bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparative study by circular dichroism of the conformation of deazapurine nucleosides and that of common purine nucleosides.
Authors: Miles D W , Townsend L B , Redington P , Eyring H
Published: May 2006

Purine nucleoside analogs modified by replacement of the nitrogen atom at the 3 position by a CH group give a characteristic circular dichroism curve that is not substantially modified by chemical substitution at the 8 position. Since it is rather well established that 8-substituted purine nucleosides are predominantly in the syn conformation in aqueous solution, it follows that the 3-deazapurine nucleosides, whether substituted at position 8 or not, also favor the syn conformation. These data are in sharp contrast to the circular dichroism data obtained on 8-halogenated and 8-alkylated derivatives of adenosine and guanosine, which give circular dichroism profiles substantially different from those obtained on the parent compounds. Certain purine-nucleoside-utilizing enzymes fail to interact effectively with either the unsubstituted 3-deaza analogs or the 8-substituted derivatives of adenosine and guanosine. The hypothesis recently given that the inactivity of the 8-substituted derivatives springs from their syn-conformational preference is tentatively accepted to explain the inactivity of the 3-deaza analogs.
Electrophoretic distributions of human peripheral blood mononuclear white cells from normal subjects and from patients with acute lymphocytic leukemia.
Authors: Smith B A , Ware B R , Weiner R S
Published: May 2006

The electrophoretic mobilities of mononuclear white cells from the peripheral blood of normal subjects and leukemic patients were measured. Analysis of both fresh and cryopreserved samples of each type showed that cryopreservation had no distinguishable effect on the electrophoretic distribution of either normal or leukemic cells. Mode mobilities (mobilities at which maximum intensities are observed) of cells from nine leukemic patients were 7-28% below the average mode mobility of normal cells. The standard deviation of the electrophoretic mobilities of normal cells was 2%. Normal cells had an asymmetric, often bimodal, electrophoretic distribution; leukemic cells consistently had a single symmetric distribution.
Lateral transport on cell membranes: mobility of concanavalin A receptors on myoblasts.
Authors: Schlessinger J , Koppel D E , Axelrod D , Jacobson K , Webb W W , Elson E L
Published: May 2006

We report measurements of the lateral mobility of fluorescent labeled concanavalin A receptor complexes on the plasma membrane of cultured myoblasts of rat. Transport rates were measured by observing the recovery of fluorescence in a small region of the cell surface initially photobleached irreversibly by an intense, focused laser light pulse. Under different conditions we measured effective diffusion coefficients of the receptor complexes in the range 8 x 10(-12) less than D less than 3 x 10(-11) cm2/sec which is two orders of magnitude lower than we found for a fluorescent lipid probe, D approximately (8 +/- 3) x 10(-9) cm2/sec. This large difference and the presence of apparently immobile concanavalin A receptors suggests that factors beyond the fluoidity of the phospholipid bilayer membrane matrix control the rate of lateral transport of the complexes. Effective mobilities of the complexes decrease with increases in the valence, dose, and occupation time of the lectin on the membrane. These properties imply an aggregation of the lectin-receptor complexes. Mobilities are not influenced by azide, colchicine or preincubation at low temperature. Cytochalasin B and low temperatures, during the time of measurement, decrease the lateral transport rate.
Ontogeny of mouse B lymphocytes and inactivation by antigen of early B lymphocytes.
Authors: Bruyns C , Urbain-Vansanten G , Planard C , Vos-Cloetens C , Urbain J
Published: May 2006

Taking advantage of recent findings about membrane fluidity, we have studied and compared the biosynthetic capacities of fetal or neonatal mouse B (bone-marrow derived) lymphocytes (until 10 days after birth) and adult B lymphocytes. Although both early and adult lymphocytes can synthesize surface immunoglobulins, they have a different physiological behavior after interaction with a ligand (anti-immunoglobulin sera or antigen), either in vivo or in vitro. Fetal and neonatal lymphocytes bearing surface immunoglobulins do not reexpress their membrane receptors after capping and endocytosis promoted by anti-immunoglobulin sera. On the other hand, adult lymphocytes resynthesize completely their receptors after the same treatment. Furthermore, intrafetal injections of hemocyanin in pregnant mice lead to a striking decrease in the number of hemocyanin-binding cells. It seems plausible that this non-reexpression of surface immunoglobulins could be the first step in tolerance establishment.
Simian virus to (SV40) tumor-specific proteins in nucleus and plasma membrane of HeLa cells infected by adenovirus 2-SV40 hybrid virus Ad2+ND2.
Authors: Deppert W , Walter G
Published: May 2006

HeLa cells infected with the nondefective adenovirus 2-SV40 hybrid virus (Ad2+ND2) have previously been shown to synthesize two SV40-specific proteins with molecular weights of 56,000 (56 K protein) and 42,000 (42 K protein). The present study demonstrates that these proteins are immunoprecipitable with serum from hamsters bearing SV40 tumors. By peptide analysis, it is shown that the 56 K and 42 K proteins share common amino acid sequences. Both proteins are metabolically stable in nuclei and plasma membranes but unstable in the cytoplasm, as shown by pulse-chase experiments and cell fractionation studies.
Evidence for regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity and cholesterol synthesis in nonhepatic tissues of rat.
Authors: Balasubramaniam S , Goldstein J L , Faust J R , Brown M S
Published: May 2006

The adenine analogue 4-aminopyrazolopyrimidine has been reported previously to reduce the hepatic secretion of plasma lipoproteins in rats, thereby lowering the plasma cholesterol level. In the current studies, reduction of the plasma cholesterol level by 90% in rats through the administration of aminopyrazolopyrimidine was found to be associated with a 5- to 30-fold increase in the activity of 3-hydroxy-3-methylglutaryl-coenzyme A reductase [mevalonate:NADP+ oxidoreductase (CoA-acylating), EC1.1.1.34] in kidney and lung. In both tissues, the enhanced activity of this microsomal enzyme was associated with a 3-fold elevation in the rate of cholesterol synthesis from either [14C]acetate or [14C]octanoate. Comparable increases were not observed in the activities of several other microsomal enzymes or in the rates of [14C]acetate incorporation into saponifiable lipids or CO2. When administration of 4-aminopyrazolopyrimidine was terminated, plasma cholesterol levels rose and 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity declined in the kidney in a reciprocal manner. These data are consistent with the hypothesis that the low levels of 3-hydroxy-3-methylglutaryl-coenzyme A reductase activity and cholesterol synthesis that are normally observed in certain nonhepatic tissues of the rat are due to an active form of feedback regulation mediated by cholesterol carried in plasma lipoproteins.
Decreased fidelity of DNA polymerase activity isolated from aging human fibroblasts.
Authors: Linn S , Kairis M , Holliday R
Published: May 2006

DNA polymerase (deoxynucleosidetriphosphate: DNA nucleotidyltransferase, EC 2.7.7.7 or DNA nucleotidyltransferase) activity, isolated from late and early passage cells of the diploid human fibroblast line, MRC-5, was compared. The level of activity dropped with increasing passage. In addition, when the fidelity of polymerization was monitored with four synthetic templates under a variety of conditions, it was observed that the enzyme from late passage cells was more error-prone. The possible relation of these observations to "senescence" of the fibroblasts is discussed.
Cytoarchitectonic mapping by microdensitometry.
Authors: Hudspeth A J , Ruark J E , Kelly J P
Published: May 2006

Functional differences among various portions of the cerebral cortex are often correlated with differing cortical layering patterns. Convenient, accurate techniques for scoring layering should therefore prove useful in electrophysiological as well as anatomical investigations. We report the application of a computer-controlled scanning microdensitometer as a means of rapid measurement of optical densities in histological sections of monkey visual cortices, areas 17 and 18. The technique readily permits recognition of the previously defined cortical layers and suggests that still finer consistent layering patterns exist; it provides objective "fingerprints" of cortical regions which facilitate comparisons of structure from area to area and from animal to animal. The procedure should serve also to score the positions of autoradiographic grains, degenerating axonal terminals, and other labeled structures, and to allow the comparison of preparations stained by various techniques.
Interferon-induced inhibition of protein synthesis in L-cell extracts: an ATP-dependent step in the activation of an inhibitor by double-stranded RNA.
Authors: Roberts W K , Clemens M J , Kerr I M
Published: May 2006

The translation of encephalomyocarditis virion RNA in extracts from interferon-treated L-cells is inhibited by the addition of double-stranded RNA (dsRNA) at 400 ng/ml. A similar inhibition in response to dsRNA is seen in control cell extracts supplemented with small amounts of a postribosomal supernatant fraction from interferon-treated cells (interferon cell sap): Neither interferon cell sap nor dsRNA alone is inhibitory in control systems. The inhibition is much reduced if translation is carried out at low ATP concentrations.Conversely, the inhibitory capacity of the interferon cell sap is increased 100-fold if it is preincubated with dsRNA and ATP prior to its addition to the protein-synthesizing system. After this preincubation all detectable dsRNA can be removed without any diminution of the inhibitory activity of the cell sap. These results are compatible with a two-step model for the inhibition in which a pre-inhibitor is activated by dsRNA, the activated inhibitor then interacting with the protein synthesis system to inhibit translation.
Control of the amplification convertase of complement by the plasma protein beta1H.
Authors: Weiler J M , Daha M R , Austen K F , Fearon D T
Published: May 2006

An inhibitory activity for an erythrocyte in termediate bearing the properdin (P)-stabilized amplification C3 convertase, PC3bBb, was recognized in whole normal human serum and separated from C3b inactivator by its distinct physicochemical and functional characteristics. The inhibitory activity was found to reside in a protein that was purified to homogeneity and elicited a monospecific antibody in a rabbit. This protein was identified as beta1H and found to have a serum concentration of 516 +/- 89 mug/ml (mean +/- 1 SD). beta1H produced a dose related, first-order loss of convertase function and release of 125I-Bb from the P-stabilized intermediate, indicating a mechanism of action by decay-dissociation of Bb from the complex, PC3bBb. beta1H exhibited only a limited capacity to accelerate decay of C3bBb sites stabilized with C3 nephritic factor or to release 125I-Bb from such sites. Amplification of C3 cleavage by C3bBb may well determine whether initial complement activation by the classical or alternative activating sequence is beneficial or detrimental to the host. Regulation of this amplifying function is now recognized to occur at at least three steps: intrinsic decay which reflects the inherent lability of the C3bBb convertase; extrinsic decay-dissociation of Bb which is mediated by the effect of beta1H; and inactivation of exposed C3b by C3b inactivator. The stabilization of C3bBb by activated properdin minimizes intrinsic decay and protects C3b in the bimolecular complex from C3b inactivator. beta1H restores control of the system by decay-dissociation of the bimolecular complex, therby exposing C3b to C3b inactivator whose irreversible action prevents regeneration of the convertase at that site.
Proofreading of the codon-anticodon interaction on ribosomes.
Authors: Thompson R C , Stone P J
Published: May 2006

The fidelity of protein synthesis is substantially greater than the specificity of codon-anticodon recognition that would be expected from the known energetics of base-pairing in solution. To test the suggestion that the specificity of recognition may be increased by "kinetic proofreading" associated with GTP hydrolysis [J. J. Hopfield (1974) Proc. Natl. Acad. Sci. USA 71, 4135-4139], we have studied the interaction of ternary complexes of polypeptide elongation factor Tu, aminoacyl-tRNA, and GTP with poly(U)-programed ribosomes. With most noncognate ternary complexes, including two that pair correctly with the 5' and 3' bases of UUU, rejection occurred without GTP hydrolysis, presumably by the reverse of the initial binding reaction. However, with complexes containing Leu- or Ile-tRNAs, which may pair correctly with the 3' and middle bases, GTP hydrolysis was stimulated though the aa-tRNA was not retained on the ribosome. These results demonstrate the existence of a GTP-dependent proofreading step in aminoacyl-tRNA recognition on ribosomes. They also suggest that the 5' base of the codon is more prone than the middle base to errors that can be corrected by proofreading.
Infectious DNA of spleen necrosis virus is integrated at a single site in the DNA of chronically infected chicken fibroblasts.
Authors: Battula N , Temin H M
Published: May 2006

The infectious DNAs of a number of avian leukosis-sarcoma and reticuloendotheliosis viruses were digested with six nucleotide-specific restriction endonucleases, and the digests were tested for infectivity. All of the enzymes inactivated the viral infectivities except for EcoRI, which did not inactivate the infectivity of the DNA of two of the reticuloendotheliosis viruses, spleen necrosis and chick syncytial viruses. The infectious DNA of spleen necrosis virus after digestion with EcoRI had a buoyant density in CsCl solution greater than the density of the high-molecular-weight infectious viral DNA. The infectious EcoRI-digested spleen necrosis virus DNA from chronically infected chicken cells was uniform in size, 10 megadaltons, which indicated a single site of integration. The infectious EcoRI-digested spleen necrosis virus DNA from acutely infected cells was heterogeneous in size, ranging from 8-14 megadaltons, which indicated multiple sites of integration. These results are consistent with the hypothesis that cells that integrate infectious spleen necrosis virus DNA at a single site survive and multiply, whereas cells that integrate infectious viral DNA at additional sites either die or selectively lose or inactivate the DNA in the additional sites.
Nonsense suppressors of yeast cause osmotic-sensitive growth.
Authors: Singh A
Published: May 2006

Many nonsense suppressors of Saccharomyces cerevisiae cause growth inhibition on hypertonic media. Eight tyrosine-inserting UAA (ochre) suppressors, eight tyrosine-inserting UAG (amber) suppressors, a leucine-inserting UAG suppressor, and a serine-inserting recessive lethal UAG suppressor cause osmotic sensitivity, whereas a serine-inserting UAA suppressor does not cause sensitivity. Although the mechanism is not understood, the growth inhibition of specific suppressors on hypertonic media is correlated with their efficiencies of suppression. This heretofore unknown property of nonsense suppressors is useful for mitotic mapping, selecting tRNA mutants, selecting antisuppressors, and scoring nonsense suppressors.
The humoral immune response of NIH Swiss and SWR/J mice to vaccination with formalinized AKR or Gross murine leukemia virus.
Authors: Lee J C , Ihle J N , Huebner R
Published: May 2006

The humoral immune responses of NIH Swiss and SWR/J mice immunized with formalin-inactivated AKR or Gross murine leukemia virus, respectively, were examined. Both immune sera had high titers of antibodies detectable in radioimmune precipitation assays using [3H]leucine-labeled AKR virus and in radioimmunoassays using purified virion components. The predominant antibody titers were directed against gp71 and pl15(E). The immune response against gp71 was predominantly type-specific, whereas the reactivity with pl15(E) was predominantly group-specific. A weak immune response against p15 was also detected. Both sera were cytotoxic against cells replicating the AKR-Gross virus type but not against cells replicating Friend murine leukemia virus. This cytotoxicity could be specifically blocked with purified gp71 of AKR murine leukemia virus. Sera from immune NIH Swiss mice neutralized AKR virus, but did not neutralize Rauscher, Scripps, or wild mouse leukemia virus.
Reduction of blood pressure and vascular collagen in hypertensive rats by beta-aminopropionitrile.
Authors: Iwatsuki K , Cardinale G J , Spector S , Udenfriend S
Published: May 2006

beta-Aminopropionitrile, a specific inhibitor of lysyl oxidase prevented the rise in blood pressure induced by deoxycorticosterone-salt in rats. In addition, after the onset of hypertension, administration of beta-aminopropionitrile lowered the blood pressure. Concomitant with the lowering of blood pressure, there was a reduction in the more highly crosslinked form of vascular collagen. These findings would indicate that increases in vascular connective tissue are not only sequelae of hypertension, but may also contribute to the maintenance of elevated blood pressure.
Transient induction of a nuclear antigen unrelated to Epstein-Barr nuclear antigen in cells of two human B-lymphoma lines converted by Epstein-Barr virus.
Authors: Fresen K O , zur Hausen H
Published: May 2006

Infection of cells of the Epstein-Barr virus (EBV)-negative human B-lymphoma lines BJAB and Ramos with EBV preparations from P3HR-1 or B 95-8 cells converted these cells to EBV genome carriers expressing Epstein-Barr nuclear antigen (EBNA) in almost 100% of these cells. Induction of these cells as well as of clones from P3HR-1 EBV-converted BJAB cells with iododeoxyuridine, aminopterin, and hypoxanthine resulted in the appearance of a nuclear antigen in about 1-6% of the cells 1-4 days after induction. The antigen is different from known EBV-induced antigens like EBNA, viral capsid antigen (VCA) or the D- and R-subspecificities of the early antigen (EA) complex. It is demonstrated by indirect immunofluorescence and inactivated after acetone fixation. The antigen was not detectable after induction of uninfected BJAB and Ramos cells nor has it been found in noninduced or induced P3HR-1 and Raji cells. Thus, it appears that EBV-infection mediates the expression of this antigen, for which the name TINA (transiently induced nuclear antigen) is suggested. Sera reacting against TINA generally contained high antibody titers against EBV-induced EA. Only a limited number of highly EA-reactive sera, however, were also positive for TINA. Among 200 sera tested thus far, TINA reactivity was most frequently observed in sera of patients with nasopharyngeal carcinoma (7 out of 28), in sera of the only two patients with immunoblastoma tested and occasionally in sera from patients with Hodgkin's disease and chronic lymphatic leukemia. Among 70 sera from nontumor patients, TINA reactivity was observed three times: two patients suffered from "chronic" infectious mononucleosis, the other revealed persistent splenomegaly.
Inhibition of erythrocyte sickling in vitro by DL-glyceraldehyde.
Authors: Nigen A M , Manning J M
Published: May 2006

Concentrations of DL-glyceraldehyde between 5 and 20 mM reduce the sickling of S/S erythrocytes even in the complete absence of oxygen; at 10 mM glyceraldehyde the increase in the number of normal cells ranges from 20 to 40%. The inhibition of sickling was both concentration- and time-dependent and was not reversed by repeated washings with buffer. Incubation of erythrocytes with increasing concentrations of glyceraldehyde resulted in only a small increase in the oxygen affinity, a moderate reduction in the Hill coefficient, a substantial increase in the minimum gelling concentration, and modification of up to two lysine residues per hemoglobin molecule.
Comparative subunit structure of HeLa, yeast, and chicken erythrocyte chromatin.
Authors: Lohr D , Corden J , Tatchell K , Kovacic R T , Van Holde K E
Published: May 2006

We have compared the chromatin subunit structure of yeast, HeLa, and chicken erythrocyte by analyzing the DNA fragments produced by in situ digestion with staphylococcal nuclease (EC 3.1.4.7) and DNase I(EC 3.1.4.5). The repeat size of the chromatin varies among (and within two of) the three organisms but the size and the structure of the most nuclease-resistant "core" of the repeat is the same. Thus, the interspecies differences in repeat size are due to different lengths of nuclease-sensitive "spacer" DNA between the cores. There also seems to be a difference in the manner of spacing of cores; the transcriptionally active (yeast and HeLa) chromatins have spacings of variable length while the transcriptionally inactive (chicken erythrocyte) has a more regular spacing of cores.
Effect of galactose oxidase, with and without prior sialidase treatment, on the viability of erythrocytes in circulation.
Authors: Bell W C , Levy G N , Williams R , Aminoff D
Published: May 2006

Previous studies have shown that sialidase-treated mammalian erythrocytes were rapidly eliminated from circulation. In contrast, chicken asialoerythrocytes remained fully viable. This investigation was undertaken to ascertain the reason for this difference in behavior as well as to determine the extent of the similarity of the physiological mechanism for the elimination from circulation of asialoglycoproteins and mammalian asialoerythrocytes. To that end, erythrocytes from dogs, rabbits, and chickens were each subjected to the action of galactose oxidase (D-galactose:oxygen 6-oxidoreductase; EC 1.1.3.9) both before and after sialidase (acylneuraminyl hydrolase; EC 3.2.1.18) treatment. The viability of the autologously transfused erythrocytes in circulation was monitored by Na2-51CrO4 labeling. Galactose oxidase had no deleterious effect on the viability of dog or chicken erythrocytes, nor did it restore the viability of dog or rabbit asialoerythrocytes. On the other hand, desialated chicken erythrocytes, which were fully viable, were rendered nonviable upon treatment with galactose oxidase. It may be concluded therefore that (a) the physiological mechanism of elimination of mammalian asialoerythrocytes from circulation is not the same as that for plasma asialoglycoproteins and (b) the treatment of chicken asialoerythrocytes with galactose oxidase results in the oxidation at carbons 6 of the galactosyl- or N-acetylgalactosaminyl residues, thereby rendering the erythrocytes nonviable.
Nucleotide sequences near the origin of replication of bacteriophage f1.
Authors: Ravetch J V , Horiuchi K , Zinder N D
Published: May 2006

The nucleotide sequence of a region related to the initiation of the reaction in which single-stranded DNA gives rise to the replicative form (complementary strand synthesis) in bacteriophage f1 has been determined. The sequence can be drawn in an extensively base-paired structure, i.e., a single hairpin-helix 55 bases long.
Semisynthetic cytochrome c.
Authors: Barstow L E , Young R S , Yakali E , Sharp J J , O'Brien J C , Berman P W , Harbury H A
Published: May 2006

Horse heart cytochrome c can be split with cyanogen bromide into a heme peptide (residues 1-65) and a nonheme peptide (residues 66-104). In a process involving (i) complex formation between the two fragments and (ii) restoration of the severed peptide linkage, a fully active cytochrome c preparation can be re-formed. Use has been made of this process to couple the heme peptide to peptide 66-104 synthesized by the Merrifield solid-phase procedure. The semisynthetic product formed in this manner is indistinguishable from reconstituted cytochrome c prepared with nonsynthetic peptide 66-104.
Structure of native
Renilla reinformis
luciferin
Authors: Hori Kazuo , Charbonneau Harry , Hart Russell C. , Cormier Milton J.
Published: May 2006

The structure of native luciferin from the bioluminescent coelenterate
Renilla reniformis
is shown to be 3,7-dihydro-2-(
p
-hydroxybenzyl)-6-(
p
-hydroxyphenyl)-8-benzylimidazo[1,2-
a
]pyrazin-3-one by mass spectral analysis of synthetic luciferin and the luciferin derived from a protein directly involved in the bioluminescent system. A previous report of the molecular weight of luciferin is shown to be incorrect by reexamination of the spectral data and by synthesis of two derivatives. Detailed analysis of kinetic, emission, and quantum yield data for the isolated and synthetic luciferins confirms this structure. Confirmation of this structure in a number of species from different phyla suggests a common substrate for a variety of bioluminescent marine organisms.
New pathway for the metabolism of pentitols
Authors: London Jack , Chace Nina M.
Published: May 2006

Certain strains of
Lactobacillus casei
can grow at the expense of one or more pentitols. These microorganisms possess a pentitol phosphate pathway that appears to be analogous to the hexitol phosphate pathway found in many facultatively anaerobic bacteria. Pentitol is transported into the cell by a phospho
enol
pyruvate phosphotransferase system that converts it to pentitol phosphate, whereupon a specific dehydrogenase oxidizes the intermediate product to ketopentose phosphate. The ketopentose phosphate is subsequently converted to xylulose-5-
P
and enters one of the pathways of central metabolism.
Gene-sized DNA molecules of the Oxytricha macronucleus have the same terminal sequence.
Authors: Lawn R M
Published: May 2006

The DNA in the macronucleus of the ciliated protozoan Oxytricha exists as small linear molecules with a number average size of about 3000 base pairs. Most, and possibly all, of these DNA molecules contain the same inverted terminal repeat sequence of approximately 26 base pairs. In addition to its terminal location, two inverted copies of this same sequence surround single-strand interruptions within these DNA molecules. This sequence arrangement may function in the processing of these molecules from large chromosomal precursors or in the subsequent replication of these small linear DNAs during cell reproduction.
DNA sequence organization in the water mold
Achlya
Authors: Hudspeth M. E. S. , Timberlake William E. , Goldberg Robert B.
Published: May 2006

Experiments are described that characterize the organization of DNA sequences in the water mold
Achlya bisexualis
. These experiments demonstrate that repetitive and single copy sequences in the
Achlya
genome are arranged in a long-period interspersion pattern. Estimates of the spacing intervals between repetitive and single copy DNA indicate, however, that the interspersion pattern in
Achlya
is longer than has been previously reported in other eukaryotes. These data and measurements of structural gene expression in
Achlya
[Timberlake, W.E., Shumard, D. S. & Goldberg, R. B. (1977)
Cell
10, 623-632] make it difficult to propose a regulatory function for repeated DNA in this eukaryote.
Explant cultures of catecholamine-containing neurons from rat brain: Biochemical, histofluorescence, and electron microscopic studies
Authors: Schlumpf M. , Shoemaker W. J. , Bloom F. E.
Published: May 2006

Norepinephrine (NE)-producing cells of the nucleus locus ceruleus and dopamine (DA)-producing cells of the substantia nigra were dissected microscopically from embryonic rat brain, explanted, and maintained in culture for up to 5 weeks. The cultured neurons of both brain regions showed normal maturation of axons and dendrites and formed ultrastructurally defined synaptic contacts. Fluorescence microscopy of cultured neurons from both brain regions showed typical
in situ
cytological features: long axonal processes with multiple varicosities for locus ceruleus cultures, and smooth, wispy nonvaricose processes in the substantia nigra cultures. All cultures processed for fluorescence microscopy contained specific catecholamine-fluorescent cells. By radioenzyme assay for catecholamines, more than half of the locus ceruleus cultures contained measurable (>10 pg) quantities of NE and DA, but, unlike results on intact brains, DA content exceeded NE content. Cultures of substantia nigra neurons retained no NE and very little DA. Media from substantia nigra and locus ceruleus cultures contained substantial quantities of DA. Addition of reserpine (10 μM) to the medium depleted locus ceruleus neurons of both amines.

The long survival time in culture of locus ceruleus cells, the normal appearance of fluorescent cell bodies and processes, the apparent development of morphologically specialized interneuronal connections, and the ability to synthesize and store NE make these cultures ideally suited for neurophysiological recording as well as morphological, biochemical, and pharmacological experiments.
Norepinephrine induces glial-specific enzyme activity in cultured plasma glioma cells.
Authors: McMorris F A
Published: May 2006

Addition of 1 micronM 1-norepinephrine to cultures of C6TK- rat glioma cells caused a 2-fold increase in specific activity of the glial-specific enzyme 2':3'-cyclic nucleotide 3'-phosphohydrolase (nucleoside-2':3'-cyclic-phosphate 3'-nucleotidohydrolase, EC 3.1.4.16). Specific activity could also be stimulated by analogues of 3':5'-cyclic AMP, and the effect of norepinephrine could be blocked by beta-adrenergic receptor antagonists but not by alpha-adrenergic antagonists. Norepinephrine or cyclic AMP analogues also increased the specific activity of this enzyme in other clones of glioma and Schwannoma cells and in glioma X neuroblastoma cell hybrids. These results show that the stimulatory effect of norepinephrine on cyclic AMP concentrations in glioma cells leads ultimately to a stimulation of glial-specific cell funtion.
In vivo and in vitro phosphorylation of rat liver fructose-1,6-bisphosphatase.
Authors: Riou J P , Claus T H , Flockhart D A , Corbin J D , Pilkis S J
Published: May 2006

Incorporation of 32P from [gamma-32P]ATP into a homogenous preparation of rat hepatic fructose-1,6-bisphosphatase (D-fructose-1,6-bisphosphatase 1-phosphohydrolase, EC 3.1.3.11) was catalyzed by a homogeneous preparation of the catalytic subunit of cyclic AMP-dependent protein kinase from bovine liver. Approximately 4 mol of phosphate were incorporated per mol of the tetrameric enzyme. This phosphorylation was associated with an increase in enzyme activity. In addition, in vivo phosphorylation of the enzyme was observed after injection of radioactive inorganic phosphate into rats and subsequent isolation of the enzyme by conventional purification methods and by immunoprecipitation. All of the labeled phosphate incorporation into the enzyme, both in vitro and in vivo, was precipitated by antibody specific for the enzyme. Furthermore, the 32Pi counts were coincident with the enzyme subunit band when the immunoprecipitates were examined by sodium dodecyl sulfate/disc gel electrophoresis. Acid hydrolysis of the immunoprecipitated enzyme that was phosphorylated in vitro revealed that only seryl residues were labeled. On the basis of the concentration of protein kinase (0.2-1.0 muM) necessary to phosphorylate physiological amounts of fructose-1,6-bisphosphatase (1.0-4.0 muM), it is suggested that cyclic AMP-dependent protein kinase may catalyze the phosphorylation of fructose-1,6-bisphosphatase in vivo.
Immunological distinction between acetylcholine receptor and the alpha-bungarotoxin-binding component on sympathetic neurons.
Authors: Patrick J , Stallcup W B
Published: May 2006

Clone PC12 of a rat sympathetic neuron cell line binds alpha-bungarotoxin and exhibits carbamoylcholine-stimulated uptake of sodium ions. Concentrations of alpha-bungarotoxin that saturate the alpha-bungarotoxin-binding site have no effect on agonist-stimulated sodium uptake. Conversely, antibodies against eel acetylcholine receptor block the agonist-induced sodium flux but fail to recognize the alpha-bungarotoxin-binding component. Detergent extracts of the PC12 clone inhibit the ability of antibody to eel acetylcholine receptor to recognize 125I-alpha-bungarotoxin-acetylcholine receptor complexes derived from muscle. These results distinguish between a ganglionic nicotinic acetylcholine receptor and an alpha-bungarotoxin-binding component on these cells, and provide evidence for antigenic similarities between muscle acetylcholine receptor and ganglionic acetylcholine receptor.
2-Aminopurine-induced mutagenesis in T4 bacteriophage: a model relating mutation frequency to 2-aminopurine incorporation in DNA.
Authors: Goodman M F , Hopkins R , Gore W C
Published: May 2006

We measured the in vivo incorporation of 2-aminopurine into DNA of T4 bacteriophage allelic for gene 43 (DNA polymerase), mutator (L56), 43+, and antimutator (L141). The magnitude of incorporation (mol/mol of Thy) was 1/1500 in L56, 1/1600 in 43+, and 1/8900 in L141. The incorporation ratio L56:43+:L141 in vivo was equal to that mediated by the purified DNA polymerases of these allelic phages in vitro. A model for 2-aminopurine-induced A-T in equilibrium G-C transitions is discussed. The model is used to predict the magnitudes of replication errors (C mispairing with a template 2-aminopurine) and incorporation errors (2-aminopurine mispairing with a template C) per round of replication and to investigate the asymmetry in 2-aminopurine-induced transitions favoring the A-T leads to G-C pathway over G-C leads to A-T. We suggest that the fidelity of L56 and L141 DNA polymerases exemplifies one-step and two-step editing, respectively.
Glucocorticoids increase the responsiveness of cells in culture to prostaglandin E
1
Authors: Foster Stephen J. , Perkins John P.
Published: May 2006

The influence of steroid hormones on the response of human astrocytoma cells (1321N1) to prostaglandin E
1
(PGE
1
) has been investigated. Responsiveness to PGE
1
was determined by measuring the conversion of [
3
H]ATP to cyclic [
3
H]AMP in cells prelabeled with [
3
H]adenine. After incubation of the cells with dexamethasone, a marked increase in both the maximal effect (2- to 3-fold) and the potency (5-fold) of PGE
1
was observed. The effect was specific for the action of PGE
1
in that no change in the response of the cells to isoproterenol was observed. The EC
50
for dexamethasone was 0.001 μM and the effect was dependent on the presence of serum. The effect of dexamethasone was first observed after a 30- to 60-min lag and was maximal by 6-8 hr. Preconfluent cultures (3 days after seeding) exhibited optimal responsiveness to glucocorticoids. Both hydrocortisone and corticosterone mimicked the effect of dexamethasone but both were less potent. The action of dexamethasone was blocked by progesterone, testosterone, and 17α-methyltestosterone. Cycloheximide, at a concentration (1.0 μg/ml) that blocked protein synthesis (>90%) in 1321N1 cells, totally prevented the effect of dexamethasone on the response of the cells to PGE
1
. Upon removal of dexamethasone from cells treated for 16 hr, responsiveness to PGE
1
returned to control levels with a half-time of 4 hr. Dexamethasone also was found to increase the response to PGE
1
of a Rous sarcoma virus-transformed human astrocytoma cell line and the WI-38 human fibroblast line. The most obvious interpretation of our findings is that glucocorticoids induce the synthesis of a protein that selectively modifies the sensitivity of adenylate cyclase to PGE
1
.
Cloned segment of Drosophila melanogaster rDNA containing new types of sequence insertion.
Authors: Glover D M
Published: May 2006

A cloned 14.3-kbase segment of Drosophila melanogaster rDNA (Dm207) is described in which only a 4-kbase region is homologous to a cloned 17-kbase rDNA repeating unit, Dm103; this 4-kbase region consists of part of the 28S rRNA gene and most but not all of the adjacent transcribed spacer that normally connects the 18S and 28S genes. The transcribed spacer in Dm207 is interrupted by a 2.2-kbase stretch of DNA that does not contain any 18S gene sequences. At the other end of the 4-kbase homology, the 28S gene is interrupted by an 8.1-kbase stretch of DNA at a position equivalent to the site of the 28S insertion found in the 17-kbase units. The question of whether the 2.2-kbase and 8.1-kbase interrupter segments in Dm207 derive from longer insertions into the transcribed spacer and 28S genes of a very long repeating unit (greater than or equal to 22 kbases) or represent a region of the chromosomal DNA into which a 4-kbase fragment of rDNA has been inserted is discussed.
Definition of subclasses of nucleoplasmic RNA
Authors: Tamm Igor
Published: May 2006

RNA synthesis in isolated HeLa cell nuclei prepared from cells pretreated with 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) is inhibited in a time-dependent manner. After 40-min pretreatment of cells with 60 μM DRB in the presence of actinomycin D (0.04 μg/ml), the rate of RNA synthesis in isolated nuclei, measured by [
3
H]UTP incorporation, is decreased by 63%. The DRB-resistant one-third of heterogeneous nuclear is distributed over the entire size range of heterogeneous nuclear RNA with some enrichment in the 18S range, as was observed earlier by pulse-labeling whole cells. A subclass of nucleoplasmic RNA molecules is defined in the approximate size range 110 to 250 × 10
3
daltons (330-740 nucleotides). By using heparin (2 mg/ml) to block the synthesis of smaller RNA, a peak in the chain-length range 330-740 nucleotides can be clearly resolved on 2.2% polyacrylamide/1% agarose gels in nuclei from control and DRB-treated cells. The synthesis of these molecules is largely (∼90%) resistant to DRB but sensitive to α-amanitin at 1 μg/ml. The
in vitro
synthesis of molecules in the 140-330 residue range is also sensitive to α-amanitin at 1 μg/ml, and it is not at all affected by pretreatment of cells with DRB. Although the synthesis of the RNA in both the 330-740 and the 140-330 residue size ranges appears to be catalyzed by RNA polymerase II, the results with heparin suggest that there may be reinitiation of molecules in the 140-330 size range but not in the 330-740 range
in vitro
. The synthesis of 4.5S RNA (∼100 nucleotides) and 5S RNA (120 nucleotides) is unaffected by pretreatment of cells with DRB and, as previously reported, is catalyzed by RNA polymerase III, with reinitiation occurring
in vitro
. Addition of DRB directly to isolated HeLa cell nuclei
in vitro
has no detectable effect on the overall rate of RNA synthesis.
Reduction of syngeneic tumor growth by an anti-I-J-alloantiserum.
Authors: Greene M I , Dorf M E , Pierres M , Benacerraf B
Published: May 2006

Highly significant suppression of the growth of S1509a and Sa-I syngeneic sarcomas was observed in A/J mice following daily intravenous injections of 2 microliter of anti-I-Jk alloantiserum. This effect persisted as long as the anti-I-Jk serum was administered (day 15), In contrast, a control anti-I-Js serum had no discernible effect on the growth of the S1509a tumor. The inhibitory activity of the anti-I-Jk serum on the growth of the tumor was absorbed specifically by B10-BR spleen cells bearing I-Jk determinants. In other experiments, we established that A/J mice treated with anti-I-Jk serum, according to the protocol described above, are no longer a source of tumor-specific suppressor cells for adoptive transfer into immune tumor-bearing recipient mice. We conclude that anti-I-Jk serum inhibits tumor growth in A/J mice by abolishing tumor-specific suppressor activity.
Three-dimensional structure of an intact human immunoglobulin.
Authors: Silverton E W , Navia M A , Davies D R
Published: May 2006

We have examined the low-resolution structure of a complete human IgG1 using known domain coordinates from crystallographic investigations of immunoglobulin fragment structures. Our results indicate that the Fc portion of this molecule has a structure similar to that of an isolated Fc fragment, with the carbohydrate moiety playing a central role as the principal contact between the CH2 domains. Carbohydrate also forms a large part of the interface between the Fc and Fab regions. The relative orientations of the variable and constant portions of the Fab regions are intermediate between those reported previously, emphasizing the flexibility of the switch region. These data do not support a two-state allosteric model such as has been proposed for antibody effector functions.
Renaturation of complementary single-stranded DNA circles: complete rewinding facilitated by the DNA untwisting enzyme.
Authors: Champoux J J
Published: May 2006

Renaturation of two complementary single-stranded circles should be limited by topological constraints against the rewinding of the DNA helix. If a mixture of complementary single-stranded rings is annealed and then treated with the DNA untwisting enzyme, the DNA circles completely renature as judged by (i) the presence of interlocked rings that sediment at 53 S in alkali, (ii) the buoyant density of the renatured DNA in CsCl gradients containing ethidium bromide, and (iii) the resistance of the product to the single-strand-specific S1 nuclease. Therefore, the DNA untwisting enzyme is able to provide a transient single-strand break that is sufficient to allow the two strands to completely rewind. The possibility that the untwisting enzyme might facilitate the initiation of the process of genetic recombination is discussed.
Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl-paper and hybridization with DNA probes.
Authors: Alwine J C , Kemp D J , Stark G R
Published: May 2006

We describe a technique for transferring electrophoretically separated bands of RNA from an agarose gel to paper strips. The RNA is coupled covalently to diazobenzyloxymethyl groups on the paper. After transfer and appropriate treatment of the paper to destroy remaining diazo groups, specific RNA bands can be detected by hybridization with 32P-labeled DNA probes followed by autoradiography. This procedure allows detection of specific RNA bands with high sensitivity and low background.
Cloning and mapping of the replication origin of Escherichia coli.
Authors: Yasuda S , Hirota Y
Published: May 2006

The replication origin of Escherichia coli has been cloned on a nonreplicating DNA fragment coding for ampicillin resistance. This recombinant DNA, named pSY211, replicates depending on the presence of the replication origin and can be recovered as a closed circular plasmid DNA of 10.7 megadaltons (Mdal). A restriction map has been constructed. EcoRI cleaves pSY211 into two fragments: one is the ampicillin fragment of 4.5 Mdal and the other is a chromosomal fragment of 6 Mdal and contains the origin. The 6 Mdal EcoRI fragment has four BamHI sites, three HindIII sites, and one Xho I site. A mutant of pSY211 has been isolated which is lacking two BamHI fragments of the chromosomal fragment. In recA hosts, pSY211 is lost at a high frequency. In recA+ hosts, pSY211 is integrated into the chromosome due to nucleotide sequence homology between pSY211 and the replication origin of the E. coli chromosome. The integration site has been mapped. We conclude that the replication origin is located at a site between uncA and rbsK, at about 83 min on the genetic map of E. coli.
Genetic dissection of the control of normal differentiation in myeloid leukemic cells
Authors: Lotem Joseph , Sachs Leo
Published: May 2006

Normal myeloid precursors and MGI
+
D
+
myeloid leukemic cells can be induced to differentiate to mature cells by the normal protein inducer MGI. The sequence of differentiation is the induction of C3 and Fc rosettes, C3 and Fc immune phagocytosis (IP), synthesis and secretion of lysozyme, and formation of mature macrophages and granulocytes. Mutant clones of myeloid leukemic cells have been isolated with differences in the time of induction of C3 and Fc rosettes and C3 and Fc IP, in which lysozyme was induced without going through the stage of Fc or C3 IP, and with differences in inducibility by MGI to mature macrophages or granulocytes. Only one out of five MGI
-
D
-
clones gave rise to MGI
+
D
+
mutants. The ability to obtain mutants from this clone was associated with its special chromosome constitution, and these mutants showed a change in their ability for cap formation by concanavalin A. The steroid inducer dexamethasone can induce in MGI
+
D
+
clones differentiation to macrophages but not to granulocytes. Differentiation by steroid inducer in different clones occurred either with or without induction of Fc rosettes and Fc IP, and induction of C3 rosettes was not always associated with induction of C3 IP. The use of mutants that differ in their competence to be induced by MGI or steroid inducer has shown that there are separate controls for the induction of C3 and Fc rosettes, C3 and Fc IP, lysozyme, macrophages, and granulocytes.
Ly phenotype of T cells cytotoxic for syngeneic mouse mammary tumors: evidence for T cell interactions.
Authors: Stutman O , Shen F W , Boyse E A
Published: May 2006

Specific cell-mediated cytotoxicity (CMC) of lymph node cells from immunized C3Hf mice, against syngeneic C3H/Umc mammary tumor cells, assayed in vitro, is effected by T lymphocytes. This CMC response is biphasic, with an early peak attained within 6 hr and a second major peak beginning at about 18 hr. Effector cells of both the early minor and late major phases of the response belong to the Ly23 set. Other T cell sets evidently play no part in the early effector response. But specifically activated Ly1 cells help or amplify the major late-phase response. Nevertheless, the mixture of specifically activated Ly1 and Ly23 sets still does not completely reconstitute the late response, which implies that the Ly123 set is also needed for maximal expression of CMC in this system. These Ly123 cells must come from specifically immunized donors. It appears, therefore, that maximal CMC is achieved by the participation of specific Ly123 cells which in the late phase directly or indirectly give rise to Ly23 killer cells. Thus, although killing of syngeneic mammary tumor cells in the CMC assay is invariably effected by cells of the Ly23 set, specifically activated cells of the Ly1 set, and probably of the Ly123 set also, are participants in the interactions needed to produce a maximal CMC response.
Neurophysiological and behavioral evidence for selective reinnervation in skin-grafted
Rana pipiens
Authors: Heidemann Merle Knudson
Published: May 2006

Rana pipiens
with skin dorosoventrally reversed can respond to stimulation of the back with forelimb wipes to the belly and to stimulation of the belly with hindlimb wipes to the back. These “misdirected wiping responses” have been explained in terms of two alternative hypotheses of nerve regeneration: nerve respecification or selective reinnervation. Experimental behavioral and neurophysiological experiments reported here support the selective reinnervation hypothesis. Severing ventral nerves, which normally innervate the belly, greatly reduced the percentage of misdirected responses on stimulation of belly skin grafted to the back, while severing dorsal nerves, which normally innervate the back, increased the percentage of misdirected responses elicited under the same circumstances. Moreover, neurophysiological recordings of grafted animals showed three effects of skin grafting on nerve distributions: (
i
) termination of dorsal and ventral nerve receptive field at graft edges; (
ii
) overlap of nonadjacent ventral nerve receptive fields; and (
iii
) dorsal coursing of ventral nerves to reinnervate target belly skin displaced to the back. These neurophysiological observations, and particularly the third effect, also support selective reinnervation as the mechanism of nerve regeneration in skin-grafted
Rana pipiens
.