bluuebunny/crossref_metadata_2025 huggingface dataset:
Extrachromosomal control of mutability in Drosophila melanogaster.
Authors: Engels W R
Published: May 2006

Hybrid dysgenesis is a syndrome of germ-line aberrations including, e.g., sterility and mutation, found in certain interstrain hybrids of Drosophila melanogaster. Previous studies of sterility have shown that elements responsible for dysgenesis may reside on all major chromosomes, but that their dysgenesis-causing ability is controlled by an unknown extrachromosomal factor. Dysgenic hybrids also give rise to unstable visible mutations thought to be DNA insertions at certain sensitive loci. One such unstable allele at the singed bristle locus, designated snw, was found to mutate at extraordinary rates exceeding 50%. This instability was shown to be under the same extrachromosomal control as hybrid dysgenesis itself. That is, the mutability of snw was reversibly suppressed when placed in the background cytotype known to prevent sterility and other characteristics of hybrid dysgenesis. These results suggest that snw may represent an insertion at the singed locus of a hypothetical gene responsible for hybrid dysgenesis.
Enkephalin- and somatostatin-like immunoreactivities in human adrenal medulla and pheochromocytoma.
Authors: Lundberg J M , Hamberger B , Schultzberg M , Hökfelt T , Granberg P O , Efendić S , Terenius L , Goldstein M , Luft R
Published: May 2006

By using the Coons indirect immunofluorescence technique, enkephalin-like immunoreactivity with a granular localization was observed in human adrenal medullary gland cells and pheochromocytomas. In two of the tumors and in a few adrenal gland cells, a somatostatin-like peptide could also be identified. Catecholamine cell types were visualized on adjacent sections with antisera to the synthesizing enzymes dopamine-beta-hydroxylase [DBH; dopamine beta-monooxygenase; 3,4-dihydroxyphenylethylamine, ascorbate: oxygen oxidoreductase (beta-hydroxylating), EC 1.14.17.1] and phenylethanolamine-N-methyltransferase (PNMT; noradrenalin N-methyltransferase; S-adenosyl-L-methionine:phenylethanolamine N-methyltransferase, EC 2.1.1.28). In the normal adrenal medulla more DBH- than PNMT-immunoreactive gland cells were observed. In the adrenal pheochromocytoma both DBH- and PNMT-positive cells were seen, whereas the two extra-adrenal tumors contained only DBH. These findings correlated well with plasma catecholamine measurements. Finally, enkephalin immunoreactive fibers and somatostatin immunoreactive cells were observed in a sympathetic ganglion extirpated together with one of the tumors.
Topological organization of proteins in an intracellular secretory organelle: the synaptic vesicle.
Authors: Wagner J A , Kelly R B
Published: May 2006

Intact synaptic vesicles prepared from the electric organ of the marine elasmobranch Narcine brasiliensis have eight major polypeptides demonstrable on sodium dodecyl sulfate gels. Six of these copurify with the synaptic vesicles during isolation of vesicles by chromatography on CPG-3000 and, by this criterion, are specific to vesicles. The other two are either shared by many membrane or are contaminants. One of these proteins comigrates with actin. Three different approaches were used to determine which proteins were exposed on the external, cytoplasmic surface of the vesicle and which were internal. The first was susceptibility to the proteases trypsin, Streptomyces griseus protease, and Pronase; the second was labeling by the membrane-impermeable reagent diazotized [125I]iodosulfanilic acid; and the third was iodination catalyzed by lactoperoxidase. In general, the three approaches give the same result: six of the eight proteins are on the external, cytoplasmic surface and two are accessible only after the vesicles are lysed by freezing and thawing or by detergents. Five of the vesicle-specific proteins are external and one is internal. The actin-like protein is internal. Proteins involved in the interaction of vesicles with the presynaptic membrane during exocytosis might be expected to be vesicle specific and external.
Enhanced nonenzymatic glucosylation of human serum albumin in diabetes mellitus.
Authors: Guthrow C E , Morris M A , Day J F , Thorpe S R , Baynes J W
Published: May 2006

Use of an ion exchange chromatographic method and a colorimetric method with thiobarbituric acid showed that levels of nonenzymatically glucosylated serum albumin were increased in patients with poorly controlled diabetes mellitus compared to controls. The two methods correlated well (r = 0.99) and clearly discriminated between normal and poorly controlled diabetic populations. The levels of glycosylated hemoglobin were also measured in both populations. Several patients apparently in good control based on glycosylated hemoglobin measurements were found to have increased levels of glycosylated albumin. Because albumin has a shorter circulating half-life than does the human erythrocyte, the plasma concentration of glucosylated albumin should be expected to reflect short-term control of hyperglycemia in diabetes. The studies reported here suggest that the level of glucosylated albumin may indeed be a sensitive indicator of moderate hyperglycemia and of early glucose intolerance.
Mechanism for circularization of linear DNAs: circular parvovirus MVM DNA is formed by a "noose" sliding in a "lasso"-like DNA structure.
Authors: Bratosin S , Laub O , Tal J , Aloni Y
Published: May 2006

During an electron-microscopic survey with the aim of identifying the parvovirus MVM transcription template, we observed previously unidentified structures of MVM DNA in lysates of virus-infected cells. These included double-stranded "lasso"-like structures and relaxed circles. Both structures were of unit length MVM DNA, indicating that they were not intermediates formed during replication; they each represented about 5% of the total nuclear MVM DNA. The proportion of these structures was unchanged after digestion with sodium dodecyl sulfate/Pronase and RNase and after mild denaturation treatment. Cleavage of the "lasso" structures with EcoRI restriction endonuclease indicated that the "noose" part of the "lasso" structure is located on the 5' side of the genomic single-stranded MVM DNA. A model is presented for the molecular nature of the circularization process of MVM DNA in which the "lasso" structures are identified as intermediates during circle formation. This model proposes a mechanism for circularization of linear DNAs.
Synthesis of a 600-nucleotide-long plus-strand DNA by virions of Moloney murine leukemia virus.
Authors: Mitra S W , Goff S , Gilboa E , Baltimore D
Published: May 2006

A discrete, 600-nucleotide-long plus-strand DNA has been identified among the products of reverse transcription by virions of Moloney murine leukemia virus. Its polarity was shown by hybridization to minus-strand DNA. It appears to be copied from the right end of minus-strand DNA because (i) its restriction endonuclease cleavage pattern corresponds to the redundant 600-base segment found at either end of the ultimate double-stranded reverse transcription products, (ii) its synthesis is actinomycin D sensitive, and (iii) its synthesis begins during the first hour of a reverse transcription reaction when only the right-hand end of minus-strand DNA is available as template. We therefore call this DNA plus-strong-stop DNA by analogy with the minus-strong-stop DNA copied from the left end of the viral RNA. Both strong-stop DNAs are made early during in vitro reactions and decline in concentration later, consistent with postulated roles as initiators of long minus- and plus-strand DNA. Unlike minus-strong-stop DNA, plus-strong-stop DNA remains as a double-stranded nucleic acid after its synthesis, as shown by S1 nuclease resistance. A primer to initiate plus-strong-stop DNA synthesis has not been identified; the product found thus far has no detectable RNA attached to it.
Involvement of long-chain acyl coenzyme A for lipid synthesis in repression of acetyl-coenzyme A carboxylase in Candida lipolytica.
Authors: Kamiryo T , Nishikawa Y , Mishina M , Terao M , Numa S
Published: May 2006

Mutant strains of Candida lipolytica defective in acyl-CoA synthetase II [acid:CoA ligase (AMP-forming), EC 6.2.1.3] have been isolated. The mutants fail to grow on fatty acid as a sole carbon source but are capable of incorporating exogenous fatty acid into cellular lipids. This observation, together with our previous finding that mutant strains defective in acyl-CoA synthetase I cannot incorporate exogenous fatty acid into cellular lipids but are able to degrade fatty acid via beta-oxidation, indicates the presence of two functionally distinct long-chain acyl-CoA pools in the cell--i.e., one for lipid synthesis and the other for beta-oxidation. Unlike the wild-type and the revertant strains as well as the mutants lacking acyl-CoA synthetase II, the mutants defective in acyl-CoA synthetase I do not exhibit the repression of acetyl-CoA carboxylase [acetyl-CoA:carbon-dioxide ligase (ADP-forming), EC 6.4.1.2] by exogenous fatty acid. Measurement of the two long-chain acyl-CoA pools with the aid of appropriate mutant strains has indicated that the long-chain acyl-CoA to be utilized for lipid synthesis, but not that to be degraded via beta-oxidation, is involved in the repression of acetyl-CoA carboxylase.
Acute stimulation of aromatization in Leydig cells by human chorionic gonadotropin in vitro.
Authors: Valladares L E , Payne A H
Published: May 2006

Aromatization of testosterone in Leydig cells purified from mature rat testes was assessed. Leydig cells incubated for 4 hr with increasing concentrations of [3H]testosterone. At saturating concentrations of testosterone, human chorionic gonadotropin (hCG) acutely stimulated aromatization. This stimulation was first observed at 1 hr, an 8-fold increase being found during a 4-hr incubation. The maximal amount of estradiol produced at saturating concentrations of testosterone and hCG was 1.8 ng per 10(6) cells. These results demonstrate that purified Leydig cells have a high capacity for aromatization and that hCG can acutely stimulate aromatization independently of stimulating testosterone synthesis in vitro.
Molecular genetics of herpes simplex virus: The terminal
a
sequences of the L and S components are obligatorily identical and constitute a part of a structural gene mapping predominantly in the S component
Authors: Knipe David M. , Ruyechan William T. , Honess Robert W. , Roizman Bernard
Published: May 2006

In herpes simplex virus 1 (HSV-1) DNA, a small sequence, designated the
a
sequence, flanks the reiterated sequences at the ends of both the L and S components. The
a
sequence is the only sequence shared by the termini of all isomeric arrangements of HSV-1 DNA that arise from inversions of the covalently linked L and S components. We report that the
a
sequence, although present in both components, is a part of a structural gene mapping predominantly in the reiterated sequences of the S component. This conclusion is based on the observations that the mutant HSV-1(13)
ts
C75 is rescued by transfection of cells with the mutant DNA and any one of the four terminal or four L-S junction fragments of wild-type DNA. Furthermore, in doubly infected cells, this mutant shows little or no recombination or complementation with other
ts
mutants previously mapped within the reiterated sequences of the S component. Because it is otherwise difficult to explain the isolation of a mutant with several independent, equivalent mutations, the data argue for a mechanism that maintains the identity of the multiple copies of the
a
sequence.

The paradox arising from the two observations that all termini rescue the
ts
mutant but that in coinfection tests the
ts
lesion is closely linked to the reiterated sequences of the S component could be accounted for by postulating that either recombination occurs while the DNA is in a circular form—in which case all
a
sequences would be adjacent to the reiterated sequence of the S component—or recombination can occur while the DNA is in a linear form. In this case the only effective substitution of the
a
sequence that is perpetuated is the one occurring at the L-S junction or in the S component.

In light of the observations that
ts
C75 and the other mutants tested in this study map in the reiterated sequences and fail to yield appreciable recombinational frequencies, it is unlikely that isomerization of the DNA occurs by intramolecular recombination between reiterated sequences.
Nicotinic acetylcholine receptor: evidence for a functionally distinct receptor on human lymphocytes.
Authors: Richman D P , Arnason B G
Published: May 2006

The presence of three distinct cholinergic receptors on human lymphocytes was suggested by the effects of carbamoylcholine on lymphocyte proliferation in vitro. The cells responded to both 0.1 nM and 1 microM carbamoylcholine by increased proliferation which was blocked by the muscarinic antagonist atropine. This effect occurred in both mitogen-stimulated cells (maximum effect at 24 hr) and nonstimulated cells (maximum effect at 72 hr). In contrast, 1--10 nM carbamoylcholine produced diminished in vitro proliferation, an effect which was blocked by the nicotinic antagonists alpha-bungarotoxin and d-tubocurarine.
Target recognition in neurogenesis: formation of the Mauthner axon cap.
Authors: Kimmel C B , Sessions S K , Kimmel R J
Published: May 2006

To learn whether presence of a specific neuron, the Mauthner (M) cell, is required for the organization, during embryogenesis, of an associated synaptic neuropil, the M-axon cap, M-cell precursors were experimentally deleted in embryos of the zebra fish (Brachydanio rerio) and the axolotl (Ambystoma mexicanum). Examination of early larvae revealed that in about half of the cases M-axon caps were present in the absence of the corresponding M-cells. The locations and structures of such caps were approximately normal. We conclude that the M-cell is not a necessary target for normal development of the axon cap.
Isolation and characterization of beta-endorphin-like peptides from bovine brains.
Authors: Swann R W , Li C H
Published: May 2006

Four beta-endorphin-like peptides from bovine brain extracts have been identified by their behavior in CM-cellulose chromatography, gel filtration, high-performance liquid chromatography, and radioimmunoassay. Two of them have been isolated in sufficient quantity for amino acid analysis, radioimmunoassay, and radioreceptor assay. One peptide has an amino acid composition nearly identical to that of beta-endorphin with 56% of the radioimmunoreactivity and 2.3% of the potency in the radioreceptor assay of beta-endorphin. The amino acid content of the other beta-endorphin-like peptide is very different from that of bovine beta-endorphin but it has 47% of the radioimmunoreactivity and 1% of the potency in the radioreceptor assay of beta-endorphin.
Reassembly of protein-lipid complexes into large bilayer vesicles: Perspectives for membrane reconstitution
Authors: Darszon A. , Vandenberg C. A. , Schönfeld M. , Ellisman M. H. , Spitzer N. C. , Montal M.
Published: May 2006

Protein-lipid complexes in apolar solvents reassemble into large bilayer protein-lipid vesicles (PLVs) with diameters of several micrometers. PLVs form spontaneously upon hydration of the protein-lipid complex residue after solvent removal. This procedure has been applied to the following membrane proteins: bovine and squid rhodopsin, reaction centers from
Rhodopseudomonas sphaeroides
, beef heart cytochrome
c
oxidase, and acetylcholine receptors from
Torpedo californica
. PLVs have a large internal aqueous space (e.g., 790 μl/mg of lipid for cattle rhodopsin vesicles). Freeze-fracture replicas of PLVs revealed that both internal and external leaflets contained numerous intramembranous particles with diameters between 80 and 120 Å, depending on the specific protein incorporated in the membrane. The optical spectral properties of rhodopsin and reaction centers in PLVs were similar to those recorded in the respective natural membrane. Furthermore, bovine rhodopsin in PLVs was chemically regenerable with 9-
cis
-retinal. Actinic illumination induced proton efflux from reaction center vesicles that was abolished by proton ionophores. Therefore, this method is suitable for the incorporation of some membrane proteins in their functional state. PLVs were penetrated with microelectrodes and visualized by the injection of a fluorescent dye. Preliminary electrical recordings were obtained by sealing PLVs to a hole in a septum separating two aqueous compartments. These studies suggest that PLVs assembled by this procedure permit the simultaneous analysis of reconstituted membranes by chemical, optical, and electrical techniques.
Isolation of a replication origin complex from Escherichia coli.
Authors: Nagai K , Hendrickson W , Balakrishnan R , Yamaki H , Boyd D , Schaechter M
Published: May 2006

A complex consisting of replicative origin DNA and several proteins was isolated from Escherichia coli. Cells of temperature-sensitive mutants were labeled at the origin and fractionated by sucrose gradient centrifugation. A complex highly purified in origin DNA sedimented as a unique band. This complex dissociated at high concentration, above 0.2 M KCl. Upon dialysis, the complex reformed, allowing further purification of its constituents. Three major protein bands were found, corresponding to proteins of the outer membrane. The complex did not sediment with membrane fractions, but adhered to the outer membrane in the presence of magnesium.
DNA crosslinking and cytotoxicity in normal and transformed human cells treated with antitumor nitrosoureas.
Authors: Erickson L C , Bradley M O , Ducore J M , Ewig R A , Kohn K W
Published: May 2006

Normal (IMR-90) and simian virus 40-transformed (VA-13) human embryo cells were treated with antitumor nitrosoureas, and the effects on cell viability and cell DNA were compared. All six nitrosoureas tested were more toxic to VA-13 cells than to IMR-90 cells as measured by decrease in cell proliferation or in colony formation. The nitrosoureas capable of generating alkylisocyanates produced a smaller difference between the cell types than did derivatives lacking this capacity. DNA damage was measured by alkaline elution in cells treated with four chloroethylnitrosoureas. Whereas VA-13 cells exhibited dose-dependent interstrand crosslinking, little or none was detected in IMR-90 cells. The IMR-90 cells, however, exhibited at least as much DNA-protein crosslinking as did VA-13 cells. The results can be interpreted in terms of a possible difference in DNA repair between the cell lines.
Relation between the local field at large distances from a charge or dipole and the dielectric constant
Authors: Pollock E. L. , Alder B. J. , Pratt L. R.
Published: May 2006

The local field at large distances from a charged or dipolar impurity in a nonpolar polarizable fluid is related to the dielectric constant of the medium. The predicted local field agrees accurately with molecular dynamics results. In contrast, the usual continuum theory prediction is only accurate to first order in the polarizability and leads to serious error for realistic values of the polarizability. At smaller distances these relations cannot be used to define a physically reasonable local dielectric constant and consequently the local field is more satisfactorily described in terms of a screening function.
Crohn disease lymph node homogenates produce murine lymphoma in athymic mice.
Authors: Das K M , Valenzuela I , Morecki R
Published: May 2006

To study the putative agent(s) related to Crohn disease, we intraperitoneally in injected mesenteric lymph node homogenates from four patients with active Crohn disease into 10-week-old athymic (nu/nu) mice. Control mice (nu/nu) were injected with homogenates of mesenteric lymph nodes from two patients with ulcerative colitis and four patients undergoing elective cholecystectomy, and with a homogenate of a cervical lymph node containing sarcoid granuloma. Thirty-four mice received filtered or unfiltered homogenates from Crohn disease lymph nodes. Thirty-two mice received homogenates or filtrates from lymph nodes of control patients. Four mice from the group injected with Crohn disease homogenates from four different patients developed generalized lymphadenopathy due to lymphoma 10-28 weeks after th injection. Two additional mice developed lymphadenopathy due to plasma cell hyperplasia. None of the control mice developed lymphomas or lymphadenopathy. Two lymphomas were homogenized, filtered, and injected intraperitoneally into a second group of nu/nu mice, which also developed lymphoma within 8 weeks of injection. Two lymphomas were cultured in vitro and B cell sur?ACE MARKERS WERE IDENTIFIED. Indirect immunofluorescence studies in two lymphomas showed cytoplasmic staining of lymphoma cells with sera from 10 patients with active Crohn disease but not with sera from 13 control subjects, including 6 with ulcerative colitis and 7 with other gastrointestinal disorders. These results suggest that a transmissible factor present in Crohn disease lymph nodes produces lymphoma in nu/nu mice. Furthermore, sera of Crohn disease patients contain an antibody that recognizes an "antigen(s)" in the murine lymphoma.
Relationships between cytokinin production, presence of plasmids, and fasciation caused by strains of
Corynebacterium fascians
Authors: Murai Norimoto , Skoog Folke , Doyle Marjorie E. , Hanson Richards S.
Published: May 2006

Cytokinin activity in the culture medium of four pathogenic strains of
Corynebacterium fascians
varied from 168 to 0.4 μg of kinetin (6-furfurylaminopurine) equivalents per liter, as compared to 0.2 in an avirulent control.
N
6
-Isopentenyladenine was the predominant cytokinin in the medium of all five strains, and its increased production was correlated with the degree of pathogenicity; however, the virulent strains also produced 8-13 times more
cis
-zeatin [6-(4-hydroxy-3-methyl-
cis
-2-butenylamino)purine] than the avirulent strain. The three most virulent strains (with the higher cytokinin contents in the medium) contained a large (
M
r
≈ 10
8
) plasmid. The barely virulent strain contained a smaller plasmid. No plasmid was detected in the avirulent control. The total cytokinin content (mole%) and biological activity (μg of kinetin equivalent per mg of tRNA) in tRNA were about the same in all three virulent strains and in the avirulent control. Five cytokinins were isolated from each strain. Four were rigorously characterized as 6-(4-hydroxy-3-methyl-
cis
-2-butenylamino)-9-β-D-ribofuranosylpurin e, 6-(3-methyl-2-butenylamino)-9-β-D-ribofuranosylpurine, and their 2-methylthio derivatives. The fifth cytokinin was tentatively identified as 6-(4-hydroxy-3-methyl-
trans
-2-butenylamino)-9-β-D-ribofuranosylpur ine. Ribosyl-
cis
-zeatin was 3-fold higher and
N
6
-isopentenyladenosine was correspondingly lower in the plasmid-containing strains than in the plasmidless control. Because the entire syndrome of fasciation caused by infection can be induced with synthetic cytokinins, the disease would appear to be caused by plasmid-induced high rates of cytokinin production by the bacteria.
Presynaptic membrane potential affects transmitter release in an identified neuron in
Aplysia
by modulating the Ca
2+
and K
+
currents
Authors: Shapiro Eli , Castellucci Vincent F. , Kandel Eric R.
Published: May 2006

We have examined the relationships between the modulation of transmitter release and of specific ionic currents by membrane potential in the cholinergic interneuron L10 of the abdominal ganglion of
Aplysia californica
. The presynaptic cell body was voltage-clamped under various pharmacological conditions and transmitter release from the terminals was assayed simultaneously by recording the synaptic potentials in the postsynaptic cell. When cell L10 was voltage-clamped from a holding potential of -60 mV in the presence of tetrodotoxin, graded transmitter release was evoked by depolarizing command pulses in the membrane voltage range (-35 mV to + 10 mV) in which the Ca
2+
current was also increasing. Depolarizing the holding potential of L10 results in increased transmitter output. Two ionic mechanisms contribute to this form of plasticity. First, depolarization inactivates some K
+
channels so that depolarizing command pulses recruit a smaller K
+
current. In unclamped cells the decreased K
+
conductance causes spike-broadening and increased influx of Ca
2+
during each spike. Second, small depolarizations around resting potential (-55 mV to -35 mV) activate a steady-state Ca
2+
current that also contributes to the modulation of transmitter release, because, even with most presynaptic K
+
currents blocked pharmacologically, depolarizing the holding potential still increases transmitter release. In contrast to the steady-state Ca
2+
current, the transient inward Ca
2+
current evoked by depolarizing clamp steps is relatively unchanged from various holding potentials.
Self-association and base pairing of guanosine, cytidine, adenosine, and uridine in dimethyl sulfoxide solution measured by 15N nuclear magnetic resonance spectroscopy.
Authors: Dyllick-Brenzinger C , Sullivan G R , Pang P P , Roberts J D
Published: May 2006

The self-association of guanosine, cytidine, and adenosine and base pairing between guanosine, cytidine, adenosine, and uridine in dimethyl sulfoxide have been investigated by the variation of their 15N NMR chemical shifts with concentration and temperature. Guanosine, cytidine, and adenosine all showed evidence of self-association by hydrogen bonding. In guanosine/cytidine mixtures, a hydrogen-bonded dimer is formed; however, no base pairing could be detected with adenosine/cytidine or adenosine/uridine mixtures.
Direct observation of the torsional dynamics of DNA and RNA by picosecond spectroscopy.
Authors: Millar D P , Robbins R J , Zewail A H
Published: May 2006

Picosecond time-dependent fluorescence depolarization techniques have been used to monitor the reorientation of ethidium bromide intercalated in DNA and RNA. The fluorescence polarization anisotropy reveals a nonexponential, exp(-at 1/2), torsional relaxation of the DNA double helix and provides an accurate value for its torsional rigidity, C = 1.3 +/- 0.2 X 10(-19) erg cm. Furthermore, from accurate measurements of the limiting anisotropy at zero time, we conclude that there is an additional fast (< 10 psec) internal motion that depends on the viscosity of the medium. Denatured DNA is considerably more flexible than the intact double helix, thus demonstrating the influence of secondary structure on internal motions.
Exons encode functional and structural units of chicken lysozyme.
Authors: Jung A , Sippel A E , Grez M , Schütz G
Published: May 2006

The nucleotide sequence was determined for the chicken egg white lysozyme mRNA and for the exons of the gene together with their flanking intron regions. The exon pattern is to some degree related to the structural subdivision of the final protein product. However, the relationship of exons to functional units of the enzyme is better established. Exon 2 codes for amino acids 28-82, which include the catalytically active residues and a cluster of amino acids which bind rings C, D, E, and F of the oligosaccharide substrate. Exon 3 codes for amino acids 82-108, which give additional substrate specificity, determine the cleavage frame for the alternating N-acetylglucosamine/N-acetylmuramic acid chain, and increase the catalytic efficiency of the active center. Exons 1 and 4, respectively, code for translational signal sequences on the mRNA, for the signal peptide of prelysozyme, and for the amino- and carboxy-terminal regions of the enzyme. These regions increase the stability of the molecule but are not directly involved in the catalytic function.
Identification of ecotropic proviral sequences in inbred mouse strains with a cloned subgenomic DNA fragment.
Authors: Chan H W , Bryan T , Moore J L , Staal S P , Rowe W P , Martin M A
Published: May 2006

A specific probe for detecting ecotropic murine leukemia virus sequences was constructed by cloning a 500-base-pair DNA segment, corresponding to a portion of the env region of the AKR ecotropic virus, in a pBR322/Escherichia coli K-12 host/vector system. This probe was used to screen the cellular DNAs of six inbred strains of mice for the presence of ecotropic retroviral DNA sequences by the Southern blot hybridization procedure. Three copies of ecotropic viral DNA were detected in AKR/N (a high-ecotropic virus strain) and two were found in BALB/c (a low-ecotropic virus strain) DNAs. As expected, no sequences reactive with this probe were found in NFS mouse DNA (a virus-negative strain). However, cellular DNA sequences that reacted strongly with the ecotropic-specific DNA probe were detected in certain NZB, C57L, and 129 mice (all virus-negative strains). In contrast to the reactive sequences in AKR and BALB/c, the reactions were chiefly associated with EcoRI segments that were subgenomic in size.
Dihydrofolate reductase: thymidylate synthase, a bifunctional polypeptide from Crithidia fasciculata.
Authors: Ferone R , Roland S
Published: May 2006

The molecular weight of dihydrofolate reductase (5,6,7,8-tetrahydrofolate:NADP+ oxidoreductase, EC 1.5.1.3) from protozoa has been reported to be 5- to 10-fold larger than the isofunctional enzyme of most other organisms studied, based on gel filtration. This enzyme from the protozoal flagellate Crithidia fasciculata has been purified to homogeneity and found to be a bifunctional protein with thymidylate synthase (5,10-methylene tetrahydrofolate:dUMP C-methyltransferase, EC 2.1.1.45) activity. The purified protein, eluted from methotrexate-Sepharose columns by dihydrofolate, migrated as a single band on both nondenaturing and denaturing polyacrylamide gel electrophoresis. The monomer Mr is 56,700 +/- 200. The native Mr was calculated to be 107,000 from a sedimentation coefficient of 5.9 and Stokes radius of 4.4 nm. Dihydrofolate reductase and thymidylate synthase activities of the rodent malaria organism Plasmodium berghei also copurified on Sephadex G-200 and methotexate-Sepharose columns, suggesting that this unique bifunctional protein might occur throughout the Protozoa.
Comparison of two delta-crystallin genes in the chicken.
Authors: Jones R E , Bhat S P , Sullivan M A , Piatigorsky J
Published: May 2006

Restriction endonuclease mapping and electron microscopy established that there are at least two nonallelic delta-crystallin genes in the chicken. The structure of a complete delta-crystallin gene has been deduced by analysis of five cloned fragments from the chicken genome. Fifteen intervening sequences compose approximately 80% of this gene; the other gene contains at least 14 intervening sequences. The mRNA gene sequences appear to be similar; at least 60% divergence was found between the intervening sequences of the two delta-crystallin genes. Insertions or deletions or both appear to be major events responsible for these differences.
Human platelet basic protein associated with antiheparin and mitogenic activities: purification and partial characterization.
Authors: Paul D , Niewiarowski S , Varma K G , Rucinski B , Rucker S , Lange E
Published: May 2006

Platelet basic protein (PBP) (isoelectric point, 10.0-10.5; apparent Mr, 11,000-15,000) has been purified to homogeneity from material secreted by fresh human platelets after stimulation by thrombin. The purification, using preparative isoelectric focusing and chromatography on heparin-Sepharose, yielded two additional peptides with antiheparin activity that were immunologically identical with PBP: low-affinity platelet factor 4 and beta-thromboglubulin. The purity of the peptides was confirmed by immuoelectrophoresis and by NH2-terminal amino acid analysis. Dansyl chloride-treated PBP yielded a single dansylated amino acid residue (glycine). By using a specific radioimmunoassay it was shown that 10(9) human platelets contain 2-3 microgram of PBP which can be released in response to specific stimulation. PBP is associated with mitogenic activity as assayed in Swiss 3T3 mouse cells cultured in low-serum (0.4-1.5%) medium at levels of about 1 ng/ml and saturating at 10-40 ng/ml. The biological activity of different PBP preparations was variable, presumably due to inhibition by the varying amounts of ampholytes that interfered with the mitogenic activity of the peptide. Mitogenic activity was eluted from NaDodSO4/polyacrylamide gels and shown to comigrate with immunoreactive material and with conventional marker proteins of 14,000-17,000 daltons or with histones of 11,000-15,000 daltons. Evidence is presented that PBP is different from cationic platelet-derived growth factor which has an apparent Mr of 30,000.
Spreading of fibroblasts in medium containing cytochalasin B: formation of lamellar cytoplasm as a combination of several functional different processes.
Authors: Bliokh Z L , Domnina L V , Ivanova O Y , Pletjushkina O Y , Svitkina T M , Smolyaninov V A , Vasiliev J M , Gelfand I M
Published: May 2006

Normal cultured mouse fibroblasts spreading on solid substrate extend and attach numerous pseudopods; lamellar cytoplasm is eventually formed from the attached pseudopods. Fibroblasts spreading in the presence of cytochasin B (CB) from de novo a system of arbor-like branched processes rather than lamellar cytoplasm. The growing and fully formed arbor-like processes, in contrast to normal lamellar cytoplasm, have low contractility and are unable to clear patched concanavalin A receptors from their surfaces; their attachement sites are not associated with microfilament bundles. The cells spreading in medium containing CB and Colcemid do not form well-organized branched structures but extend and attach numerous unstable pseudopods. It is suggested that normal formation of lamellar cytoplasm can be regarded as a combination of several functionally different processes: (a) of rudimentary pseudopodial reactions resistant to CB and Colcemid; (b) of CB-sensitive lamellization; and (c) of Colcemid-sensitive stabilization.
Error-free excision of the cytotoxic,mutagenic N2-deoxyguanosine DNA adduct formed in human fibroblasts by (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene.
Authors: Yang L L , Maher V M , McCormick J J
Published: May 2006

The ability of normal diploid human fibroblasts and excision repair-deficient xeroderma pigmentosum cells (XP12BE, complementation group A) to excise potentially cytotoxic or mutagenic lesions induced in DNA by (+/-)-7 beta, 8 alpha-dihydroxy-9 alpha, 10 alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BzaP-diol epoxide) was determined. Large populations of cells were prevented from replicating by being grown to confluence; after 3 days they were exposed to tritiated BzaP-diol epoxide for 2 hr. One set of cultures was immediately released and assayed for the number of residues covalently bound to DNA, percent survival of colony-forming ability, and frequency of induced mutations. After various periods of time in confluence, other sets were similarly released and assayed. The normal cells exhibited a gradual increase in survival with time held in confluence (recovery from potentially cytotoxic lesions) which was directly correlated with a gradual loss of radioactivity from their DNA and a gradual decrease in the frequency of induced mutations. In contrast, no loss of radioactively labeled carcinogen from the DNA of the XP12BE cells could be detected during a 6-day period and their percent survival and frequency of induced mutations did not change. DNA from normal cells harvested immediately after treatment or after 2, 4, or 8 days in confluence was enzymatically hydrolyzed and analyzed by high-pressure liquid chromatograhy. Only a single peak was detected that cochromatographed with a standard prepared from deoxyguanosine treated with BzaP-diol epoxide. The kinetics of decrease of tritium label in this specific peak corresponded to the decrease in radioactivity of the total DNA with time and with the kinetics of recovery of the cells from the potentially cytotoxic and mutagenic effects of BzaP-diol epoxide. These results suggest that the N2-deoxyguanosine adduct is responsible for these biological effects and indicate that excision repair of this lesion by the normal human cells is "error free."
Manufacture of diploid/tetraploid chimeric mice.
Authors: Lu T Y , Markert C L
Published: May 2006

Tetraploid mouse embryos were produced by cytochalasin B treatment. These embryos usually die before completion of embryonic development and are abnormal morphologically and physiologically. The tetraploid embryos can be rescued to develop to maturity by aggregating them with normal diploid embryos to produce diploid/tetraploid chimeric mice. The diploid/tetraploid chimeric embryos are frequently abnormal: the larger the proportion of tetraploid cells, the greater the abnormality. By karyotype analysis and by the use of appropriate pigment cell markers, we have demonstrated that two of our surviving chimeras are in fact diploid/tetraploid chimeras. One surviving chimera is retarded in growth and displays neurological abnormalities. The coat color chimerism suggests that this chimera is about 50% tetraploid. Another chimera with about 10% tetraploid pigment cells in the coat is only slightly retarded in growth and is a fertile male. Tetraploid cells are distributed in many, if not all, tissues of embryos but evidently are physiologically inadequate to support completely normal development and function in the absence of substantial numbers of normal diploid cells.
Transformation in
Agmenellum quadruplicatum
Authors: Stevens S. Edward , Porter Ronald D.
Published: May 2006

A DNA-mediated transformation system for the blue-green alga
Agmenellum quadruplicatum
, strain PR-6, is described and characterized for DNA concentration dependence, dependence on time of exposure to DNA, phenotypic expression, sensitivity to various enzymes, and competence. The stability of the transformants has been investigated, and genetic backcross and selfing experiments have been performed. This system fulfills all of the criteria established for the well-characterized transformation systems in heterotrophic bacteria and demonstrates significant similarities to at least one of these systems for all characteristics examined. The efficiency of transformation is high. This system fills a need for a well-characterized genetic system in an oxygen-evolving photoautotroph. We have used it to transform a strain with a mutational lesion in assimilatory nitrogen metabolism to a wild-type genotype.
Resurgence of responding after the cessation of response-independent reinforcement.
Authors: Epstein R , Skinner B F
Published: May 2006

In an autoshaping experiment, food-deprived pigeons pecked rapidly at a moving dot that preceded the delivery of food. When the moving dot and food were no longer correlated, the rate of pecking dropped nearly to zero. When, subsequently, no food was given, pecking reappeared at a high rate (nearly 200 pecks per min for each subject), the rate dropping again in subsequent sessions. In two other experiments, designed to clarify relevant variables, the effect was replicated. The data suggest that although response-independent reinforcement produces a decrement in responding, it does not reduce a tendency to respond under other conditions.
Permeabilization of animal cells for kinetic studies of intracellular enzymes: in situ behavior of the glycolytic enzymes of erythrocytes.
Authors: Aragón J J , Felíu J E , Frenkel R A , Sols A
Published: May 2006

Intracellular enzymes in erythrocytes can be made accessible for in situ kinetic studies by treating the cells with bifunctional reagents to crosslink proteins, thus creating a network that allows subsequent permeabilization by delipidation without escape of intracellular proteins. Dimethyl suberimidate, dimethyl 3,3'-dithiobispropionimidate, and toluene-2,4-diisocyanate have been used successfully as crosslinking reagents, and digitonin has been used for delipidation. In a systematic study of the in situ behavior of the 11 glycolytic enzymes of rat erythrocytes, it was observed that Km and Vmax values for the majority of the enzymes are essentially the same in situ as in vitro. Lactate dehydrogenase (L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) is inhibited by excess of pyruvate as much in situ as in vitro. Hexokinase (ATP:D-hexose 6-phosphotransferase, EC 2.7.1.1) was allosterically inhibited by glucose 6-phosphate nearly as much in situ as in vitro but was not affected by 2,3-biphosphoglycerate. The allosteric properties of 6-phosphofructokinase (ATP:D-fructose 6-phosphate 1-phosphotransferase, EC 2.7.1.11), glyceraldehyde-phosphate dehydrogenase [D-glyceraldehyde-3-phosphate:NAD+ oxidoreductase (phosphorylating), EC 1.2.1.12], and pyruvate kinase (ATP: pyruvate 2-O-phosphotransferase, EC 2.7.1.40) in situ were qualitatively similar to those observed in vitro, but some important quantitative differences were noticed. Particularly striking was the much greater activity of phosphofructokinase in situ compared to that in vitro at physiological concentrations of effector metabolites.
Redox conformation changes in refined tuna cytochrome c.
Authors: Takano T , Dickerson R E
Published: May 2006

Tuna ferrocytochrome c and ferricytochrome c have been refined independently at high resolution (1.5 A and 1.8 A) to crystallographic residual errors of 17.3% and 20.8%, respectively. Small but significant conformational differences are seen surrounding a buried water molecule that is hydrogen bonded to Asn-52, Tyr-67, and Thr-78. In the oxidized state, this water molecule is 1.0 A closer to the heme and the heme has moved 0.15 A out of its heme crevice; both changes lead to a more polar microenvironment for the heme. Chemical modification studies, patterns of evolutionary conservatism, structural differences in bacterial cytochromes, and x-ray studies all agree that the "active site" for cytochrome c is bounded by lysines 8, 13,27, 72, 79, 86, and 87 (thus containing the evolutionary conservative 72-87 loop) and has the buried water molecule just below its surface and the opening of the heme crevice slightly to one side.
Conformational flexibility of DNA: polymorphism and handedness.
Authors: Gupta G , Bansal M , Sasisekharan V
Published: May 2006

It is shown that left-handed duplexes are possible for A, B, and D forms of DNA. These duplexes are stereochemically satisfactory and are consistent with the observed x-ray intensity data. On scrutiny the refined right-handed models of B and D DNA by Arnott and coworkers are found to be stereochemically unacceptable. It was possible to formulate a stereochemical guideline for molecular model building based on theory and analysis of single-crystal structure data of dinucleoside monophosphate and higher oligomers. This led to both right- and left-handed DNA duplexes. The right-handed B and D DNA duplexes so obtained are stereochemically superior to earlier models and agree well with the observed x-ray intensity data. The observation that DNA can exist in either handedness for all the polymorphous forms of DNA at once explained A in equilibrium B and B in equilibrium D transitions. Hence it is confirmed that polymorphism of DNA is a reflection on the conformational flexibility inherent in DNA, the same cause that ultimately allows DNA in either handedness. The possibility of various types of right- and left-handed duplexes generated by using dinucleoside monophosphate and trinucleoside diphosphate as repeating units resulted in a variety of models, called RL models. All these models have alternating right and left helical segments and inverted stacking at the bend region as suggested by us earlier. It turns out that the B-Z DNA model of Wang et al. is only an example of RL models.
Autonomous movements of cytoplasmic fragments.
Authors: Albrecht-Buehler G
Published: May 2006

Tiny fragments from the cytoplasm of human skin fibroblasts with about 2% of the original cell volume ("microplasts") were prepared by treatment with cytochalasin B, vigorous pipetting, and trypsinization of the attached fragments. They remained alive for 8 hr or longer. Some of the microplasts were able to produce and move filopodia, ruffle, or both; others blebbed continuously. Slow flattening was observed in the larger microplasts. In all cases tested, microplasts avoided contact with other cells or microplasts. The observations suggest that the cytoplasmic matrix and the membranes of animal cells are so constructed as to express locally and autonomously any one of the elementary amoeboid movements listed above. More importantly, whatever types of motile surface projections a microplast expressed, it continued to produce and move them in a stereotypical way as if there were long-lived structural or material determinants for each type. The microplasts were unable to locomote autonomously. Therefore, it is conceivable that directional movement of whole cells may require a supervising mechanism that confers a certain coordination and strategy on its component cytoplasmic bits. Otherwise they would continue to move in stereotypical and autonomous ways without ever displacing themselves, as suggested by the behavior of the microplasts.
Sanfilippo disease type D: deficiency of N-acetylglucosamine-6-sulfate sulfatase required for heparan sulfate degradation.
Authors: Kresse H , Paschke E , von Figura K , Gilberg W , Fuchs W
Published: May 2006

Skin fibroblasts from two patients who had symptoms of the Sanfilippo syndrome (mucopolysaccharidosis III) accumulated excessive amounts of heparan sulfate and were unable to release sulfate from N-acetylglucosamine-6-sulfate linkages in heparan sulfate-derived oligosaccharides. Keratan sulfate-derived oligosaccharides bearing the same residue at the nonreducing end and p-nitrophenyl-6-sulfo-2-acetamido-2-deoxy-beta-D-glucopyranoside were degraded normally. Kinetic differences between th sulfatase activities of normal fibroblasts were found. These observations suggest that N-acetylglucosamine-6-sulfate sulfatase activities degrading heparan sulfate and keratan sulfate, respectively, can be distinguished. It is the activity directed toward heparan sulfate that is deficient in these patients; we propose that this deficiency causes Sanfilippo disease type D.
Generation of human monoclonal antibodies reactive with human mammary carcinoma cells.
Authors: Schlom J , Wunderlich D , Teramoto Y A
Published: May 2006

Lymphocytes from lymph nodes obtained at mastectomy in breast cancer patients have been fused with murine nonproducer myeloma cells to obtain human-mouse hybridoma cultures that synthesize human monoclonal antibodies. To date, 52 hybridoma cultures synthesizing either human IgG or human IgM have been obtained from lymph nodes of 13 patients. Ig production was stable in many of these cloned cultures through 60-200 days of observation. Levels of human Ig synthesis ranged from 0.1 to 20 microgram/ml of supernatant fluid. The immunological reactivities of the human Igs were assayed on tissue sections by using the immunoperoxidase technique. Several of the human monoclonal antibodies demonstrated preferential binding to human mammary tumor cells. One human IgM monoclonal antibody was used to discriminate between mammary carcinoma cells (from 55 of 59 patients) and normal mammary epithelial cells, stroma, or lymphocytes of the same breast. Decreased binding to some of the benign breast tumors tested and to selected non-breast adenocarcinomas was also observed. This same human monoclonal antibody, however, reacted significantly with metastatic mammary carcinoma cells in lymph nodes of breast cancer patients, with no binding to normal lymphocytes or to stroma of the same node. These studies demonstrate that stable clones of human-mouse hybridomas can be generated by using lymph nodes of mastectomy patients and that clones can be selected which synthesize human monoclonal antibodies reactive with human mammary carcinoma cells.