bluuebunny/crossref_metadata_2025 huggingface dataset:
Aphelion clustering of “new” comets: Star tracks through Oort's cloud
Authors: Biermann L. , Huebner W. F. , Lüst Rh.
Published: May 2006

An explanation is proposed for the observed and often discussed clustering of long-period comet aphelia on the sky. Poisson and several multinomial distributions are applied to the most conspicuous cluster, considering only the 80 “new” and 59 “intermediate” comets with the best-determined orbits. The observed number of aphelion points in adjacent areas in two tests with 24 and 36 equal sky areas, respectively, shows a large deviation from a random distribution. The expected probability frequency for this to happen by chance is <0.1% for the “new” comets alone; it is further reduced when “new” and “intermediate” comets are combined. When these comets are analyzed separately from others, it is found that the clustering is the result of perturbations of their original orbits by the passage of a star, or a stellar system, through Oort's cloud a few million years ago. Because the statistical effect on the change of a comet's angular momentum about the sun is proportional to the square of the ratio of the mass to the velocity of a star relative to the sun, it is
a priori
probable that passages by a few stars should produce clustering of the aphelia, as is discussed in detail.
Isolation of opiate binding components by affinity chromatography and reconstitution of binding activities.
Authors: Cho T M , Ge B L , Yamato C , Smith A P , Loh H H
Published: May 2006

Rat brain membranes exhibiting stereospecific opiate binding activity were solubilized by sonication and detergent treatment. The active material could be bound to an affinity column containing 6-succinylmorphine but could not be eluted with free agonist. Although two protein peaks could be eluted with NaCl, neither possessed binding activity; however, one of the peaks (A), in combination with an acidic lipid fraction, eluted subsequently from the column, showed stereospecific binding. Opiate ligands of the mu type bound to this protein/lipid mixture with an order of affinities closely correlating with those of the original membrane but one to two orders of magnitude lower; binding of delta, kappa, and sigma prototype opioids was considerably less. The protein/lipid mixture also competed with the membranes for mu ligands. These results suggest that the isolated protein-lipid complex may be a component of the opiate receptor and, specifically, the mu receptor or binding site. However, because of the lower affinities of mu opiates for this complex, it is conceivable that some essential membrane component is still missing. Preliminary analysis of peak A indicates that it contains a broad spectrum of protein bands, but it remains to be seen which of these are essential for activity.
Nucleotide sequence of a streptomycete aminoglycoside phosphotransferase gene and its relationship to phosphotransferases encoded by resistance plasmids.
Authors: Thompson C J , Gray G S
Published: May 2006

The DNA sequence of an aminoglycoside phosphotransferase gene (aph) from Streptomyces fradiae ATCC 10745 (a neomycin producer) was determined. The gene was localized by in vitro subcloning and insertional inactivation. Molecular weight, amino acid composition, and amino-terminal analysis of the purified aph gene product confirmed the accuracy and position of the aph gene sequence. Pairwise comparisons of S. fradiae aph with the aph genes encoded by bacterial transposons Tn5 and Tn903 showed significant nucleotide and amino acid homologies which indicated a common evolutionary origin for these antibiotic-resistance genes.
Inhibition of metabolic cooperation by cigarette smoke condensate and its fractions in V-79 Chinese hamster lung fibroblasts.
Authors: Hartman T G , Rosen J D
Published: May 2006

This study was performed to determine the usefulness of the intercellular metabolic cooperation assay for analysis of a complex mixture and to compare the results obtained with previously conducted in vivo tumor promoter assays. One hundred 2R1 cigarettes were smoked according to Federal Trade Commission guidelines and the resulting condensate was separated into a water/methanol-soluble fraction (which was further partitioned into acidic and basic components) and an organic solvent-soluble fraction (which was then chromatographed on silicic acid with petroleum ether, benzene/petroleum ether, benzene, ether, and methanol). The following fractions were positive in the metabolic cooperation assay (in decreasing order of activity): organic solvent-soluble, acidic, whole condensate, and water/methanol-soluble fractions as well as the ether, benzene, and benzene/petroleum ether eluates. The basic fraction and the petroleum ether and methanol eluates were negative. In general, the metabolic cooperation assay results were comparable to previously published results obtained on mouse skin.
Control of malaria virulence by alpha 1-acid glycoprotein (orosomucoid), an acute-phase (inflammatory) reactant.
Authors: Friedman M J
Published: May 2006

During malaria and other infections, the plasma concentration of alpha 1-acid glycoprotein (AGP) increases 3- to 4-fold, but the function of this glycoprotein has been unknown. This study demonstrates, by in vitro culture of the malaria parasite Plasmodium falciparum, that the AGP concentration achieved during malaria is sufficient to inhibit parasite multiplication by 80%. It was found that the inhibitory activity of AGP depends on and is a function of its sialic acid complement (12-16 mol/mol) and its higher-order structure. AGP acts by blocking parasite-erythrocyte interaction during the invasion process. These findings indicate a function for AGP with definite in vivo significance. Moreover, they reveal an important protective response to malaria and perhaps other infectious diseases.
Deficiency of an erythrocyte membrane protein with complement regulatory activity in paroxysmal nocturnal hemoglobinuria.
Authors: Pangburn M K , Schreiber R D , Müller-Eberhard H J
Published: May 2006

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired hemolytic anemia in which the erythrocytes are abnormally sensitive to lysis by complement. A functional deficiency of membrane-associated complement regulators has been demonstrated on PNH erythrocytes. The two factor H-like proteins, the C3b receptor (CR1) and the decay-accelerating factor (DAF), were isolated from normal human erythrocytes, and specific antisera were prepared. Selective inhibition of the two proteins on normal erythrocytes by the antisera demonstrated (i) that the factor responsible for accelerated decay of erythrocyte-bound C3 convertase is DAF and (ii) that the cofactor required for inactivation of erythrocyte-bound C3b by factor I is CR1. PNH erythrocytes were deficient in both of these activities. Erythrocytes deficient in CR1, which were obtained from an apparently healthy individual, exhibited normal DAF activity but no factor I cofactor activity. These cells were not susceptible to complement-mediated lysis in acidified human serum, whereas PNH erythrocytes and Pronase-treated human erythrocytes (which lack DAF and CR1 activities) were lysed by this treatment. It is suggested that the protein primarily responsible for preventing complement activation on normal human erythrocytes is DAF. AMr 73,000 protein isolated from the normal erythrocyte membranes of one PNH patient by using anti-DAF IgG was largely absent from the abnormal erythrocytes of this individual, suggesting that PNH cells lack the DAF protein. CR1 antigen, however, was present on the abnormal PNH erythrocytes. The results suggest that the primary molecular defect underlying the clinical manifestations of PNH may be the lack of the membrane-associated DAF protein and that the abnormal cells may also exhibit impaired CR1 function.
Mammalian pancreatic preproglucagon contains three glucagon-related peptides.
Authors: Lopez L C , Frazier M L , Su C J , Kumar A , Saunders G F
Published: May 2006

We have isolated cDNA clones encoding bovine pancreatic preproglucagon. Twenty-five putative preproglucagon clones were selected by screening 3,100 clones of a fetal bovine pancreas cDNA library with a synthetic oligodeoxynucleotide probe. The probe was a mixture of synthetic 17-base DNA oligomers constructed to correspond to the six carboxyl-terminal amino acids (residues 24-29) of mature glucagon. Restriction mapping of six of these clones suggested that they represented a single mRNA species. Primary sequence analysis of one clone containing a 1,200-base-pair DNA insert revealed that it contained an essentially full-length copy of glucagon mRNA. Analysis of the cDNA suggested a protein coding sequence of 540 nucleotides and 5'- and 3'-untranslated regions of 90 and 471 nucleotides, respectively. This cDNA sequence encoded a 20-amino acid signal sequence followed by one for glicentin, a 69-amino acid polypeptide containing an internal glucagon moiety that has been found in porcine intestines. Glicentin is followed by two additional glucagon-like peptides, each flanked by paired basic amino acids (Lys, Arg) characteristic of prohormone processing. These polypeptide sequences show striking homology with those for glucagon and other members of the glucagon family of peptides.
Isolation of the nitrogen assimilation regulator NR
I
, the product of the
glnG
gene of
Escherichia coli
Authors: Reitzer Lawrence J. , Magasanik Boris
Published: May 2006

The product of the
glnG
gene, a member of the complex
glnALG
operon, is an essential component in the response of
Escherichia coli
K-12 and other enteric bacteria to nitrogen-limited growth. We have purified this protein which we propose to call “NR
I
,” for
n
itrogen
r
egulator I, to about 95% purity from an overproducing strain. Purified NR
I
was identified as a dimer by gel filtration. NR
I
specifically inhibited initiation of transcription from a DNA fragment containing the
glnL
promoter but was without effect on
lacZ
transcription. We determined the intracellular concentration of NR
I
under different growth conditions by using immunological techniques. The ratio of glutamine synthetase polypeptides, the product of the
glnA
gene, to NR
I
polypeptides was about 80:1. NR
I
was not rapidly degraded after ammonia shock, even though the ability to activate nitrogen-controlled systems was lost.
Flat revertants isolated from Kirsten sarcoma virus-transformed cells are resistant to the action of specific oncogenes.
Authors: Noda M , Selinger Z , Scolnick E M , Bassin R H
Published: May 2006

Two flat revertants have been isolated from mutagen-treated populations of Kirsten murine sarcoma virus (Ki-MuSV)-transformed NIH/3T3 cells. These revertants, which appear to be cellular variants resistant to transformation by the Ki-MuSV oncogene v-Ki-ras, contain Ki-MuSV-specific DNA, elevated levels of the v-Ki-ras gene product p21, and rescuable transforming virus. Cell hybridization studies indicated that the revertant phenotype is dominant in hybrids between revertant cells and cells transformed by Ki-MuSV or the closely related Harvey MuSV and BALB MuSV. Analysis of hybrid cells resulting from the fusion of these revertants to cell lines transformed by other retroviruses showed that the action of certain oncogenes structurally unrelated to v-Ki-ras also could be suppressed. Thus, there appear to be functional relationships and diversities among transforming genes (oncogenes) not readily apparent from their structural characteristics.
Amphotericin B selection of mutant Chinese hamster cells with defects in the receptor-mediated endocytosis of low density lipoprotein and cholesterol biosynthesis.
Authors: Krieger M , Martin J , Segal M , Kingsley D
Published: May 2006

This paper describes a rapid and efficient two-step procedure for the isolation of mutant cells with defects in receptor-mediated endocytosis. The procedure takes advantage of two fungal metabolites, compactin (ML236B), a potent inhibitor of cholesterol biosynthesis, and amphotericin B, a polyene antibiotic that forms toxic complexes with sterols in membranes. Mutagen-treated Chinese hamster ovary cells were preincubated overnight in a medium containing mevalonate, low density lipoprotein (LDL), and compactin (Mev/LDL/Com). At the end of the preincubation period, wild-type cells were cholesterol replete while mutant cells that could not utilize the cholesterol in LDL were cholesterol deficient. Subsequent incubation with amphotericin B for 6 hr killed most of the wild-type cells. After a second round of Mev/LDL/Com-amphotericin B selection, endocytosis-defective clones appeared at a frequency of approximately equal to 2.6 X 10(-5). Some of these clones expressed LDL receptor-defective phenotypes and fell into one of two previously defined classes of mutation. Sensitivity of the mutants to infection by vesicular stomatitis virus suggested that the mutations do not disrupt the coated pit-coated vesicle pathway of endocytosis. Minor modifications in the Mev/LDL/Com-amphotericin B selection permit the isolation of cholesterol auxotrophs and might allow the isolation of conditional-lethal mutations. Because LDL can be coupled to ligands that bind to receptors other than the LDL receptor, Mev/LDL/Com-amphotericin B selection may permit the isolation of mutant cells with defects that specifically disrupt other endocytic pathways.
Antibodies to the epidermal growth factor receptor block the biological activities of sarcoma growth factor.
Authors: Carpenter G , Stoscheck C M , Preston Y A , DeLarco J E
Published: May 2006

The role of the epidermal growth factor (EGF) receptor system in mediating the biological activities of sarcoma growth factor (SGF) has been assessed by using specific anti-EGF receptor antibodies. There are two classes of anti-EGF receptor antibodies, those that block binding of 125I-labeled EGF (125I-EGF) and those that do not block binding but do interact with a portion of the EGF receptor on the surface of intact cells. Antisera of both types have been assayed for their capacity to affect the biological activities of SGF. The antisera that block 125I-EGF binding to its receptor block the induction of DNA synthesis in human fibroblasts by either EGF or SGF but not by other polypeptide mitogens. Titration of the anti-EGF receptor antiserum indicates the presence of one population of antibody that blocks the site of both EGF and SGF action. Antisera to the EGF receptor that block 125I-EGF binding also inhibited the SGF-dependent anchorage-independent growth of normal cells in soft agar. The antisera to the EGF receptor that does not block 125I-EGF binding or EGF activity did not inhibit any of the biological activities of SGF. The results suggest that occupation of the EGF receptor is required for both the mitogenic and colony-forming activity of SGF.
Partial purification and characterization of an antigen-specific helper factor synthesized by a T-cell continuous line.
Authors: Lifshitz R , Apte R N , Mozes E
Published: May 2006

Antigen-specific factors produced by the T-cell growth factor-dependent T-cell continuous line E-9M(+) were partially purified. Gel analysis of the twice-affinity-purified eluate of a poly(Tyr,Glu)-poly(DLAla)--poly(LLys) [(T,G)-A--L] column revealed the existence of iodinated bands with molecular weights of 17,000 and 15,000, in addition to a diffuse band of high molecular weight. The specific helper activity of the E-9M(+) supernatants was associated with a precipitate from 65-75% ammonium sulfate. Gel electrophoresis of either the eluate of a (T,G)-A--L column or of the 65-75% salt precipitate indicated that in both preparations two fractions contained the biological activity of the factor, one of a high (less than 67,000) molecular weight and the other of a low (15,000-17,000) one. Culture supernatants of the internally [35S]methionine-labeled E-9M(+) line were subjected to a combined purification of sequential ammonium sulfate precipitations, followed by affinity chromatography. Polyacrylamide gel patterns obtained of the eluates of the different salt precipitates demonstrated that the 65-75% ammonium sulfate precipitate contained two 35S-labeled bands with apparent molecular weights in the range of 60,000 and 15,000, similar to the activity patterns obtained by the gel electrophoresis fractionation experiments. Thus, it is suggested that a fraction of low molecular weight preserves the antigen specificity and the helper activity of the factor produced by the T-cell line.
Isolation and characterization of monoclonal antibodies directed against the DNA repair enzyme uracil DNA glycosylase from human placenta.
Authors: Arenaz P , Sirover M A
Published: May 2006

A series of monoclonal antibodies has been prepared against the base excision repair enzyme uracil DNA glycosylase isolated from human placenta. Spleen cells from BALB/c mice immunized with purified human placental uracil DNA glycosylase were fused with either P3X63 Ag8.653 or SP2/0 myeloma cells. Hybridomas producing antibodies directed against the placental glycosylase were identified in an enzyme-linked immunosorbent assay. Each positive hybridoma was cloned twice by limit dilution and tested for anti-glycosylase activity in an enzyme immunoprecipitation assay. Each of the four clones examined in detail precipitated enzyme activity in an immunoprecipitation reaction only in the presence of rabbit anti-mouse IgG as a second antibody. No anti-uracil DNA glycosylase activity was observed in a spontaneous hybridoma used as a control. Each monoclonal antibody immunoprecipitated uracil DNA glycosylases isolated from several human tissues. Partial crossreactivity was observed with rat liver glycosylase and with a hamster enzyme. In contrast, no crossreactivity was observed with yeast or Escherichia coli glycosylase. Glycerol gradient sedimentation analysis demonstrated that one of the antibodies bound to the glycosylase at a site that did not diminish its catalytic activity. A second monoclonal antibody bound at a determinant that affected catalytic activity. Analysis of antibody-glycosylase interactions suggests that human cells contain antigenically distinct glycosylase species that may be encoded by individual uracil DNA glycosylase genes. The potential use of these monoclonal antibodies in studies examining the regulation of glycosylase isoenzymes during cell proliferation in normal human cells and in cells from cancer-prone individuals is considered.
A host protein (La) binds to a unique species of minus-sense leader RNA during replication of vesicular stomatitis virus.
Authors: Wilusz J , Kurilla M G , Keene J D
Published: May 2006

Baby hamster kidney cells infected with the minus-strand RNA virus vesicular stomatitis virus (VSV) were found to contain three small viral leader RNA species of the minus sense. The longest minus-strand leader RNA was 54 nucleotides long and was complexed with the host cell La protein that was immunoprecipitated by antisera from patients with systemic lupus erythematosus. The La protein is normally found associated with RNA polymerase III transcripts in their unprocessed form. Shorter minus-strand leader RNA species of 45-48 nucleotides were more abundant but were not associated with the La protein. Unlike the plus-strand leader RNA of VSV, the minus-strand leader RNAs were not detected in the nucleus in any form. The minus-strand leader RNAs accumulated gradually throughout the infection and could not be found in association with the viral nucleocapsid protein. The sequence required for La protein binding on the 54-nucleotide-long minus-strand leader is similar to that at the 3' end of the La protein binding-plus-strand leader RNA and, thus, we propose a role for the La protein in the replication of VSV.
Identification of a molecular weight 43,000 protein kinase in acetylcholine receptor-enriched membranes.
Authors: Gordon A S , Milfay D , Diamond I
Published: May 2006

A photoaffinity ATP ligand is used to identify the protein kinase present in acetylcholine receptor-enriched membranes from Torpedo californica. Incubation of these membranes with 8-azido-[alpha-32P]ATP and subsequent irradiation with UV light resulted in covalent labeling of a major band of Mr 43,000. Alkali-stripped membranes that show a selective reduction in the Mr 43,000 polypeptide also show a corresponding reduction in incorporation of photoaffinity label. In addition, the neutralized alkaline extract also showed one band at Mr 43,000 when labeled with the photoaffinity ligand. After alkali extraction, endogenous protein kinase activity decreased in the membranes in proportion to the loss of Mr 43,000 peptide. Moreover, the alkaline extract was able to phosphorylate casein in an exogenous assay system. These results suggest that a Mr 43,000 polypeptide in acetylcholine receptor-enriched membranes is the acetylcholine receptor kinase.
Molecular cloning and characterization of hepatitis A virus cDNA.
Authors: Ticehurst J R , Racaniello V R , Baroudy B M , Baltimore D , Purcell R H , Feinstone S M
Published: May 2006

Double-stranded cDNA was synthesized from hepatitis A virus (HAV) RNA and inserted into the Pst I site of pBR322. Restriction endonuclease digestion and cross-hybridization of fragments yielded a map of overlapping cloned cDNAs that included at least 99% of the viral genome. Molecular clones containing HAV cDNA were identified by hybridizing cloned cDNA to electrophoretically resolved RNA from uninfected and HAV-infected tissue culture cells. Cloned cDNA probes specifically hybridized to RNA from infected cells, and the predominant species identified had the characteristic genomic length of picornaviral RNA (approximately equal to 7,500 nucleotides). A partial sequence from the 3' end of the genome revealed 414 bases in an open reading frame followed by two closely spaced stop codons, a 60-base noncoding region, and a tract of poly(A).
Z-DNA immunoreactivity in fixed metaphase chromosomes of primates.
Authors: Viegas-Péquignot E , Derbin C , Malfoy B , Taillandier E , Leng M , Dutrillaux B
Published: May 2006

Antibodies against Z-DNA bind to fixed metaphase chromosomes of man and Cebus albifrons (Platyrrhini, Primate). By indirect immunofluorescence and indirect immunoperoxidase techniques, a heavy staining is detected in some segments of chromosomes of C. albifrons. These segments correspond to R-band-positive heterochromatin, which has a high G + C-base content. Euchromatin of human and Cebus chromosomes show a weak and heterogeneous staining that consistently reproduces an R- and T-banding pattern in both species. Because chromosome homologies previously were demonstrated between these distantly related species by chromosome banding, our results suggest that Z-DNA has been conserved during the course of primate evolution.
Two classes of mouse mast cells delineated by monoclonal antibodies.
Authors: Katz H R , LeBlanc P A , Russell S W
Published: May 2006

A panel of rat monoclonal antibodies was used to classify mouse mast cells from four different sources on the basis of their surface determinants. Three populations of mast cells were derived by culturing normal bone marrow, spleen, and peripheral blood cells, whereas the fourth population was isolated directly from the peritoneal cavity. For each population, a surface antigen phenotype was determined by indirect immunofluorescence microscopy and compared to the phenotypes of the other populations of mast cells. The results showed that culture-derived mast cells, regardless of origin, had nearly identical antigen phenotypes, suggesting that these populations were comparable. By contrast, peritoneal mast cells had a different phenotype, suggesting that they represented a class distinct from culture-derived mast cells. These findings, which corroborate classification schemes based on nonimmunologic methods, demonstrate that subpopulations of mouse mast cells can be distinguished antigenically.
On the optimal sex ratio.
Authors: Karlin S , Lessard S
Published: May 2006

The equilibrium structures for various multiallele sex-determining genetic models of panmictic populations with sex expression depending on the genotypes of either the zygote or a parent are described. Even-sex-ratio equilibrium surfaces can exist apart from equilibrium points having coincident male and female allelic frequency sets. The latter (symmetric) equilibrium states entail biased sex ratios. A stable symmetric equilibrium and an even-sex-ratio equilibrium segregating the same alleles cannot coexist. The tendency to evolve toward an even-sex ratio is embodied by the following optimality property: starting from an equilibrium having a biased sex ratio, mutant sex-determining alleles can accumulate only in the case in which in the augmented system all attainable equilibrium states have a sex ratio closer to one to one.
Restoration of Ca
2+
influx and degranulation capacity of variant RBL-2H3 cells upon implantation of isolated cromolyn binding protein
Authors: Mazurek N. , Bashkin P. , Loyter A. , Pecht I.
Published: May 2006

Recently, variants of the rat basophilic leukemia cells (RBL-2H3), deficient in their binding capacity for the antiallergic drug cromolyn but displaying unimpaired ability to bind IgE, were selected and cloned [Mazurek, N., Bashkin, P., Petrank, A. & Pecht, I. (1983)
Nature
(
London
) 303, 528-530]. Although the histamine content and the number of IgE receptors in these variants are similar to those of the parental cells, they cannot be stimulated immunologically to allow Ca
2+
influx and to degranulate. In contrast, the Ca
2+
ionophore A23187 causes these variants to degranulate, indicating that the mechanism distal to the Ca
2+
gating is intact in the variants. The cromolyn binding protein (CBP), present in the membranes of RBL-2H3 cells, has recently been isolated by affinity chromatography under nondenaturing conditions. In the current study we have used Sendaivirus envelopes as fusogenic carriers to implant the purified CBP into the membrane of variant basophils that were defective in it. This fusion leads to the restoration of Ca
2+
uptake and degranulation capacity of the variants after IgE-mediated stimulation. These restored activities seem to show a sigmoidal dependence on the amount of incorporated CBP. Saturation values comparable to those of the parental line are reached when the level of implanted CBP approaches its density on the latter line. The restored capacity is due to the implanted CBP, because the reinstated immunological response can be blocked by the inhibitory drug cromolyn and by monoclonal antibodies specific to CBP, both shown to prevent Ca
2+
uptake and degranulation in mast cells and parental RBL-2H3 cells. These results point out that CBP plays an important role in the Ca
2+
gating process resulting in degranulation.
Renal mineralocorticoid receptors and hippocampal corticosterone-binding species have identical intrinsic steroid specificity.
Authors: Krozowski Z S , Funder J W
Published: May 2006

There is current evidence for two classes of hippocampal glucocorticoid receptors (GR)--one classical, [3H]dexamethasone [( 3H]Dex)-binding sites in glial cells, and the other [3H]corticosterone-preferring sites in neuronal cells. In the presence of 1 microM of the synthetic glucocorticoid RU26988 (11 beta, 17 beta-dihydroxy-17 alpha-propynylandrost-1,4,6,-trien-3-one) to exclude tracer from [3H]Dex sites, hippocampal cytosol from adrenalectomized/ovariectomized Sprague-Dawley rats binds [3H]Dex to sites (Kd at 4 degrees C, 0.77 X 10(-9) M; 65 fmol/mg of protein) with the following order of specificity: aldosterone (Aldo) = 9 alpha-fluorocortisol (9 alpha F-cortisol) = deoxycorticosterone (DOC) = corticosterone greater than cortisol much greater than Dex; [3H]Aldo, [3H]DOC, and [3H]corticosterone binding show identical specificity in the presence of RU26988. Addition of 1% adrenalectomized/ovariectomized rat plasma (but not plasma heated at 56 degrees C for 30 min) alters the specificity to: 9 alpha F-cortisol greater than or equal to Aldo greater than or equal to DOC much greater than Dex greater than or equal to corticosterone greater than or equal to cortisol, consistent with sequestration of DOC, corticosterone, and cortisol by transcortin and similar to classical mineralocorticoid receptor (MR) binding of [3H]Aldo in renal cytosol (9 alpha F-cortisol greater than or equal to Aldo greater than or equal to DOC much greater than corticosterone greater than or equal to cortisol greater than or equal to Dex). Separation of other renal binders from transcortin by hydroxylapatite adsorption established the intrinsic specificity of [3H]Aldo binding to MR as: DOC greater than or equal to Aldo greater than or equal to 9 alpha F-cortisol greater than or equal to corticosterone greater than cortisol much greater than Dex, parallel to that of the [3H]corticosterone-binding sites in hippocampus. These studies suggest (i) that hippocampal [3H]corticosterone-binding sites and renal MR may have identical intrinsic specificity for steroids, with apparent specificity differences the result of tissue-specific sequestration of naturally occurring steroids other than Aldo and (ii) that an identical steroid-binding species may thus be occupied under physiological conditions by a mineralocorticoid in one tissue (kidney) and a glucocorticoid in another (hippocampus).
Establishment and characterization of 10 cell lines derived from patients with adult T-cell leukemia.
Authors: Hoshino H , Esumi H , Miwa M , Shimoyama M , Minato K , Tobinai K , Hirose M , Watanabe S , Inada N , Kinoshita K , Kamihira S , Ichimaru M , Sugimura T
Published: May 2006

By using human T-cell growth factor (TCGF), 10 cell lines were established from tissue samples of 10 patients with adult T-cell leukemia (ATL). Three cell lines were adapted to growth in medium lacking TCGF. The surface markers of all cell lines were characteristic of inducer/helper T cells, i.e., OKT3+, OKT4+, OKT6-, OKT8-, OKIa1+, and human Lyt2+ and Lyt3+, except that one cell line was OKT3-. The expression of the viral antigen was examined during establishment of 8 of the 10 cell lines. The viral antigen was not expressed in leukemic cells before cultivation. In 5 lines, the viral antigen was detected by immunofluorescent staining after a short period of cultivation. However, 3 cell lines, ATL-6A, ATL-9Y, and ATL-1K did not express the viral antigen during short-term culture: the ATL-6A and ATL-9Y cell lines became positive for the viral antigen after 5 and 2 months of cultivation, respectively; the ATL-1K cell line remained antigen-negative throughout a culture period of 13 months. Southern blot hybridization assay showed that all of the cell lines, including the viral antigen-negative ATL-1K cell line, contained the viral genome. Thus, the retrovirus was associated with all 10 cell lines established from ATL patients, but there was a heterogeneity in the expression time of the retroviral antigen in leukemic cells maintained in vitro. Our findings suggested that the expression of the viral antigen was not required for maintenance of the leukemic state in vivo and for growth of leukemic cells in vitro.
Isolation and preliminary characterization of the transforming gene of a human neuroblastoma cell line.
Authors: Shimizu K , Goldfarb M , Perucho M , Wigler M
Published: May 2006

DNA from the human neuroblastoma cell line SK-N-SH is capable of inducing foci of transformed NIH 3T3 cells after DNA-mediated gene transfer. Using genetic selection with the Escherichia coli sup F gene, we have isolated human sequences from mouse cells responsible for the oncogenic transformation. These sequences are present in all human DNAs surveyed and no gross rearrangements of these sequences are found in SK-N-SH cells. Although clearly distinct from two other human transforming genes present in bladder, lung, and colon carcinoma cell lines, all three transforming gene sequences may be related members of the ras gene family.
Isolation and characterization of a full-length expressible cDNA for human hypoxanthine phosphoribosyl transferase.
Authors: Jolly D J , Okayama H , Berg P , Esty A C , Filpula D , Bohlen P , Johnson G G , Shively J E , Hunkapillar T , Friedmann T
Published: May 2006

We have cloned a full-length 1.6-kilobase cDNA of a human mRNA coding for hypoxanthine phosphoribosyltransferase (HPRT; IMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.8) into a simian virus 40-based expression vector and have determined its full nucleotide sequence. The inferred amino acid sequence agrees with a partial amino acid sequence determined for authentic human HPRT protein. Transfection of HPRT-deficient mouse LA9 cells with the purified plasmid leads to the expression of human HPRT enzyme activity in cells stably transfected and selected for enzyme activity in hypoxanthine/aminopterin/thymidine medium.
Structural and transcriptional evidence for related thrS and infC expression.
Authors: Mayaux J F , Fayat G , Fromant M , Springer M , Grunberg-Manago M , Blanquet S
Published: May 2006

The nucleotide sequence of thrS, the gene encoding dimeric Escherichia coli threonyl-tRNA synthetase [L-threonine:tRNAThr ligase (AMP forming), 6.1.1.3], has been determined. The structural part of the gene is found upstream of and on the same DNA coding strand as infC, the gene for translational initiation factor IF3. thrS is composed of 1,926 base pairs and accounts for a protein of molecular weight 73,906. In addition, a 336-base-pair sequence 5' to the thrS structural gene has been determined. There are only three nucleotides between the stop codon of thrS and the initiator codon of infC. The only potent transcriptional terminator structure is 55 base pairs downstream of the infC coding sequence. This implies that thrS and infC can be expressed from a polycistronic mRNA originating from a promoter upstream to thrS. Although sequence data indicate that thrS and infC are cotranscribed, in vitro transcription and RNA sequence analyses reveal the existence of a promoter within thrS. This promoter can account for the independent expression of infC as reported [Springer, M., Plumbridge, J. A., Trudel, M., Graffe, M. & Grunberg-Manago, M. (1982) Mol. Gen. Genet. 186, 247-252]. A second promoter has been located within infC and could link the expression of infC and that of the next downstream gene, pdzA. Whether these promoters function normally in vivo is an open question.
Vanadium ions stimulate DNA synthesis in Swiss mouse 3T3 and 3T6 cells.
Authors: Smith J B
Published: May 2006

Vanadyl sulfate and sodium orthovanadate in the concentration range between 5 and 50 microM are shown to be mitogenic for quiescent cultures of Swiss mouse 3T3 and 3T6 cells. The compounds caused a striking shift in the dose-response for the effect of serum on [3H]thymidine incorporation and DNA synthesis. In the absence of serum the effect of vanadium was greatly potentiated by insulin. Vanadium ions produced no more than additive increases in [3H]thymidine incorporation when combined with epidermal growth factor, cholera toxin, or phorbol 12-myristate 13-acetate. Both vanadium compounds stimulated ouabain-inhibitable 86Rb+ uptake, indicating that the vanadium ions increase, rather than inhibit, Na+/K+ pump activity in the intact cell. Neither vanadium compound had any effect on cellular cAMP under a variety of different conditions. The mitogenic effect of the vanadium compounds was similar to that of colchicine. Taxol, which stabilizes cytoplasmic microtubules, prevented the stimulation of DNA synthesis by vanadium.
Protein-associated intercalator-induced DNA scission is enhanced by estrogen stimulation in human breast cancer cells.
Authors: Zwelling L A , Kerrigan D , Lippman M E
Published: May 2006

Estrogen-responsive human breast cancer cells (MCF-7) displayed a higher frequency of intercalator-induced protein-associated DNA scission after treatment with 17 beta-estradiol (E2) than did cells that had not received estrogen treatment. This effect was dependent on estrogen concentration (maximum enhancement at approximately equal to 1 nM E2) and time (maximum effect seen approximately equal to 24 hr after E2 addition). Human breast cancer cells lacking estrogen receptors did not display the enhanced response. Antiestrogens produced a slight decrease in intercalator-induced DNA scission, whereas insulin produced an enhanced effect. The DNA breaks produced by the intercalators 5-iminodaunorubicin and 4'-(9-acridinylamino)methanesulfon-m-anisidide (m-AMSA) in these cells were undetectable without enzymatic deproteinization of cell lysates prior to quantification by alkaline elution. Intercalator-induced DNA-protein crosslinking also was enhanced in E2-treated MCF-7 cells. Studies with m-[14C]AMSA revealed no estrogen-associated increases in drug uptake. The data suggest that E2 treatment, either by specifically and directly increasing active transcription in chromatin or through secondary effects on DNA that accompany alterations in cell growth or cell cycle distribution, alters the susceptibility of DNA to intercalator-induced protein-associated DNA scission. If this enhanced protein-associated scission is selectively localized to transcriptionally active chromatin, the adsorption of the DNA-bound proteins to membrane filters (DNA-protein crosslinking) may allow identification and isolation of estrogen-regulated gene sequences.
Macrophage-mediated cytotoxicity: role of a soluble macrophage cytotoxic factor similar to lymphotoxin and tumor necrosis factor.
Authors: Zacharchuk C M , Drysdale B E , Mayer M M , Shin H S
Published: May 2006

Guinea pig peritoneal macrophages, when activated for cytotoxicity by the calcium ionophore A23187 or lipopolysaccharide, produce a cytotoxic factor [macrophage cytotoxic factor (M phi-CF)] that is not blocked by catalase or protease inhibitors. Fractionation of culture supernates containing M phi-CF by gel filtration revealed one peak of cytotoxic activity of Mr approximately 45,000, the same as guinea pig lymphotoxin (LT). Antiserum prepared against purified guinea pig LT completely neutralized the cytotoxic activity of M phi-CF. In addition, the cytotoxic factor in guinea pig tumor necrosis serum was found to have a Mr of 45,000 and was neutralized by anti-LT. Thus, M phi-CF is physicochemically and immunochemically similar to LT and tumor necrosis factor, if not identical. To investigate the role of M phi-CF in macrophage-mediated cytotoxicity, anti-LT was added to A23187- or lipopolysaccharide-activated macrophages before addition of L-929 target cells. In 10 of 16 experiments, the inhibition of macrophage-mediated cytotoxicity was 100%. In the others, cytotoxicity was blocked partially, the lowest inhibition being 49%. The effectiveness of inhibition appeared to be inversely related to the intensity of macrophage activation. These results indicate that M phi-CF plays a significant role in macrophage-mediated cytotoxicity but involvement of another mechanism cannot be excluded.
Suppression of liver uptake of liposomes by dextran sulfate 500.
Authors: Patel K R , Li M P , Baldeschwieler J D
Published: May 2006

The effect of dextran sulfate (DS, 500,000 Mr) and multilamellar vesicles (MLV) as liver blockade agents has been investigated in mice. Intravenous injection of unlabeled MLV prior to radioactive MLV caused moderate reduction in the liver uptake and increased tibia, lung, and spleen uptake. More drastic differences were observed with intraperitoneal injection of DS. When tested in the range of 0-50 mg of DS per kg of body weight, maximal liver blockade occurred at a dose of 50 mg. By using 50 mg of DS per kg, maximal liver blockade occurred at 12 hr after DS injection. The liver blockade was temporary, ending within 48 hr. The intraperitoneal route of injection for DS was more effective for liver blockade than the intravenous route.
Avian carcinoma virus MH2 contains a transformation-specific sequence, mht, and shares the myc sequence with MC29, CMII, and OK10 viruses.
Authors: Kan N C , Flordellis C S , Garon C F , Duesberg P H , Papas T S
Published: May 2006

Avian carcinoma virus MH2 has been grouped together with MC29, CMII, and OK10, because all of these viruses share a transformation-specific sequence termed myc. A 5.2-kilobase (kb) DNA provirus of MH2 has been molecularly cloned. The complete genetic structure of MH2 is 5'-delta gag(1.9-kb)-mht(1.2-kb)-myc(1.3-kb)-delta env(?) and noncoding c-region (0.2-kb)-3'. delta gag, delta env, and c are genetic elements shared with nondefective retroviruses, whereas mht is a unique, possibly MH2 transformation-specific, sequence. Hybridizations with normal chicken DNA and cloned chicken c-myc DNA indicate that the mht sequence probably derives from a normal cellular gene that is distinct from the c-myc gene. The genetic structure of MH2 suggests that the delta gag and mht sequences function as a hybrid gene that encodes the p100 putative transforming protein. The myc sequence of MH2 appears to encode a second transforming function. Therefore, it seems that MH2 contains two genes with possible oncogenic function, whereas MC29, CMII, and OK10 each carries a single hybrid delta gag-myc transforming gene. It is remarkable that, despite these fundamental differences in their primary structures and mechanisms of gene expression, MH2 and MC29 have very similar oncogenic properties.
Transformation of human skeletal muscle cells by simian virus 40.
Authors: Miranda A F , Babiss L E , Fisher P B
Published: May 2006

Molecular studies of the biochemical alterations involved in human myopathies have been restricted because of the finite life-span and slow growth rate of cultures derived from primary tissue. Because the tumor virus simian virus 40 (SV40) can alter both the growth properties and longevity of human cells, we have infected skeletal muscle cultures derived from four biopsies with a small-plaque variant of SV40 and analyzed the biological and biochemical properties of cloned myoblast derivatives. At early times after infection, myoblasts fused normally into multinucleated myotubes, and both unfused and fused cells contained SV40 tumor antigen (T antigen). After six to eight subcultures after infection, the ability of myoblasts to fuse diminished, and clonal cell lines were generated with increased growth rates and saturation densities. Transformed cultures also lost contact inhibition of growth and became anchorage independent. Unlike untransformed myoblasts, SV40-transformed clones did not undergo an increase in creatine kinase activity or a transition of creatine kinase isoenzymes from the BB form to the muscle-specific MM form. Analysis of the pattern of SV40 DNA integration by Southern blotting hybridization analysis in two cloned SV40-transformed myoblast cell lines (KJ-SV40 and PK-SV40) indicated that KJ-SV40 contained at least one site of SV40 DNA integration into chromosomal DNA and PK-SV40 contained at least three sites of SV40 DNA covalently linked to cellular DNA. Cell lysates and growth medium from PK-SV40 transformants contained infectious small-plaque variant SV40, whereas KJ-SV40 did not contain or produce detectable virus. These studies demonstrate that human myoblasts can be immortalized by SV40. This procedure may prove useful for generating large quantities of genetically deficient human cells for biochemical and molecular analysis.
Evidence for two suppressor factors secreted by a single cell suggests a solution to the J-locus paradox.
Authors: Ikezawa Z , Baxevanis C N , Arden B , Tada T , Waltenbaugh C R , Nagy Z A , Klein J
Published: May 2006

The hybridoma produced by the fusion of lactate dehydrogenase-B (LDH-B)-primed B10.A(2R) mouse suppressor T (Ts) cells with the BW5147 thymoma secretes two kinds of T suppressor factors (TsF), TsF-A and TsF-E. The TsF-A suppresses A beta-restricted and the TsF-E, E beta-restricted helper T (Th) cells. Each of the two factors consists of two polypeptide chains, an antigen-binding chain (ABC) and a major histocompatibility complex (MHC) chain. The ABC binds LDH-B, which is then recognized by one of the two receptors of the Th cell and an antigen bridge is formed between the factor and the Th cell. This chain is presumably identical in both factors. The MHC chain of the TsF-A carries antigenic determinants recognized by three sets of monoclonal antibodies: antibodies specific for the A beta chain, antibodies specific for class II determinants expressed in T cells and controlled by the A beta-E beta chromosomal segment, and antibodies crossreacting with J determinants. The MHC chain of the TsF-E carries determinants recognized by E beta-specific and by J-specific antibodies. Only some of these serologically detectable determinants reside in the region of the TsF molecule recognized by Th cells. These findings suggest that the J determinants are carried by the modified E beta and also by the modified A beta chains.
Direct stimulation of beta 2-adrenergic receptors in rat anterior pituitary induces the release of adrenocorticotropin in vivo.
Authors: Mezey E , Reisine T D , Palkovits M , Brownstein M J , Axelrod J
Published: May 2006

Previous work in our laboratory has shown that stimulation of beta 2-adrenergic receptors on mouse anterior pituitary tumor cells causes the secretion of immunoreactive adrenocorticotropin (ACTH). The present study was designed to test the hypothesis that catecholamines can cause the release of ACTH in vivo by the direct stimulation of beta 2-adrenergic receptors in the rat anterior pituitary. Systemic administration of a beta-adrenergic receptor agonist (-)-isoproterenol resulted in an increase in plasma ACTH levels in intact animals and in rats with transected pituitary stalks. This effect could be blocked by the beta-adrenergic receptor antagonist, propranolol, but not by the specific beta 1-adrenergic receptor antagonist, practolol. Salmefamol, a beta 2-adrenergic receptor agonist also elevated plasma ACTH levels in stalk-sectioned animals. Dexamethasone, a glucocorticoid that inhibits the synthesis and release of ACTH from the anterior pituitary but not the intermediate lobe, prevented the elevation of ACTH secretion by (-)-isoproterenol in stalk-transected rats. These data indicate that beta 2-adrenergic receptors are present on anterior pituitary cells and suggest that catecholamines can directly stimulate ACTH secretion.
Modifiers of mutation rate: Evolutionary optimum with complete selfing
Authors: Holsinger Kent E. , Feldman Marcus W.
Published: May 2006

It often has been assumed that, for infinite random mating populations in a constant environment, natural selection will favor genotypes at a neutral modifier locus that minimize the mutation rate. Mathematical modeling of this process confirms this assertion, independent of the selection regime. The same model under conditions of complete selfing can produce, under certain nondegenerate overdominance conditions, an optimum mutation rate below which increased mutation is favored and above which decreased mutation is favored. This occurs with unidirectional mutation models and a class of reversible mutation models with fitness overdominance. This is the first time that such a modifier optimum has been produced analytically for an infinite population in a constant environment.
Location of eight ribosomal proteins on the surface of the 50S subunit from Escherichia coli.
Authors: Stöffler-Meilicke M , Noah M , Stöffler G
Published: May 2006

Eight ribosomal proteins, L9, L11, L15, L17, L18, L19, L23, and L29, have been localized on the surface of the 50S subunit from Escherichia coli by immunoelectron microscopy. The specificity of the antibody binding site was demonstrated by stringent absorption experiments. For each protein, the antibody attachment site was localized on the two characteristic views of the 50S subunit. Thus, each protein could be located in a confined region on the three-dimensional structural model of the 50S subunit.