bluuebunny/crossref_metadata_2025 huggingface dataset:
Quantitative visualization of γ-aminobutyric acid receptors in hippocampus and area dentata demonstrated by [
3
H]muscimol autoradiography
Authors: Chan-Palay Victoria
Published: May 2006

Muscimol, a potent γ-aminobutyric acid (GABA) agonist, was used in a radioactively labeled form for the quantitative localization of GABA receptors in the rat's hippocampus (CA
1
to CA
4
) and area dentata. [
3
H]Muscimol was injected directly
in vivo
or used in the incubation medium of tissue slices, and the tissues were then fixed and prepared for autoradiography. [
3
H]Muscimol-bound GABA receptors are weakly though evenly distributed over the fimbria of the fornix. There was a laminar distribution in CA
1
to CA
4
and the area dentata, with an increasing density of the GABA receptors in that order. The lowest density was found in the alveus of CA
1
and CA
2
and the highest in the stratum granulosum of the area dentata. The greatest density was found in the neuropil between granule cells, in which are found dendrites and the basket-like plexuses of the inhibitory GABA-containing local circuit neurons. The molecular layers of the area dentata, CA
1
, and CA
2
also have a high density of GABA receptors, indicating a probable distribution over the dendrites of granule and pyramidal cells. The laminar distribution of GABA receptors in the hippocampus and area dentata is similar to the distributions of the GABA-synthesizing enzyme, glutamate decarboxylase, and of GABA previously published. Neurons with label of various density are found in the polymorphic cell layer of the area dentata, in the stratum radiatum of CA
3
, and in CA
4
. These are possibly the GABA-containing basket local circuit neurons.
Photoprocesses in covalently linked pyrochlorophyllide dimer: Triplet state formation and opening and closing of hydroxylic linkages
Authors: Periasamy N. , Linschitz H. , Closs G. L. , Boxer S. G.
Published: May 2006

Covalently linked pyrochlorophyllide
a
dimers exist in solution in either “pinned” or unfolded form, depending on the concentration of hydroxylic (H bonding) or non-H-bonding Lewis bases. Laser flash photolysis studies have been made on these dimer forms and on pyrochlorophyllide monomer at 30-nsec resolution. Spectra and recovery kinetics, including observation of both the triplet decay and ground state folding reactions, show that the flash transient obtained from the pinned form consists of a triplet and ground state moiety in the unpinned configuration. A charge-transfer excited state is not seen.
Structural polymorphism within the amino-terminal region of MM, NN, and MN glycoproteins (glycophorins) of the human erythrocyte membrane
Authors: Blumenfeld Olga O. , Adamany Anthony M.
Published: May 2006

MM, NN, and MN glycoproteins of human erythrocytes from single donors were cleaved by cyanogen bromide into three fragments—A, B, and C—which, upon gel electrophoresis, appeared to be common to the three antigens. Phenol/aqueous urea partitioning and gel filtration were used to separate the peptides quantitatively. Peptide C lacked carbohydrate and homoserine and represented the carboxyl-terminal portion of the glycoproteins. Peptides A and B contained one homoserine each and accounted for all the carbohydrate of the glycoproteins. The peptide portion of glycopeptide A from MM, NN, or MN antigens consisted of eight amino acid residues, of which six were homologous and two varied according to blood type. The variants were serine and glycine in glycopeptide A(MM), leucine and glutamic acid in A(NN), and half-residues of serine, glycine, leucine, and glutamic acid in A(MN). Serine was the amino-terminal residue in A(MM), leucine in A(NN), and one half residue of serine and leucine in A(MN). Each glycopeptide carried two tetrasaccharides (2 NANA, 1 Gal, 1 GalNAc) and one trisaccharide (NANA, Gal, GalNAc) linked
O
-glycosidically to one serine and two threonines as determined by β-elimination and sulfite addition. The carbohydrate units were attached to serine and threonine located in the invariant region, because the amino-terminal serine residue could be oxidized by periodate. The M-N antigens are believed to be products of allelic genes which are expressed exclusively in homozygotes and equimolarly in heterozygotes.
Endocytosis of cholera toxin into neuronal GERL.
Authors: Joseph K C , Kim S U , Stieber A , Gonatas N K
Published: May 2006

Cholera toxin linked covalently by glutaraldehyde to horseradish peroxidase was incubated with cultured chicken sympathetic neurons at 4 degrees. Cells were washed and brought to 37 degrees to permit endocytosis of bound toxin on plasma membranes. Massive internalization of the ligand into vesicles and cisterns of the Golgi--endoplasmic reticulum--lysosome (GERL) system was demonstrated by the cytochemical reaction for the enzyme. Surface binding and subsequent endocytosis of the cholera toxin--enzyme conjugate was inhibited when conjugate and monosialoganglioside (GM1) were simultaneously applied to cells at 4 degrees. Cholera toxin is not toxic to neurons at the levels used. These results indicate that GERL is the primary site of endocytosis of presumed complexes of cholera toxin with its plasma membrane receptor (GM1 ganglioside-containing moieties). It is suggested that, in neurons, plasma-membrane bound ligands are taken up primarily into GERL.
Liver cell-mediated mutagenesis of mammalian cells by liver carcinogens.
Authors: Langenbach R , Freed H J , Huberman E
Published: May 2006

A cell-mediated mutagenesis assay using primary cultures of rat liver cells and V79 Chinese hamster cells has been developed. Liver carcinogens and their structural analogues were studied. Mutations in the V79 cells were characterized by resistance to ouabain. Cocultivation of the liver cells and V79 cells in the presence of the carcinogens N-nitrosodimethylamine, N-nitrosodiethylamine, and aflatoxin B1 caused the induction of ouabain-resistant mutants of V79 cells. In the absence of liver cells, the carcinogens did not induce ouabain resistance. The analogues N-nitrosomethyl-tert-butylamine and aflatoxin G2 were not mutagenic. The carcinogens exhibited a dose-dependent enhancement of mutation frequency. The mutation frequency also increased with increasing numbers of liver cells seeded. It is suggested that such an experimental system may be useful for screening for chemical carcinogens.
Structure-function relationships of heparin species
Authors: Rosenberg Robert D. , Armand Gerard , Lam Lun
Published: May 2006

We have fractionated porcine heparin species of low molecular weight, with an average specific anticoagulant activity of 96 units/mg by affinity chromotography. Highly active and relatively inactive preparations of similar size were obtained with specific anticoagulant activities of 360 and 4 units/mg, respectively. The highly active heparin fraction possesses 1.1 additional residues of glucuronic acid and 1.5 fewer residues of
N
-sulfated glucosamine per molecule compared to the relatively inactive species. This decrease in
N
-sulfated glucosamine appears to be secondary to a corresponding increase in
N
-acetylated glucosamine. This form also contains a tetrasaccharide sequence with a
N
-sulfated glucosamine at its reducing end as well as equivalent amounts of glucuronic acid and iduronic acid. Furthermore, the internal glucosamine residue of this sequence appears to be
N
-acetylated. Sufficient amounts of this tetrasaccharide sequence are present within the highly active preparation such that each molecule may be endowed with this structure. The relatively inactive product contains a significantly decreased quantity of this tetrasaccharide sequence such that only [unk]20% of these molecules may possess this structure. The mean distance between nonsulfated uronic acid residues of the highly active species is smaller than that separating similar residues of the relatively inactive product. In addition, a larger number of the nonsulfated uronic acid residues of the highly active material appears either to be present in a restricted region of the molecule separated only by glucosamine residues or to be located at penultimate positions within the polysaccharide chain.
Insulin receptor: Interaction with nonreceptor glycoprotein from liver cell membranes
Authors: Maturo Joseph M. , Hollenberg Morley D.
Published: May 2006

In crude receptor preparations (either particulate or soluble) of rat liver membranes, the insulin receptor exhibits complicated binding kinetics (two binding plateaus, half-saturated at approximately 60 pM and 700 pM insulin) and an apparent chromatographic heterogeneity, suggested by the presence of two detectable, soluble insulin-binding components with apparent Stokes radii of 72 Å and 38 Å. In contrast, the insulin receptor isolated by affinity chromatography exhibits a simple binding isotherm (half-maximal saturation of binding at 700 pM insulin) without evidence for negative cooperativity and behaves as a single component (apparent Stokes radius of 38 Å) upon chromatography on Sepharose 6B. The apparent discrepancies between the properties of the unpurified insulin receptor and the affinity-purified receptor can be attributed to the presence in crude preparations of a nonreceptor constituent(s) having properties consistent with those of a membrane glycoprotein. A glycoprotein fraction from such crude soluble membrane preparations, freed from insulin receptor and subsequently partially purified using concanavalin-A-agarose, when combined with affinity-purified insulin receptor, causes both a reappearance of the complicated binding kinetics and an increase in the receptor's apparent Stokes radius from 38 Å to 72 Å. Similar results are observed for a glycoprotein fraction obtained from rat adipocyte membranes but are not observed for an identical fraction isolated from human erythrocyte membranes. We conclude that the insulin receptor in rat liver membranes can interact with another nonreceptor membrane glycoprotein that may represent either a nonrecognition moiety of the receptor oligomer or an effector molecule to the biological action of insulin.
Functional covalent complex between elongation factor Tu and an analog of lysyl-tRNA.
Authors: Johnson A E , Miller D L , Cantor C R
Published: May 2006

Complex formation between elongation factor Tu, GTP, and Nepsilon-bromoacetyl-Lys-tRNA results in the cross-linking of the protein and the modified Lys-tRNA. The efficiency of affinity labeling is greater than 50%. In the presence of unmodified Lys-tRNA, the amount of crosslinking is greatly decreased. There is no covalent reaction with elongation factor Tu in the absence of complex formation. Substantial purification of the crosslinked ternary complex can be achieved by gel filtration at low Mg2+ concentration and passage through nitrocellulose filters. The crosslinked complex exhibits message-dependent binding to ribosomes which is accompanied by the hydrolysis of the associated GTP, as shown by both filter assays and gel filtration profiles. The crosslinked complex therefore appears to function normally except for its inability to dissociate. These experiments demonstrate that the ternary complex is the true intermediate in the binding of aminoacyl-tRNA to the ribosomes.
Binding to chromosomes of herpes simplex-related antigens in biochemically transformed cells.
Authors: Kit S , Kurchak M , Wray W , Dubbs D R
Published: May 2006

Association of herpes simplex virus (HSV)-related antigens with chromosomes was demonstrated in human and mouse cells biochemically transformed by HSV that had been irradiated with ultraviolet light. This was accomplished by using peroxidase-anti-peroxidase immunological staining with rabbit antisera that had high neutralizing titers against both HSV-specific thymidine kinase activity and virus infectivity. Antisera-against HSV did not react with chromosomes of uninfected cells nor did normal sera react with any of the constitutents of biochemically transformed cells. Methanol/acetic acid treatment of biochemically transformed cells eliminated their nuclear staining for HSV-related antigens. In vitro binding of HSV-related antigens to chromosomes was demonstrated by incubating soluble antigens from high salt extracts of HSV-infected cells with methanol/acetic acid-fixed chromosomes of biochemically transformed or uninfected cells, followed by exposure to antiserum against HSV and peroxidase-anti-peroxidase staining. There was no staining when soluble extracts from uninfected cells were substituted for those from HSV-infected cells. The results show that cells biochemically transformed and lytically infected by HSV, respectively, contain antigens, which like the Epstein-Barr virus-associated nuclear antigen (EBNA), bind to chromosomes in vivo and in vitro.
Metabolic cooperation between mouse embryonal carcinoma cells and their differentiated derivatives.
Authors: Nicolas J F , Jakob H , Jacob F
Published: May 2006

Metabolic cooperation has been used as an index of intercellular communication between mouse embryonal carcinoma (EC) and other cell types. EC cells do not cooperate with differentiated cells of various properties or origins. In contrast they cooperate with cells of all other EC lines tested, including a human teratocarcinoma line, but with different efficiencies. This might reflect differences either in cell-cell interactions or in the formation of gap junctions. During in vitro differentiation, EC cells appear to remain isolated from differentiated cell types.
Different regulation of insulin receptors in intracellular (Golgi) and plasma membranes from livers of obese and lean mice.
Authors: Posner B I , Raquidan D , Josefsberg Z , Bergeron J J
Published: May 2006

Insulin binding to Golgi and plasmalemma fractions from the liver of obese (ob/bo and db/db) and lean control mice was studied. The specficiity of binding in Golgi fractions (Golgi vesicles and Golgi cisternae) was identical to that observed previously in rat liver Golgi fractions and plasmalemma. Binding to plasmalemma was much lower in obese compared to lean mice, but binding to Golgi cisternae was either not changed (ob/ob) or slightly increased (db/db) in obese compared to lean littermates. These observations suggest that intracellular (Golgi cisternae) insulin receptors are regulated differently from those of the plasmalemma. We propose that insulin affects hepatic receptor content in two different ways. On the one hand plasmalemma receptor loss is accelerated, and on the other hand intracellular receptor production is increased. This model can explain in one framework the effect on insulin in reducing and prolactin in augmenting the level of their respective hepatic receptors.
Identification of stage-specific polypeptides synthesized during murine preimplantation development.
Authors: Levinson J , Goodfellow P , vadeboncoeur M , McDevitt H
Published: May 2006

[35S]Methionine-labeled extracts of mouse ova and preimplantation embryos were analyzed by two dimensional polyacrylamide gel electrophoresis. Of the 400-600 molecular species that have been resolved as distinct spots on autoradiograms of gels for every stage of development from unfertilized eggs to early blastocysts, particular attention has been paid to the identification of 36 of these proteins, each of which is expressed only for a portion of the period under investigation. These molecules are referred to as stage-specific polypeptides and they are biochemical markers of early embryonic development and differentiation.
Tuftsin (an Ig-associated tetrapeptide) triggers the immunogenic function of macrophages: implications for activation of programmed cells.
Authors: Tzehoval E , Segal S , Stabinsky Y , Fridkin M , Spirer Z , Feldman M
Published: May 2006

The immunoglobulin heavy-chain-associated tetrapeptide, tuftsin (Thr-Lys-Pro-Arg), known for its phagocytosis-stimulating activity, was found to augment the antigen-specific, macrophage-dependent education of T lymphocytes. The investigation of stereospecific characteristics of the tetrapeptide, by use of structural analogs with different modifications, revealed strict structural requirements for eliciting the immunogenic activity of macrophages. It was found that the most important moiety for its activity is the dipeptide Pro-Arg. This finding is of interest in view of the appearance of this particular dipeptide in other bioregulatory peptides, including many of the peptide hormones. The significance of the appearance of a common structure in such molecules, which may act through specific receptors on different target cells, is discussed.
Activated human T lymphocytes display new surface glycoproteins.
Authors: Andersson L C , Gahmberg C G , Kimura A K , Wigzell H
Published: May 2006

We have analyzed the surface glycoproteins of resting and in vitro activated human T lymphocytes by the galactose oxidase/NaB3H4 and the periodate/NaB3H4 labeling techniques. The labeled glycoproteins were separated by polyacrylamide slab gel electrophoresis and visualized by fluorography. A "new" glycoprotein with an apparent molecular weight of 130,000 (GP130) was strongly labeled on alloantigen-activated T blasts but only weakly or not at all on mitogen-stimulated T blasts and resting T lymphocytes. These results demonstrate that human T cells, as earlier found in the mouse system, express different surface molecules in relation to the particular mode of activation and stage of differentiation.
Internal charge transfer in proteins to the Schiff bases of their lysine side chains
Authors: Otto Peter , Ladik János , Laki Koloman , Szent-Györgyi Albert
Published: May 2006

Stereochemical investigations carried out with the aid of molecular models show that the Schiff bases formed between methylglyoxal and the primary amino groups of the lysine side chains of proteins can bend back in such a way to the N atoms of the peptide groups that a charge transfer can occur between these groups and the oxygen atoms of the Schiff bases.
Ab initio
molecular orbital calculations support this finding.
Inactivation of phage repressor in a permeable cell system: role of recBC DNase in induction.
Authors: Oishi M , Smith C L
Published: May 2006

UV light causes inactivation of phage (phi80) repressor molecules in a plasmolyzed, permeable cell preparation of Escherichia coli. Induction without UV irradiation occurs when the permeable cells are incubated in the presence of four deoxyribonucleoside triphosphates and ATP. The induction triggered by dNTP's requires a functional recBC gene product and is associated with degradation of the DNA replication fork. The role of recBC DNase in the induction of prophage and SOS functions in general is discussed.
Amino acid sequence of tyrosinase from Neurospora crassa.
Authors: Lerch K
Published: May 2006

The amino-acid sequence of tyrosinase from Neurospora crassa (monophenol,dihydroxyphenylalanine:oxygen oxidoreductase, EC 1.14.18.1) is reported. This copper-containing oxidase consists of a single polypeptide chain of 407 amino acids. The primary structure was determined by automated and manual sequence analysis on fragments produced by cleavage with cyanogen bromide and on peptides obtained by digestion with trypsin, pepsin, thermolysin, or chymotrypsin. The amino terminus of the protein is acetylated and the single cysteinyl residue 96 is covalently linked via a thioether bridge to histidyl residue 94. The formation and the possible role of this unusual structure in Neurospora tyrosinase is discussed. Dye-sensitized photooxidation of apotyrosinase and active-site-directed inactivation of the native enzyme indicate the possible involvement of histidyl residues 188, 192, 289, and 305 or 306 as ligands to the active-site copper as well as in the catalytic mechanism of this monooxygenase.
Mechanism of integrating foreign DNA during transformation of Bacillus subtilis.
Authors: Duncan C H , Wilson G A , Young F E
Published: May 2006

Genes encoding thymidylate synthetase from Bacillus subtilis bacteriophages were cloned in Escherichia coli. Chimeric plasmids pCD1 and pCD3 were constructed from site-specific endonuclease digests of bacteriophage phi3T DNA cloned in pMB9 in E. coli. Similar cloning techniques with bacteriophage beta22 DNA yielded chimeric plasmids pCD4, pCD5, and pCD6. Endonuclease digests of DNA from pCD1 and pCD3 propagated in E. coli or from DNA isolated from bacteriophage phi3T propagated in B. subtilis transformed B. subtilis from Thy- to Thy+. Intact DNA from bacteriophage beta22, endonuclease digests of beta22 DNA, and a chimeric plasmid (pCD5) composed only of the thybeta22 gene and pMB9 did not transform B. subtilis from Thy- to Thy+ even though pCD5 could transform Thy- E. coli to Thy+. However, if the thybeta22 fragment from pCD5 was introduced into another chimeric plasmid, pCD2, that contains a region of homology to the chromosome of B. subtilis in addition to pMB9, transformation of Thy- clones of B. subtilis was possible. Furthermore, Southern hybridization analyses of the digests of chromosomal DNA from the Thy+ transformants established that the entire chimeric plasmid was incorporated into the chromosome of B. subtilis. Treatment of these plasmids with site-specific endonucleases abolished transformation. These results indicated that the entire chimeric plasmid can be incorporated into the chromosome of B. subtilis by a Campbell-like model. Therefore, an additional mechanism for transformation exists whereby plasmids can be integrated if sufficient chromosomal homology is maintained.
The ovalbumin gene: Cloning of the natural gene
Authors: Woo Savio L. C. , Dugaiczyk Achilles , Tsai Ming-Jer , Lai Eugene C. , Catterall James F. , O'Malley Bert W.
Published: May 2006

The structural ovalbumin DNA sequences are not contiguous and are separated by multiple “intervening regions” in native chicken DNA.
Eco
RI, a restriction endonuclease that does not cleave the structural ovalbumin DNA sequences, digests the natural ovalbumin gene into three distinct fragments of 2.4, 1.8, and 9.5 kilobase pairs in length by cleaving within these “intervening regions.” The 2.4-kilobase pair fragment contains only about 450 nucleotide pairs of coding sequence, with the rest being intervening sequences. This DNA fragment was cloned in bacteria by using the certified EK2 vector λgtWES·λB after enrichment from total
Eco
RI-digested chicken DNA by a combination of RPC-5 column chromatography and preparative agarose gel electrophoresis. Five out of approximately 20,000 recombinant phage plaques were capable of hybridizing with a
32
P-labeled
Hha
I fragment of a recombinant plasmid pOV230 containing the entire structural ovalbumin gene. DNA amplified in these recombinant phages, λgtWES·OV2.4, was shown to contain the same restriction endonuclease cleavage sites as in the 2.4-kilobase pair
Eco
RI fragment previously determined by restriction mapping of total genomic chicken DNA. The intervening sequences were allowed to hybridize with excess total chicken DNA and oviduct nuclear RNA after nick-translation. They were found to be unique chicken DNA sequences, and appeared to be transcribed in their entireties during gene expression. Like the structural gene sequences, the expression of the intervening sequences is also inducible by steroid hormones.
Detection of an early surface change during oncogenic transformation.
Authors: Parry G , Hawkes S P
Published: May 2006

Fluorescamine, which can label surface components of cells grown as monolayers in culture, has been used to probe alterations in chicken embryo fibroblasts infected with a temperature-sensitive mutant of Rous sarcoma virus, Prague A, LA24. The fluorescence of bound fluorescamine on cells at the permissive temperature (35 degrees) was found to be about 1/3 that of cells cultured at the nonpermissive temperature (41 degrees). During development of the transformed phenotype, i.e., after transfer of the cells from 41 degrees to 35 degrees, the decrease in surface fluorescence was observed to be an early event occurring within the first 4-8 hr after temperature shift. This alteration took place on a time scale similar to that of changes in 2-deoxyglucose transport and an increased rate of DNA synthesis, but before any major morphological changes. The change was related to cell transformation rather than to growth differences of the cells at the two temperatures. Further, it was found that fluorescamine was not monitoring the loss of LETS glycoprotein from the surface or the loss of any other surface components that could be detected by lactoperoxidase-catalyzed iodination of surface proteins. When fluorescamine-labeled components were resolved by polyacrylamide gel electrophoresis, significant differences were seen between components from cells cultured at 35 degrees compared with those from cells cultured at 41 degrees. Based on these results, possible mechanisms accounting for the fluorescence differences are suggested.
Calcification of isolated matrix vesicles and reconstituted vesicles from fetal bovine cartilage.
Authors: Hsu H H , Anderson H C
Published: May 2006

Ca deposition by isolated matrix vesicles from fetal calf growth plate cartilage and by a deoxycholate extract from matrix vesicles that included their phosphatase was studied under defined in vitro conditions. Electron microscopy showed that after removal of deoxycholate and lyophilization of the vesicle extract, new vesicles were reconstituted, often with multiple membrane layers. Both intact calf vesicles and reconstituted vesicles initiated Ca deposition maximally when supplied with ATP, GTP, CTP, or UTP. Only nucleoside triphosphates supported Ca deposition well; mono- and diphosphoesters, although hydrolyzed, were ineffective as substrates. Nucleoside triphosphates supported Ca deposition even if the final [Ca] X [P] reached in the reaction mixture was below a metastable level (3.5 mM2), suggesting that matrix vesicles or reconstituted vesicles promote calcification by localizing Ca or PO4 or both. ATP or GTP supported Ca deposition readily at concentrations ranging from 0.25 to 1.0 mM but, at 2.5 and 5.0 mM, Ca deposition was inhibited. The ATPase of intact matrix vesicles and reconstituted vesicles was stimulated by addition of Ca2+ and Mg2+. Ca deposition did not require additional Mg2+. These results lend support to the hypothesis that matrix vesicles and their phosphatases play an important role in mineralization.
Fluorescent light-induced DNA crosslinkage and chromatid breaks in mouse cells in culture.
Authors: Gantt R , Parshad R , Ewig R A , Sanford K K , Jones G M , Tarone R E , Kohn K W
Published: May 2006

A single 20-hr exposure of mouse cells derived from embryonic or lung tissue to cool-white fluorescent light (4.6 W/m2) causes both DNA damage and chromosome aberrations including chromatid breaks, exchanges, and minutes. In Kohn's alkaline elution technique, the DNA from exposed cells elutes more slowly than that from shielded cells. Because larger molecular weight DNA elutes slower than smaller, we interpret these results to mean that the DNA in cells exposed to light is crosslinked. The estimated frequency of crosslinks is sufficient to account for the number of chromatid breaks observed. The types of chromosome aberrations produced by light indicate that the primary lesion results in chromatid rather than chromosome breaks, and the results suggest an influence of cell density in that cells in densely populated cultures showed few or no chromatid breaks after irradiation. The present results, together with observations from the literature, suggest that the DNA crosslinkage and the chromosome aberrations produced by light may be related.
Inhibition of dimethyl sulfoxide-stimulated Friend cell erythrodifferentiation by hydrocortisone and other steroids.
Authors: Scher W , Tsuei D , Sassa S , Price P , Gabelman N , Friend C
Published: May 2006

Erythrodifferentiation and hemoglobin synthesis in dimethyl sulfoxide-stimulated Friend erythroleukemia cells were inhibited by hydrocortisone (HC) and four other steroids: dexamethasone, deoxycorticosterone, corticosterone, and aldosterone. The effect was specific, because no significant cytotoxicity occurred with any of these compounds at the concentrations that were inhibitory. The mechanism of action of HC was studied in detail. In the absence of dimethyl sulfoxide, it had no effect on hemoglobin levels; but, in the presence of this inducer, the synthesis of heme and globin were each inhibited by approximately 90%. There was no alteration in the synthesis of any major protein other than globin, as determined by gel electrophoresis of cell lysates. The activities of two enzymes in the heme biosynthetic pathway, delta-aminolevulinate dehydratase and uroporphyrinogen-I synthase, were inhibited by 80% and 70%, respectively. Globin mRNA induction was reduced by approximately 90%. This demonstrated that the HC inhibition of globin synthesis occurred at a pretranslational step. The dimethyl sulfoxide-induced single-stranded breaks in DNA, which have been suggested to play a role in Friend leukemia cell differentiation, were reduced in number but not eliminated. HC reduced the dimethyl sulfoxide-stimulation of virus release into the medium by approximately 50%. HC treatment in the absence of dimethyl sulfoxide doubled the production of virus.
Calcitonin and calcium ionophores: cyclic AMP responses in cells of a human lymphoid line.
Authors: Moran J , Hunziker W , Fischer J A
Published: May 2006

Receptors for calcitonin, as assayed by the specific binding of 125I-labeled salmon calcitonin and stimulation of cyclic AMP formation, were found in 8866 cells derived from a human lymphoid line. The affinity of calcitonin from different species and of various analogues of human calcitonin for the binding sites and their ability to stimulate cyclic AMP formation were closely related to their hypocalcemic activity and presumably reflected biological properties of the hormones. Besides calcitonin, prostaglandin E1 and beta-adrenergic catecholamines stimulated cyclic AMP formation in these cells. The calcium ionophores, A23187 and Br-X-573A, did not influence the specific binding of 125I-labeled salmon calcitonin. A23187, however, suppressed basal and calcitonin-stimulated formation of cyclic AMP in the presence of at least 0.6 mM calcium in the incubation medium. Br-X-537A did not require extracellular calcium to suppress basal and calcitonin-stimulated formation of cyclic AMP, suggesting that the release of calcium from internal stores may regulate adenylyate cyclase activity in 8866 cells.
High molecular weight opioid-containing proteins in striatum.
Authors: Lewis R V , Stein S , Gerber L D , Rubinstein M , Udenfriend S
Published: May 2006

Striatal extracts of guinea pigs, rats, and cattle were found to contain two large proteins (greater than 40,000 and greater than 100,000 daltons) that on treatment with trypsin yield opioid peptides differing chromatographically from the opioid nonapeptide generated by trypsin digestion of endorphins, beta-lipotropin, or pro-opiocortin. Furthermore, the large opioid proteins found in the pituitary do not appear to be present in the striatum. These and other findings indicate that the striatal enkephalins are produced via a pathway from the one deduced from studies on the pituitary.
Characterization of the blue copper site in oxidized azurin by extended x-ray absorption fine structure: Determination of a short Cu—S distance
Authors: Tullius Thomas D. , Frank Patrick , Hodgson Keith O.
Published: May 2006

The primary coordination environment of the “blue” copper ion in oxidized azurin has been elucidated by x-ray absorption spectroscopy. The most striking feature is the unambiguous presence of a very short copper-sulfur distance at 2.10 ± 0.02 Å. Nitrogen ligands, presumed to be from imidazoles, are found at 1.97 Å. There is some evidence that the copper coordination sphere may be completed by a second sulfur, the distance of which is determined with much less certainty.
Modification of DNA by aflatoxin B1 creates alkali-labile lesions in DNA at positions of guanine and adenine.
Authors: D'Andrea A D , Haseltine W A
Published: May 2006

The damage to DNA by the hepatocarcinogen aflatoxin B1 was investigated. A DNA fragment of known sequence of the lactose promoter-operator region was used as a substrate for modification by aflatoxin B1. The DNA was incubated with aflatoxin B1 in crude mammalian liver extracts or with purified microsomes. Treatment of the DNA incubated in the complete system with either 1 M piperidine or 0.1 M NaOH at 90 degrees revealed alkali-labile lesions in the DNA. The exact location of the cleavage site was determined by comparison of the length of the cleavage products with the known sequence on polyacrylamide gels. The lengths of the cleavage products were the same as those produced by alkali-induced breakage of the same sequence of DNA that had been modified with dimethyl sulfate. The major cleavage products of the aflatoxin B1-modified DNA were at positions of guanine and the minor cleavage products were at positions of adenine. These studies show that modification of DNA by aflatoxin B1 creates alkali-labile sites at positions of guanine and, to a lesser extent, adenine.
Electron microscopic heteroduplex analysis of "killer" double-stranded RNA species from yeast.
Authors: Fried H M , Fink G R
Published: May 2006

Wild-type and mutant double-stranded RNA (dsRNA) species from the yeast Saccharomyces cerevisiae were studied by electron microscopic heteroduplex mapping to determine the sequence relationships among the different RNA molecules. Three mutant dsRNAs, 1.5, 1.4, and 0.73 kilobase, were found to be derived by the same internal deletion of the wild-type (I83 kilobases) molecule. This deletion includes a wild-type (1.83 kilobases) molecule. This deletion includes a segment of about 200 base pairs that was estimated to be nearly 100% A+U. In addition, the sequences of the two larger mutant RNA species are tandem, direct duplications. One of the duplicated molecules appears to have a second internal deletion that occurred after the duplication. The mutant dsRNAs are functionally similar to the defective interfering virus particles of animal viruses--all of the mutant species prevent the propagation of the wild-type dsRNA when both are present in the same cell. The four dsRNAs share the same sequences at their termini, a finding that may suggest that these sequences are important for the replication of the dsRNAs.
High concentration of thyrotropin-releasing hormone in pancreatic islets.
Authors: Martino E , Lernmark A , Seo H , Steiner D F , Refetoff S
Published: May 2006

The concentration of thyrotropin-releasing hormone (TRH, thyroliberin) in rat islets of Langerhans is 30-fold higher than in whole rat pancreas, indicating that the islets are the main source of pancreatic TRH. The TRH extracted from islets is indistinguishable from synthetic TRH in its immunological and biological properties and in its inactivation by human serum. The physiologic function of islet TRH is unknown. However, because TRH is antagonistic to somatostatin in other systems, and somatostatin also is concentrated in islets in high concentrations, it is possible that islet TRH may serve a similar antagonistic function in the regulation of islet cell secretory activity.
Nucleotide sequence of the leader region of the phenylalanine operon of
Escherichia coli
Authors: Zurawski Gerard , Brown Keith , Killingly David , Yanofsky Charles
Published: May 2006

The
pheA
structural gene of the phenylalanine operon of
Escherichia coli
is preceded by a transcribed leader region of about 170 nucleotide pairs.
In vitro
transcription of plasmids and restriction fragments containing the
phe
promoter and leader region yields a major RNA transcript about 140 nucleotides in length. This transcript,
pheA
leader RNA, has the following features: (
i
) a potential ribosome binding site and AUG translation start codon about 20 nucleotides from its 5′ end; (
ii
) 14 additional in phase amino acid codons and a UGA stop codon after the AUG; 7 of these 14 are Phe codons; (
iii
) a 3′-OH terminus about 140 nucleotides from the 5′ end (transcription termination occurs in an A·T-rich region which is subsequent to a G·C-rich region; just beyond the site of transcription termination there is a sequence corresponding to a ribosome binding site and the AUG translation start codon of the
pheA
structural gene); (
iv
) a sequence which would permit extensive intrastrand stable hydrogen bonding. In addition to G·C-rich stem structures, highly analogous to those proposed for the leader RNAs of the tryptophan operons of
E. coli
and
Salmonella typhimurium
[Lee, F. & Yanofsky, C. (1977)
Proc. Natl. Acad. Sci. USA
74, 4365-4369], there is also extensive base-pairing possible between the
phe
codon region and a more distal region of the leader transcript. The roles of synthesis of the Phe-rich leader peptide and secondary structure of the leader transcript in the regulation of transcription termination at the attenuator of the
phe
operon are discussed.
Synthesis of catalase in two cell-free protein-synthesizing systems and in rat liver
Authors: Robbi Mariette , Lazarow Paul B.
Published: May 2006

Rat liver polysomal RNA was translated in the rabbit reticulocyte lysate and in the wheat germ cell-free protein-synthesizing systems, using [
35
S]methionine as label. The catalase (hydrogen-peroxide:hydrogen-peroxide oxidoreductase, EC 1.11.1.6) that was synthesized was isolated by immunoprecipitation and characterized by electrophoresis in sodium dodecyl sulfate/polyacrylamide gels followed by fluorography. The catalase made in both systems migrated more slowly during electrophoresis than did purified peroxisomal catalase. By comparison with standards of known molecular mass, the cell-free products were estimated to be about 4000 daltons larger than the purified enzyme. We also investigated the biosynthesis of catalase
in vivo
by injecting [
35
S]methionine into rats. The precursor of catalase known to be synthesized in liver and found in the high-speed supernatant 8 min later [Lazarow, P. B. & de Duve, C. (1973)
J. Cell Biol.
59, 491-506] was isolated immunochemically. For comparison, 1-day-old completed catalase was immunoprecipitated from peroxisomes. The migrations in sodium dodecyl sulfate gels of the 8-min-old precursor and the subunit of the day-old enzyme were indistinguishable and approximately the same as the migration of the cell-free products. These results indicate that catalase's apparent size does not change when it enters peroxisomes but rather decreases during the chemical purification procedure.
Simian virus 40 A gene function: DNA content analysis of Chinese hamster cells transformed by an early temperature-sensitive virus mutant.
Authors: Robinson C C , Lehman J M
Published: May 2006

Replication of two Chinese hamster embryo cell lines transformed by an early temperature-sensitive mutant of simian virus 40, tsA58, was examined by flow microfluorometry and autoradiography of [3H]thymidine-labeled cells in order to determine whether transformed cell DNA synthesis is initiated by the virus A gene. At the permissive temperature (37 degrees), cells transformed by the mutant were like the wild-type virus transformants in appearance, colony-forming ability, high saturation density, and rapid replication. At the nonpermissive temperature (40.5 degrees), the tsA58 transformed cells resembled normal embryo fibroblasts and seem to return to normal growth patterns. Although both mutant transformed cell lines at 40.5 degrees appeared to cease growth at low saturation density, the cells did not enter a resting state, but continued to replicate. The cultures were maintained at low densities by a balance among cell replication, cell death, and sloughing of dead cells into the supernatant. These results suggest that the simian virus 40 A gene function effected by the tsA58 mutation does not prevent Chinese hamster embryo transformed cells from entering a resting state, although the gene may control other phenotypic characteristics of transformation.
Synapses involving auditory nerve fibers in primate cochlea.
Authors: Bodian D
Published: May 2006

The anatomical mechanisms for processing auditory signals are extremely complex and incompletely understood, despite major advances already made with the use of electron microscopy. A major enigma, for example, is the presence in the mammalian cochlea of a double hair cell receptor system. A renewed attempt to discover evidence of synaptic coupling between the two systems in the primate cochlea, postulated from physiological studies, has failed. However, in the outer spiral bundle the narrow and rigid clefts seen between pairs of presumptive afferent fibers suggest the possibility of dendro-dendritic interaction confined to the outer hair cell system. The clustering of afferent processes within folds of supporting cells subjacent to outer hair cells is in contrast to the lack of such close associations in the inner hair cell region. The difference reinforces the suggestion of functional interaction of some sort between the outer hair cell afferent nerve processes.
Low and high density lipoproteins and chylomicrons as regulators of rate of cholesterol synthesis in rat liver
in vivo
Authors: Andersen John M. , Turley Stephen D. , Dietschy John M.
Published: May 2006

The steady-state levels of plasma cholesterol carried in high and low density lipoproteins and in chylomicrons were varied over a wide range by use of a constant-infusion technique. After 40 hr, the rates of hepatic cholesterol synthesis and levels of hepatic cholesterol esters were measured and were related to the plasma level of each of the lipoprotein fractions. From the rates of infusion and the steady-state plasma levels attained, the whole animal clearance rates for cholesterol carried in low density and high density lipoproteins and in chylomicrons were calculated to be 0.53, 0.61, and 42.6 ml/hr, respectively. Hepatic cholesterol ester content increased by 0.8 μg/g for each 1.0 mg/dl increase in the steady-state level of plasma low density lipoprotein cholesterol and by 1.4 μg/g for a similar elevation in plasma high density lipoprotein cholesterol. In contrast, the increase in ester content was 300-fold greater when chylomicrons were infused (330 μg/g). The rate of hepatic cholesterol synthesis was inhibited by a factor of 0.004 and 0.007, respectively, per 1.0 mg/dl increase in the steady-state level of plasma cholesterol carried in either low density or high density lipoprotein but the inhibition was by a factor of 0.255, 50-fold greater, when chylomicrons were infused. Thus, in the steady state, cholesterol carried in either low density or high density lipoproteins is apparently taken up by the liver and regulates the rate of hepatic cholesterol synthesis; however, cholesterol carried in chylomicrons is at least 50-fold more effective in this regard. This marked difference can be attributed to the much higher rates of transport of chylomicron cholesterol into the hepatocyte than of cholesterol carried in either low density or high density plasma lipoproteins.
Steroid diamine-nucleic acid interactions: partial insertion of dipyrandium between unstacked base pairs of the poly(dA-dT) duplex in solution.
Authors: Patel D J , Canuel L L
Published: May 2006

We report on an NMR investigation of steroid diamine.nucleic acid complexes as a function of phosphate-to-drug ratios in aqueous solution in order to evaluate the structural and kinetic aspects of the binding of a nonintercalative drug to a synthetic DNA in solution. The nonexchangeable proton chemical shift parameters for the dipyrandium.poly(dA-dT) complex demonstrate unstacking of base pairs and partial insertion of the steroid diamine at the complexation site. The chemical shifts and linewidths of the exchangeable protons as a function of pH demonstrate that the base pairs are intact but partially exposed to solvent at the steroid diamine binding site. The phosphorus chemical shifts suggest that the base pairs unstack upon complex formation without changes in the omega,omega' polynucleotide backbone torsion angles. The NMR line shape parameters require rapid exchange of the steroid diamine among potential binding sites and are consistent with greater segmental flexibility in the complex compared to the synthetic DNA in solution. The NMR experiments are discussed in relation to Sobell's proposed model [Sobell, H.M., Tsai, C.C., Gilbert, S.G., Jain, S.C. & Sakore, T.D. (1976) Proc. Natl. Acad. Sci USA 73, 3068-3072] for the steroid diamine-DNA complex.
Regulation of alkaline phosphatase expression in human choriocarcinoma cell lines.
Authors: Hamilton T A , Tin A W , Sussman H H
Published: May 2006

The coincident expression of two structurally distinct isoenzymes of human alkaline phosphatase was demonstrated in two independently derived gestational choriocarcinoma cell lines. These proteins were shown to have enzymatic, antigenic, and physical-chemical properties resembling those of isoenzymes from term placenta and adult liver. The regulation of these isoenzymes has been studied during the exposure of both cell lines to 5-bromodeoxyuridine and dibutyryl cyclic AMP. The responses of the alkaline phosphatase isoenzymes to these agents have also been compared with the response of another protein phenotypic to placenta, the alpha subunit of chorionic gonadotropin. The results show that (i) the separate structural genes coding for placental and liver alkaline phosphatases are regulated in a noncoordinate fashion; (ii) both alkaline phosphatase genes respond independently of the alpha subunit; and (iii) the induction of the placental type isoenzyme occurs via at least two independent pathways.
Monoclonal antibodies reacting with murine teratocarcinoma cells.
Authors: Goodfellow P N , Levinson J R , Williams V E , McDevitt H O
Published: May 2006

Monoclonal antibodies were produced in vitro by fusing mouse myeloma cells with spleen cells from a rat immunized with the C3H mouse teratocarcinoma C86-S1. After the fusion two clones were chosen for further analysis. The first clone, 3C4-10, produced an antibody recognizing an antigen with a distribution restricted to teratocarcinoma cell lines, an endoderm cell line, and a neuroblastoma. The second clone, 4A1-9, produced an antibody that reacted with all cultured murine cells tested and adult brain. Neither antibody reacted with preimplantation embryos. The 3C4-10 antibody recognized an antigen associated with proteins. The apparent molecular weight of the 3C4-10 antigen was greater than 100,000.
Interaction of peanut agglutinin with normal human lymphocytes and with leukemic cells.
Authors: Reisner Y , Biniaminov M , Rosenthal E , Sharon N , Ramot B
Published: May 2006

The interaction of peanut agglutinin (PNA) with human thymocytes, peripheral blood lymphocytes, and peripheral blood cells of various types of leukemia was investigated by using fluorescein isothiocyanate-conjugated PNA. The majority of human thymocytes (60-80%) bind the lectin. The major subpopulation of thymocytes that is PNA-positive was separated from the PNA-negative cells by differential agglutination with the lectin. The two thymocyte subpopulations were tested in the mixed lymphocyte reaction and with the phytohemagglutinin of Phaseolus vulgaris. The poor response of the PNA-positive thymocytes to these stimuli indicates that these thymocytes are functionally immature. The fluorescein isothiocyanate-PNA-binding test with peripheral blood lymphocytes of leukemic patients revealed that in most acute leukemias the PNA receptor is exposed on the blastic cells, whereas in most cases of chronic leukemia the peripheral blood lymphocytes are PNA-negative. The validity of PNA as a marker of immature blood cells and its potential clinical application are discussed.
Distribution of binding sites for 125I-labeled alpha-bungarotoxin in normal and deafferented antennal lobes of Manduca sexta.
Authors: Hildebrand J G , Hall L M , Osmond B C
Published: May 2006

125I-Labeled alpha-bungarotoxin has been used to determine the distribution of putative acetylcholine receptors in normal and chronically deafferented antennal lobes in the brain of the moth Manduca sexta. Toxin-binding sites are confined to synaptic regions in deafferented lobes. These findings suggest that receptors can develop in the insect central nervous system independently of normal synaptic influences.