleric acid + H2CO + HCN. Isobutyraldehyde is the Strecker precursor to valine and alpha-ketoisovaleric acid is the valine transamination product. Mg2+ and Ca2+ as well as several transition metals are catalysts for the alpha-ketoisovaleric acid reaction. Pantothenic acid is produced from pantoyl lactone (easily formed from pantoic acid) and the relatively high concentrations of beta-alanine that would be formed on drying prebiotic amino acid mixtures. There is no selectivity for this reaction over glycine, alanine, or gamma-amino butyric acid. The components of coenzyme A are discussed in terms of ease of prebiotic formation and stability and are shown to be plausible choices, but many other compounds are possible. The gamma-OH of pantoic acid needs to be capped to prevent decomposition of pantothenic acid. These results suggest that coenzyme A function was important in the earliest metabolic pathways and that the coenzyme A precursor contained most of the components of the present coenzyme.

4-Butyrolactone↗
PubMed1991

A deficiency of the homeotic complex of the beetle Tribolium.

In Drosophila, the establishment of regional commitments along most of the anterior/posterior axis of the developing embryo depends on two clusters of homeotic genes: the Antennapedia complex (ANT-C) and the bithorax complex (BX-C). The red flour beetle has a single complex (HOM-C) representing the homologues of the ANT-C and BX-C in juxtaposition. Beetles trans-heterozygous for two particular HOM-C mutations spontaneously generate a large deficiency, presumably by an exchange within the common region of two overlapping inversions. Genetic and molecular results indicate that this deficiency spans at least the interval between the Deformed and abdominal-A homologues. In deficiency homozygous embryos, all gnathal, thoracic and abdominal segments develop antennal appendages, suggesting that a gene(s) has been deleted that acts to distinguish trunk from head. There is no evidence that beetles have a homologue of the segmentation gene fushi tarazu of similar genomic location and function. On the basis of the genetic tractability, convenient genome size and organization of Tribolium, and its relatively long phylogenetic divergence from Drosophila (>300 million years), we have integrated developmental genetic and molecular analyses of the HOM-C. We isolated about 70 mutations in the complex representing at least six complementation groups. The homeotic phenotypes of adults and lethal embryos lead us to believe that these beetle genes are homologous with the Drosophila genes indicated in Fig. 1 (see text).

Animals↗
PubMed1995

A possible prebiotic synthesis of pantetheine, a precursor to coenzyme A.

The involvement of coenzyme A in many enzyme reactions suggests that it acted in this capacity very early in the development of life on Earth. Particularly relevant in this regard is its role in the activation of amino acids and hydroxy acids in the biosynthesis of some peptide antibiotics--a mechanism of peptide synthesis that forms the basis for the proposal that a thioester world could have preceded the RNA world. The components of coenzyme A have been shown to be probable prebiotic compounds: beta-alanine, pantoyl lactone and cysteamine and possibly adenosine. We show here that the pantetheine moiety of coenzyme A (which also occurs in a number of enzymes) can be synthesized in yields of several per cent by heating pantoyl lactone, beta-alanine and cysteamine at temperatures as low as 40 degrees C. These components are extremely soluble and so would have been preferentially concentrated in evaporating bodies of water, for example on beaches and at lagoon margins. Our results show that amide bonds can be formed at temperatures as low as 40 degrees C, and provide circumstantial support for the suggestion that pantetheine and coenzyme A were important in the earliest metabolic systems.

4-Butyrolactone↗
PubMed1996

Synthesis of long prebiotic oligomers on mineral surfaces.

Most theories of the origin of biological organization assume that polymers with lengths in the range of 30-60 monomers are needed to make a genetic system viable. But it has not proved possible to synthesize plausibly prebiotic polymers this long by condensation in aqueous solution, because hydrolysis competes with polymerization. The potential of mineral surfaces to facilitate prebiotic polymerization was pointed out long ago. Here we describe a system that models prebiotic polymerization by the oligomerization of activated monomers--both nucleotides and amino acids. We find that whereas the reactions in solution produce only short oligomers (the longest typically being a 10-mer), the presence of mineral surfaces (montmorillonite for nucleotides, illite and hydroxylapatite for amino acids) induces the formation of oligomers up to 55 monomers long. These are formed by successive 'feedings' with the monomers; polymerization takes place on the mineral surfaces in a manner akin to solid-phase synthesis of biopolymers.

Amino Acids↗
PubMed1993

Unexpected substrate specificity of T4 DNA ligase revealed by in vitro selection.

We have used in vitro selection techniques to characterize DNA sequences that are ligated efficiently by T4 DNA ligase. We find that the ensemble of selected sequences ligates about 50 times as efficiently as the random mixture of sequences used as the input for selection. Surprisingly many of the selected sequences failed to produce a match at or close to the ligation junction. None of the 20 selected oligomers that we sequenced produced a match two bases upstream from the ligation junction.

Base Sequence↗
PubMed1997

A simple procedure for constructing 5'-amino-terminated oligodeoxynucleotides in aqueous solution.

A rapid method for the synthesis of oligodeoxynucleotides (ODNs) terminated by 5'-amino-5'-deoxythymidine is described. A 3'-phosphorylated ODN (the donor) is incubated in aqueous solution with 5'-amino- 5'-deoxythymidine in the presence of N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), extending the donor by one residue via a phosphoramidate bond. Template- directed ligation of the extended donor and an acceptor ODN, followed by acid hydrolysis, yields the acceptor ODN extended by a single 5'-amino-5'-deoxythymidine residue at its 5'terminus.

Base Sequence↗
PubMed1996

Diminished forearm vasomotor response to central hypervolemic loading in aerobically fit individuals.

The aim of this study was to test the hypothesis that cardiopulmonary baroreflex control of forearm vascular resistance (FVR) during central hypervolemic loading was less sensitive in exercise trained high fit individuals (HF) compared to untrained average fit individuals (AF). Eight AF (age: 24 +/- 1 yr and weight: 78.9 +/- 1.7 kg) and eight HF (22 +/- 1 yr 79.5 +/- 2.4 kg) voluntarily participated in the investigation. Maximal aerobic power (determined on a treadmill), plasma volume and blood volume (Evans blue dilution method) were significantly greater in the HF than AF (60.8 +/- 0.7 vs. 41.2 +/- 1.9 ml.kg-1.min-1, 3.96 +/- 0.17 vs 3.36 +/- 0.08 1, and 6.33 +/- 0.23 vs 5.28 +/- 0.13 1). Baseline heart rate (HR), central venous pressure (CVP), mean arterial pressure (MAP, measured by an intraradial catheter or a Finapres finger cuff), forearm blood flow (FBF, plethysmography), and FVR, calculated from the ratio (MAP-CVP)/FBF, were not different between the HF and the AF. Lower body negative pressure (LBNP, -5, -10, -15, and -20 torr) and passive leg elevation (LE, 50 cm) combined with lower body positive pressure (LBPP, +5, +10, and +20 torr) were utilized to elicit central hypovolemia and hypervolemia, respectively. Range of CVP (from LBNP to LE+LBPP) was similar in the AF (from -3.9 to +1.9 mm Hg) and HF (from -4.0 to +2.2 mm Hg). However, FVR/CVP was significantly less in the HF (-1.8 +/- 0.1 unit.mm Hg-1) than AF (-34 +/- 0.1 unit.mm Hg-1). The FVR decrease in response to increase in CVP was significantly diminished in the HF (-1.46 +/- 0.45 unit.mm Hg-1) compared to the AF (-4.40 +/- 0.97 unit.mm Hg-1), and during LBNP induced unloading the FVR/CVP of the HF (-2.01 +/- 0.49 unit.mm Hg-1) was less (P < 0.08) than the AF (-3.28 +/- 0.69 unit.mm Hg-1). We concluded that the cardiopulmonary baroreceptor mediated FVR reflex response was significantly less sensitive to changes in CVP in individuals who practice exercise training.

Adult↗
PubMed1996

Extraterrestrial helium trapped in fullerenes in the Sudbury impact structure.

Fullerenes (C60 and C70) in the Sudbury impact structure contain trapped helium with a 3He/4He ratio of 5.5 x 10(-4) to 5.9 x 10(-4). The 3He/4He ratio exceeds the accepted solar wind value by 20 to 30 percent and is higher by an order of magnitude than the maximum reported mantle value. Terrestrial nuclear reactions or cosmic-ray bombardment are not sufficient to generate such a high ratio. The 3He/4He ratios in the Sudbury fullerenes are similar to those found in meteorites and in some interplanetary dust particles. The implication is that the helium within the C60 molecules at Sudbury is of extraterrestrial origin.

Carbon↗