bluuebunny/crossref_metadata_2025 huggingface dataset:
(E)-5-(2-Bromovinyl)-2'-deoxyuridine: a potent and selective anti-herpes agent.
Authors: De Clercq E , Descamps J , De Somer P , Barr P J , Jones A S , Walker R T
Published: May 2006

Of a series of five newly synthesized 2'-deoxyuridine derivatives, including 5-vinyl-dUrd, 5-ethynyl-dUrd, 5-(1-chlorovinyl)-dUrd, (E)-5-(2-bromovinyl)-dUrd, and (E)-5-(2-iodovinyl)-dUrd, the last two compounds were found to exert a marked inhibitory effect on the replication of herpes simplex virus type 1 [ID50 (mean inhibitory dose), 0.004-0.02 microgram/ml]. Both (E)-5-(2-bromovinyl)-dUrd and (E)-5-(2-iodovinyl)-dUrd were highly selective in their anti-herpes activity in that they did not affect the growth or metabolism of the host (primary rabbit kidney) cells unless drug concentrations were used that were 5,000- to 10,000-fold greater than those required to inhibit virus multiplication. In this sense (E)-5-(2-bromovinyl)-dUrd and (E)-5-(2-iodovinyl)-dUrd proved more selective in their activity against herpes simplex virus type 1 than all other anti-herpes compounds that have been described so far. In animal model systems (namely, cutaneous herpes infections of athymic nude mice), (E)-5-(2-bromovinyl)-dUrd suppressed the development of herpetic skin lesions and mortality therewith associated, whether the compound was administered topically or systemically. Under the same conditions, the standard anti-herpes drug 5-iodo-dUrd (Idoxuridine) offered little, if any, protection. Although the precise mechanism of action of (E)-5-(2-bromovinyl)-dUrd and (E)-5-(2-iodovinyl)-dUrd remains to be established, preliminary findings indicate that they do not specifically act at the thymidylate synthetase step.
Physical map of the recA gene.
Authors: Sancar A , Rupp W D
Published: May 2006

We have cloned the recA gene of Echerichia coli K12 and some of its restriction fragments on the plasmid cloning vehicle pBR322. The recA gene was mapped with regard to the restriction sites of EcoRI, BamHI, Pst I, Hha I, Hae III, HinfI, and Taq I restriction endonucleases. The recA promoter was localized by the binding of RNA polymerase to restriction fragments. The initiation point of transcription of recA mRNA and the direction of transcription were determined from in vitro transcription of recA gene fragments and from analysis of the polypeptides made in maxicells that contain plasmids carrying only part of the recA gene.
Purification and distribution of a major protein in rat prostate that binds estramustine, a nitrogen mustard derivative of estradiol-17 beta.
Authors: Forsgren B , Björk P , Carlström K , Gustafsson J A , Pousette A , Högberg B
Published: May 2006

A protein in rat ventral prostate cytosol that binds estramustine [estradiol 3-bis(2-chloroethyl)carbamate) was purified to homogeneity by using chromatography on DEAE-cellulose, Sephadex G-100 (superfine), octyl-Sepharose (CL-4B, and polyacrylamide gel electrophoresis. The estramustine-binding protein was found to have a Mr of 46,000 as estimated by gel filtration. After analysis by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, the protein was found to consist of two subunits with Mr of about 20,000 and 18,000. After reduction of disulfide bridges, the protein was decomposed into three components with Mr of about 12,000, 11,000, and 8000. Amino acid analysis indicated that the protein is a glycoprotein. Antibodies against the protein were raised in rabbits and a radioimmunoassay was developed for it. The estramustine-binding protein constituted about 18% of the total protein in rat ventral prostate cytosol, was present in the dorsal and lateral lobes of the prostate, and was also detected in the pituitary gland, cerebral cortex, submaxillary gland, thyroid gland, adrenal gland, seminal vesicle, coagulating gland, epididymis, and preputial gland of the male rat. In female rats the protein was detected in cerebral cortex. Because the estramustine-binding protein is predominantly found in the accessory sexual glands of the male rat, it may be of importance for maintaining male fertility.
beta-Endorphin: synthesis of analogs modified at the carboxyl terminus with increased activites.
Authors: Li C H , Yamashiro D , Tseng L F , Chang W C , Ferrara P
Published: May 2006

Three analogs of human beta-endorphin (beta h-EP) have been synthesized: [Gly31]beta h-EP, [Gly31]beta h-endorphinamide, and [Gly31]beta h-endorphinylglycine. All are more active than beta h-EP in both the guinea pig ileum bioassay and the opiate receptor binding assay. The last two analogs are about twice as active as beta h-EP in an assay for analgesia. Modification at position 31 and extension at the COOH terminus may afford a route toward analogs with even greater biological activity.
Ganglioside inhibition of fibronectin-mediated cell adhesion to collagen.
Authors: Kleinman H K , Martin G R , Fishman P H
Published: May 2006

Fibronectin mediates the adhesion of cells to collagen by first binding to the collagen substrate, followed by attachment of the cells to the fibronectin-collagen complex. Bovine brain gangliosides were found to block fibronectin-mediated cell adhesion to collagen in a concentration-dependent manner. The gangliosides did not block the binding of fibronectin to collagen but did prevent the attachment of the cells to the fibronectin-collagen complex. Of the individual gangliosides tested, GT1 and GD1a were the most effective inhibitors followed by GD1b greater than GM1 greater than GM2; GM3 was not an inhibitor. The inhibition of cell adhesion also was observed with the oligosaccharide portion of the gangliosides, but not with ceramides or with a variety of free sugars or glycosaminoglycans. Mild periodate oxidation of mixed gangliosides or of GD1a modified their sialic acid residues and the oxidized gangliosides were no longer inhibitory; subsequent reduction with NaBH4 did not restore the inhibitory activity of the modified gangliosides. These results suggest that specific gangliosides or related sialic acid-containing glycoconjugates on the cell surface may act as the receptors for fibronectin.
Transport of tricarboxylic acid cycle intermediates by membrane vesicles from renal brush border.
Authors: Kippen I , Hirayama B , Klinenberg J R , Wright E M
Published: May 2006

The uptake of citrate and alpha-ketoglutarate by membrane vesicles from rabbit renal brush border was studied by a rapid filtration technique. Both compounds exhibited transport characteristics similar to those seen for the sodium-dependent cotransport systems previously described for sugars and amino acids in brush border membranes. The estimated sodium-dependent Vmax and Km were 17 nmol per mg of protein per min and 0.18 mM for citrate and 17 nmol per mg of protein per min and 1.0 mM for alpha-ketoglutarate. The initial rate of citrate transport was 5 times that of sugars and amino acids under comparable conditions. Uptake rates of 0.1 mM citrate and alpha-ketoglutarate were inhibited by greater than 90% by 10 mM succinate, malate, fumarate, or oxaloacetate, indicating the presence in the brush border membrane of a transport system highly specialized for the renal conservation of intermediates of the tricarboxylic acid cycle.
Tyrosine administration reduces blood pressure and enhances brain norepinephrine release in spontaneously hypertensive rats.
Authors: Sved A F , Fernstrom J D , Wurtman R J
Published: May 2006

Administration of L-tyrosine to normotensive or spontaneously hypertensive rats reduces blood pressure. The effect is maximal within 2 hr of injection. In spontaneously hypertensive rats, a dose of 50 mg/kg, intraperitoneally, reduces blood pressure by about 12 mm Hg (1 mm Hg = 1.33 x 10(2) pascals); a dose of 200 mg/kg produces the maximal effect, a reduction of about 40 mm Hg. Tryptophan injection (225 mg/kg) also lowers blood pressure in spontaneously hypertensive rats, but only by about half as much as an equivalent dose of tyrosine. Other amino acids tested (leucine, isoleucine, valine, alanine, arginine, and aspartate) do not affect blood pressure. Tyrosine injection appears to reduce blood pressure via an action within the central nervous system, since the effect can be blocked by co-administering other large neutral amino acids that reduce tyrosine's uptake into the brain. That tyrosine's antihypertensive action is mediated by an acceleration in norepinephrine or epinephrine release within the central nervous system is suggested by the concurrent increase that its injection produces in brain levels of methoxyhydroxyphenylethylglycol sulfate.
Detection of glucocorticoid receptors on Friend erythroleukemia cells.
Authors: Golde D W , Bersch N , Lippman M E , Friend C
Published: May 2006

The inhibition of dimethyl sulfoxide-induced differentiation of Friend erythroleukemia cells by steroids led us to examine these cells for the presence of glucocorticoid receptors. Direct assessement of dexamethasone binding revealed high-affinity dexamethasone receptors on the untreated cells. The specific binding of [3H]dexamethasone was dose dependent. At a concentration of 10(-8) M, almost all binding sites were occupied. The mean number of binding sites per cell in two separate experiments was 8045 and 7191, respectively, and the Kd varied between 3.38 and 3.49 x 10(-9) M. Dimethyl sulfoxide treatment led to a decrease in the number of dexamethasone binding sites on the cells induced to differentiate. After 5 days of treatment, the mean number of sites per cell was reduced to 1216 and 896 in two experiments, with a Kd of 5 x 10(-9) M. Dexamethasone treatment resulted in a moderate decrease in the efficiency of colony formation within 72 hr after the cells were plated in methylcellulose. The mechanism of this inhibitory effect is unknown. However, it was also dose dependent and could be abrogated by appropriate concentrations of progesterone or 11-deoxycortisone. These results suggest that the steroid effects on growth and differentiation of the erythroleukemia cells may be mediated via glucocorticoid receptors.
Different rat-derived transforming retroviruses code for an immunologically related intracellular phosphoprotein.
Authors: Young H A , Shih T Y , Scolnick E M , Rasheed S , Gardner M B
Published: May 2006

Kirsten sarcoma virus (Ki-MSV) and Harvey sarcoma virus (Ha-MSV) are mouse-rat recombinant viruses that were originally isolated by experimental inoculation of rats with helper-independent mouse type C viruses. We have recently identified in cells transformed by Ki-MSV or Ha-MSV, a phosphoprotein, p21, coded for by Ki-MSV and Ha-MSV [Shih, T.Y., Weeks, M.O., Young, H.A. & Scolnick, E.M. (1979) Virology 95, in press]. The p21, which is not a virion structural protein, was identified with antisera prepared by transplantation in rats of syngeneic Ha-MSV- or Ki-MSV-transformed nonproducer cells. In this study, we have applied the same methodology to examine a purely rat sarcoma virus (RaSV), which was isolated in cell culture by using helper-independent rat type C viruses [Rasheed, S., Gardner, M.B. & Huebner, R.J. (1978) Proc. Natl. Acad. Sci. USA 75, 2972-2976]. We report here that this new, purely rat sarcoma virus apparently codes for a p29, which shares immunological determinants and common V-8 protease-generated peptides with the p21 of Ha-MSV. The data suggest that the RaSV has acquired genetic information with similar coding capacity to some rat genetic information with similar coding combinant viruses, Ki-MSV and Ha-MSV. Based on data obtained on the p21 of a mutant of Ki-MSV temperature-sensitive for the maintenance of transformation, we suggest that the gene in RaSV that codes for the p29 is also required for the maintenance of RaSV-induced fibroblast transformation.
Effect of molecular rotation on the vibration-vibration energy transfer in condensed media
Authors: Lin S. H. , Hays T. R. , Eyring H.
Published: May 2006

The purpose of the present communication has been to present the derivation of the vibrational relaxation rate constant for the case in which both phonons and the molecular rotation (three-dimensional) participate in the vibrational relaxation and to study the effect of molecular rotation on the vibration-vibration energy transfer in the condensed phase. The experimental results of vibration-vibration energy transfer between donors, NH and ND, and acceptors, CO and
13
CO, have been analyzed.
Brasilenyne and
cis
-dihydrorhodophytin: Antifeedant medium-ring haloethers from a sea hare (
Aplysia brasiliana
)
Authors: Kinnel R. B. , Dieter R. K. , Meinwald J. , Van Engen D. , Clardy J. , Eisner T. , Stallard M. O. , Fenical W.
Published: May 2006

Two straight-chain C
15
fish antifeedants have been isolated from the sea hare
Aplysia brasiliana
. Chemical, spectral, and x-ray diffraction studies led to the characterization of these medium-ring ethers as brasilenyne (2) and
cis
-dihydrorhodophytin (3). The oxonin ring system of 2 is novel in nature. Biosynthetic considerations permit the postulation that a third compound, a noncrystalline congener of these compounds, is
cis
-isodihydrohodophytin (4).
Reconstitution and purification by "transport specificity fractionation" of an ATP-dependent calcium transport component from synaptosome-derived vesicles.
Authors: Papazian D , Rahamimoff H , Goldin S M
Published: May 2006

A synaptosomal ATP-dependent Ca uptake system was reconstituted into artificial vesicles by a cholate dialysis procedure using an 80-fold excess of exogenous phospholipid. Under these conditions, most of these vesicles would be expected to have only one or, at most, a few membrane proteins. The vesicles containing an ATP-dependent Ca transport system were purified from the bulk of the preparation on density gradients by increasing their density by the ATP-dependent intravesicular precipitation of Ca oxalate; a approximately 100-fold purification resulted. The purified Ca-transporting vesicles contained two major protein components, of Mr 94,000 and 140,000 according to sodium dodecyl sulfate gel electrophoresis. These components are believed to be responsible for Ca transport in this synaptosome-derived membrane fraction.
UV-endonuclease from calf thymus with specificity toward pyrimidine dimers in DNA.
Authors: Waldstein E A , Peller S , Setlow R B
Published: May 2006

We describe the partial purification of an endonuclease from calf thymus that nicks phage PM2 DNA irradiated with UV doses producing only a few pyrimidine dimers per molecule. It has much less activity on DNA that has been subjected to enzymatic photoreactivation after UV irradiation. The calf thymus endonuclease is different from other mammalian UV-endonucleases so far described in that it seems to be dimer specific. The enzyme is stimulated by Mg2+ and is inactive in the presence of EDTA. It binds to UV-irradiated DNA-Sepharose from which it is released by low concentrations of KCl. Gel filtration data indicate that the endonuclease may belong to a high molecular weight protein or protein complex. The enzyme is very labile and freezing increases its lability.
Reactivation of silent rRNA genes by simian virus 40 in human-mouse hybrid cells.
Authors: Soprano K J , Dev V G , Croce C M , Baserga R
Published: May 2006

Mouse-human hybrid cells were used to study the ability of simian virus 40 to regulate the expression of rRNA genes in vivo. In these hybrid cells, only the rRNA genes of the dominant species are expressed; the genes for the rRNA of the recessive species are silent. Simian virus 40 infection of these hybrids led to the production of two distinct 28S rRNA species as analyzed by agarose/2.4% polyacrylamide gel electrophoresis. These species were identified as human and mouse rRNAs. This result was confirmed by histochemical studies which indicated that the nucleolus organizer regions of both mouse and human chromosomes were actively synthesizing rRNA in the virus-infected hybrid cells. These results indicate that simian virus 40 infection can induce the expression of otherwise silent rRNA genes.
Proliferation of Rous sarcoma virus-infected, but not of normal, chicken fibroblasts in a medium of reduced calcium and magnesium concentration
Authors: Balk Samuel D. , Polimeni Philip I. , Hoon Baldev Singh , LeStourgeon Dana N. , Mitchell Richard S.
Published: May 2006

Both normal and Rous sarcoma virus-infected chicken fibroblasts proliferate actively in a culture medium containing physiological concentrations of calcium (1.2 mM) and magnesium (0.7 mM). In the presence of a physiological concentration of magnesium, reduction of the calcium concentration to 0.125 mM resulted in a significant decrease in the proliferation of the normal, but not of the neoplastic, fibroblasts. Reduction of the magnesium concentration to 0.05 mM in the presence of a physiological concentration of calcium had a similar effect. In a culture medium containing reduced concentrations of both calcium (0.20 mM) and magnesium (0.05 mM), the normal fibroblasts were maintained without proliferation, whereas the Rous sarcoma virus-infected fibroblasts continued to proliferate actively. The cytosol concentrations of ionized calcium and magnesium are known to be regulated by a balance between net passive influx and active extrusion and sequestration. On the basis of this consideration and the findings described above it can be hypothesized that: (
i
) Fibroblast replication is initiated when cytosolic concentrations of calcium, magnesium, or both rise above a critical level. (
ii
) Autonomous initiation of replication of neoplastic fibroblasts is a result of failure of cytoplasmic divalent cation homeostasis; alternatively, sarcoma virus infection may endow cells with a divalent cation-independent mechanism that bypasses an initiation mechanism that is, normally, divalent cation-dependent. (
iii
) Proliferation of normal fibroblasts is controlled by extracellular matrix components that interact with cell surfaces in a manner that limits the permeability of plasma membranes to divalent cations or otherwise functions to lower cytosol divalent cation concentrations.
Ultraviolet light induction of diphtheria toxin-resistant mutants of normal and xeroderma pigmentosum human fibroblasts.
Authors: Glover T W , Chang C C , Trosko J E , Li S S
Published: May 2006

The UV induction of diphtheria toxin-resistant (DTr) mutants in normal and xeroderma pigmentosum human fibroblasts has been quantitatively characterized. A concentration of diphtheria toxin at which DTr cells are cross-resistant to Pseudomonas aeruginosa exotoxin A was determined and used in the selection of resistant mutants. Recovery of mutants was not influenced by the presence of wild-type cell densities of 1-8 x 10(5) per 9-cm plate, indicating no metabolic cooperation exists, in contrast to what is seen in the selection of some other variant phenotypes. Expression periods for UV-induced mutations differed with the severity of mutagen treatment and cell strain used. A relatively long (10-15 days after UV treatment) expression period was required for the maximum recovery of DTr mutants. Maximum recovery was followed by a decrease in mutation frequency on subsequent days evaluated. An apparent linear dose response within the dose range used was observed for UV-induced mutations in both normal and xeroderma pigmentosum fibroblasts. Our results indicate that xeroderma pigmentosum fibroblasts have higher UV-induced mutation frequencies per unit UV dose but similar frequencies per unit survival compared to normal cells within the range of UV doses tested.
Induction, establishment in vitro, and characterization of functional, antigen-specific, carrier-primed murine T-cell lymphomas.
Authors: Finn O J , Boniver J , Kaplan H S
Published: May 2006

T cells were isolated from spleens of C57BL/Ka/Thy 1.1/Lb mice immunized with 2,4-dinitrophenylated keyhole limpet hemocyanin (DNP-KLH). The mice were infected in vitro with radiation leukemia virus and injected intrathymically into congenic C57BL/Ka (Thy 1.2) recipients. Within 3-4 months, seven thymic lymphomas developed, five of which were of donor type. The donor lymphomas were explanted and permanent cell lines were established in vitro. These lymphoma cells are capable of providing antigen-specific carrier (KLH)-primed help to DNP-primed B lymphocytes in secondary antibody production to DNP-KLH. They also enhance the secondary antibody response of whole period spleen cell populations to DNP-KLH. The availability of these immortal clonable populations of immunofunctional neoplastic T lymphocytes should facilitate biological and biochemical investigations of lymphocyte interactions during synthesis of antibody to thymus-dependent antigens.
Evaluation of a common receptor-complement concept for predicting the antileukemic activity of nucleosides.
Authors: Townsend L B , Schram K H , Beránek J
Published: May 2006

The triangulation arrangement of one nitrogen and two oxygen atoms proposed as a requirement for antileukemic activity involving a "common receptor-complement feature" has been reevaluated specifically as it applies to nucleosides. This re-evaluation has resulted in a rejection of the "receptor-complement feature" as a working model for the a priori design of nucleosides as compounds active against leukemia L-1210.
Toad urinary bladder epithelial cells in culture: maintenance of epithelial structure, sodium transport, and response to hormones.
Authors: Handler J S , Steele R E , Sahib M K , Wade J B , Preston A S , Lawson N L , Johnson J P
Published: May 2006

Epithelial cells from the toad urinary bladder have been grown in continuous culture. Many of the cells resemble the granular cell type of the urinary bladder. They form an epithelium with typical tight junctions and gap junctions. The transport properties of two cell lines have been examined. When cells of the line designated TB-M or of line TB-6c are grown on collagen-coated nucleopore filters, epithelia are formed that have transepithelial potential differences of 40 and 20 mV, resistances of 5000 and 10,000 omega-cm2, and short-circuit currents (ISC) of 8.5 and 2.5 muA/cm2, respectively. Net mucosa to serosa sodium transport accounts for all of ISC in line TB-M and for 70% of ISC in line TB-6c. Vasopressin, which stimulates adenylate cylase and ISC in the intact bladder, has no effect on the cells in culture. Cyclic AMP stimulates ISC and lowers resistance in both lines. Aldosterone stimulates ISC in both lines. This is accompanied by a fall in resistance in line TB-M and no change in resistance in line TB-6c. Amiloride inhibits ISC in TB-M cells under basal conditions and after stimulation by aldosterone. In line TB-6c amiloride has no effect under basal conditions but lowers ISC of aldosterone-treated cells to the basal level. Thus, the cells have retained the ability to form oriented, high-resistance epithelial membranes that manifest hormone-sensitive transepithelial sodium transport.
A theory of light-induced explosion
Authors: Lin S. H. , Richardson W. B. , Eyring H.
Published: May 2006

This paper presents a theoretical analysis, based on a simplified one-dimensional model, of the response of an explosive to a wide range of light intensities. A primary result has been the identification and development of a relationship among the important critical parameters. Numerical results illustrating the relationship among the critical light intensity, sample dimension, and ambient temperature have been obtained, with lead azide as the example.
Orientation of rhodopsin alpha-helices in in retinal rod outer segment membranes studied by infrared linear dichroism.
Authors: Michel-Villaz M , Saibil H R , Chabre M
Published: May 2006

Frog retinal rod outer segments, oriented by a magentic field, were shown to contain rhodopsin alpha-helical segments preferentially aligned perpendicular to the plane of the disc membrane, by the technique of infrared linear dichroism. Infrared absorption parallel and perpendicular to the rod axes by peptide C parallel to O groups, whose absorption band contains alpha-helical and random coil components at slightly different frequencies, showed positive dichroism centered on the alpha-helix frequence. We conclude that the alpha-helical portion of the protein has an average orientation in the transmembrane direction. Furthermore, infrared spectra of rods in 2H2O Ringer's solution exhibit two distinct peptide amino group absorption bands: the unexchanged N-2H band, which is nondichroic. This implies that the oriented part of the protein is in the lipid bilayer, supporting a model for rhodopsin with a hydrophobic core containing partially oriented alpha-helices and hydrophilic ends consisting of unoriented polypeptide.
Cloned pairs of variable region genes for immunoglobulin heavy chains isolated from a clone library of the entire mouse genome.
Authors: Kemp D J , Cory S , Adams J M
Published: May 2006

To investigate the organization of immunoglobulin genes, we have constructed a clone library containing 10(6) randomly generated fragments of mouse embryo DNA, corresponding to eight equivalents of the genome. The cloning method involved methylation of embryo DNA at EcoRI recognition sites, partial digestion by EcoRI* endonclease activity, and direct ligation of the resulting large fragments to the lambda phage vector Charon 4A. The library was searched for sequences homologous to a cloned complementary DNA copy of a mu heavy chain mRNA. Nine clones bearing variable heavy chain (VH) sequences were isolated, representing at least eight distinct VH genes. Thus, multiple related VH genes are available in the genome to contribute to immunoglobulin diversity. Each of the two clones carries a pair of VH genes, one pair separated by 15 +/- 1 kilobase pairs of mouse DNA and the other by 14 +/- 2 kilobase pairs. This indicates that related VH genes are clustered and may occur in a tandem array having a repeating unit of 14--16 kilobase pairs. The large spacer sequences between VH genes cannot, however, be highly conserved.
Differential affinities of simian virus 40 large tumor antigen for DNA.
Authors: Oren M , Winocour E , Prives C
Published: May 2006

The binding of simian virus 40 (SV40) large tumor antigen (T antigen) to DNA was analyzed by using the salt-sensitive affinities of the protein for various DNAs immobilized on cellulose. At least two types of interactions could be distinguished that differed in their stability. Higher salt concentrations were required to elute T antigen from SV40 DNA than from calf thymus DNA; and even greater salt concentrations were required for the lution of T antigen from multiorigin SV 40 DNA compared to wild-type SV40 DNA. This would indicate that T antigen can bind weakly or strongly to DNA, depending on the DNA sequence. It was also found that a greater proportion of rapidly labeled or newly synthesized T antigen binds more efficiently and tightly to multiorigin SV40 DNA than to long-labeled or older forms of T antigen. This approach can be utilized not only to distinguish between different forms of T antigens which vary in their affinities for DNA but also for rapidly obtaining highly enriched T antigen preparations.
Ovalbumin is synthesized in mouse cells transformed with the natural chicken ovalbumin gene.
Authors: Lai E C , Woo S L , Bordelon-Riser M E , Fraser T H , O'Malley B W
Published: May 2006

The entire chicken ovalbumin gene, accompanied by genomic DNA sequences flanking both termini of the gene and three copies of the herpes simplex virus thymidine kinase gene, has been cloned in plasmid pBR322. This recombinant plasmid was linearized and used to transform thymidine kinase-deficient mouse cells. Thymidine kinase-positive transformants were selected by their ability to grow in the hypoxanthin/aminopterin/thymidine (HAT) medium. The entire ovalbumin gene integrated into high molecular weight DNA within all the transformants and retained its original sequence organization. In all of the transformants examined, a protein identified as chicken ovalbumin by immunoreactivity was detected within the cells. It is estimated that between 1000 and 100,000 molecules of chicken ovalbumin were produced per mouse cell in each of these transformants. Our results demonstrate that the mouse cellular machinery can be utilized to accurately express genetic information encoded in a cloned gene from a different eukaryotic organism into its specific protein product.
Charge selectivity at the lipid-protein interface of membranous Na,K-ATPase.
Authors: Brotherus J R , Jost P C , Griffith O H , Keana J F , Hokin L E
Published: May 2006

Lipid interactions with the integral membrane protein Na,K-ATPase (ATP phosphohydrolase, EC 3.6.1.3) purified from the electric organ of Electrophorus electricus were studied by spin labeling. A protein-associated component (boundary layer) in equilibrium with the fluid bilayer is clearly evident in the electron spin resonance spectra. The influence of charge on this equilibrium was determined by varying the head group of the lipid while maintaining the chain length and the position of the label constant. The lipid spin labels were 14-proxylstearylmethyl phosphate and the corresponding dimethylphosphate, alcohol, and quaternary amine. By using a pairwise spectral analysis, as well as a conventional spectral analysis, the binding affinity was found to decrease in the order of negative greater than neutral greater than positive charges. The fraction bound decreased from about 0.57 for the negatively charged phosphate to 0.25 for the positively charged quaternary amine. The amount of each bound lipid was nearly constant over the temperature range investigated (5-35 degrees C). High salt concentrations reversibly abolished the selectivity between the labels, confirming the role of charge in the binding equilibria.
Alternative method for identifying reconstituted cells
Authors: Shay Jerry W. , Clark Mike A.
Published: May 2006

Enucleation techniques combining mild centrifugation in the presence of cytochalasin B permit cells to be separated into nuclear fragments (karyoplasts) and cytoplasmic fragments (cytoplasts). These fragments, though stable for a short time, will ultimately degenerate by the procedures described in this report. One can, however, fuse cytoplasts to karyoplasts by using polyethylene glycol and obtain viable reconstituted cells whose properties may be useful for understanding some aspects of the nuclear-cytoplasmic interactions associated with tumorigenicity and steroidogenesis. However, the presence of cybrids, hybrids, and parental whole cell contaminants along with the reconstituted cell population make it necessary to have genetic markers that reside in both the nucleus and cytoplasm in order to preferentially identify reconstituted cells derived from a karyoplast fused to a cytoplast. By utilizing the Y-1 cell line, which is tumorigenic and responds to corticotropin by secreting steroids, and the AMT-BU-A1 (AMT) cell line, which is nontumorigenic and does not respond to corticotropin but has a nuclear marker,

BrdUrd
r

, and a cytoplasmic marker,

CAP
r

, we have reconstituted cells containing Y-1 karyoplasts and AMT cytoplasts. In this report we extend our previous techniques by describing an identification procedure that allowed us to isolate cells reconstituted from AMT karyoplasts fused to Y-1 cytoplasts. The results of these experiments support the concept that with these cell lines the nucleus (karyoplast) is ultimately sufficient to control the phenotypic expression or suppression of tumorigenicity and steroidogenesis.
Induction of plasminogen activator secretion in macrophages by electrochemical stimulation of the hexose monophosphate shunt with methylene blue.
Authors: Schnyder J , Baggiolini M
Published: May 2006

Resident peritoneal macrophages were obtained from untreated mice and were cultured in medium 199 with or without 5% acid-treated fetal bovine serum. Three hours after harvesting, redox compounds--i.e., methylene blue, methyl viologen, or nitro blue tetrazolium--were added to the cultures of adherent cells. After 1 hr, the cells were washed and culturing was continued in the absence of redox compounds. The effects of the redox compounds were tested by assaying for hexose monophosphate (HMP) shunt activity and for plasminogen activator secretion, and the results were compared with the effects induced by phagocytic stimuli. Methylene blue caused a concentration-dependent stimulation of the HMP shunt, whereas methyl viologen and nitro blue tetrazolium were ineffective. Shunt stimulation by methylene blue was followed, after a lag of 2-4 days, by plasminogen activator secretion. The rate of secretion was dependent on the methylene blue concentration used. Methyl viologen and nitro blue tetrazolium were again ineffective, whereas phagocytosis of zymosan or sheep erythrocytes, which stimulates the HMP shunt, induced plasminogen activator secretion at rates similar to those induced by methylene blue. These results add further evidence to our hypothesis that the HMP shunt-dependent metabolic burst is involved in macrophage activation. Because methylene blue mimics the action of zymosan it appears that shunt stimulation by itself initiates the activation process independently of phagocytosis.
Enhanced drug-metabolizing capacity within liver adjacent to human and rat liver tumors.
Authors: Sultatos L G , Vesell E S
Published: May 2006

Cytochrome P-450 content (nmol/g of liver) differed within regions of rat liver according to proximity to intrahepatically implanted Morris hepatoma 7795 or 5123D. Liver adjacent to tumor had higher microsomal cytochrome P-450 content, decreased DNA content (mg/g of liver), and unaltered cytochrome c reductase activity compared to histologically indistinguishable liver far-removed from the tumor. Liver either adjacent to or far-removed from tumor contained markedly more cytochrome P-450 and higher cytochrome c reductase activity but less DNA than transplanted Morris hepatomas 7795 and 5123D that were grown intrahepatically. Compared to intramuscular implants of these same tumors, intrahepatically implanted Morris hepatomas 7795 and 5123D had increased cytochrome P-450 content. Tumor-containing liver from two human subjects revealed regional changes in cytochrome P-450-mediated monooxygenases similar to those observed in rats. These results suggest that histomorphically nontumorous mammalian liver directly adjacent to intrahepatic tumors exhibits previously unsuspected biochemical alterations.
Interplexiform cell in cat retina: identification by uptake of gamma-[3H]aminobutyric acid and serial reconstruction.
Authors: Nakamura Y , McGuire B A , Sterling P
Published: May 2006

After intravitreal injection of gamma-[3H] aminobutyric acid (GAB), 2% of the neurons at the outer margin of the inner plexiform layer were intensely labeled. Reconstructions of these neurons from serial electron microscope autoradiograms showed that they are interplexiform cells, which synapse on bipolar processes in the outer plexiform layer and on amacrine and bipolar processes in the inner plexiform layer.
Stress-induced release of prolactin: blockade by dexamethasone and naloxone may indicate beta-endorphin mediation.
Authors: Rossier J , French E , Rivier C , Shibasaki T , Guillemin R , Bloom F E
Published: May 2006

Basal levels of immunoreactive (ir) beta-endorphin, corticotropin (ACTH), and prolactin (PRL) in plasma of male rats decrease after dexamethasone pretreatment (400 microgram/kg at 24 hr and 200 microgram/kg at 2 hr before). Inescapable electric footshocks increase ir-beta-endorphin, ACTH, and PRL plasma levels and this effect is blocked by dexamethasone pretreatment. Morphine (20 mg/kg) also increases ir-beta-endorphin, ACTH, and PRL levels. Dexamethasone pretreatment blocks the morphine-induced release of ir-beta-endorphin but does not prevent the morphine-induced release of PRL. Naloxone, the opiate antagonist, decreases basal plasma levels of PRL and partially blocks the stress-induced increase of PRL, but it has no effect on the basal or stress-induced release of ir-beta-endorphin. These results are consistent with the proposal that beta-endorphin may interact with an opiate receptor involved in the regulation of PRL secretion.
Differences in glycosylation patterns of closely related murine leukemia viruses
Authors: Rosner Marsha Rich , Grinna Lynn S. , Robbins Phillips W.
Published: May 2006

The nature of the carbohydrate chains in the major envelope glycoprotein of murine leukemia virus, gp70, and its cellular precursor has been investigated. A difference in the oligosaccharide composition of gp70 from an ecotropic murine leukemia virus (Akv) and three recombinant dual-tropic viruses [mink cell focus-inducing viruses (MCFs)] derived from Akv was demonstrated. Glycosidase digestion and gel filtration were utilized to identify the two classes of
N
-asparagine-linked oligosaccharides, high-mannose and complex. The gp70 of the ecotropic virus contained only
N
-linked oligosaccharides of the complex type. In contrast, the gp70s of the dual-tropic viruses contained both high-mannose and complex oligosaccharides. Analysis of gp70 glycopeptides from an MCF-related xenotropic virus showed an elution profile similar, but not identical, to profiles of the MCFs. The gp70 precursors isolated from cells infected with Akv or MCF virus contained
N
-linked oligosaccharides that were exclusively of the high-mannose type. Comparison of the high-mannose oligosaccharides of the MCF gp70 precursors with those of the corresponding gp70s indicated that very little further processing of the high-mannose residues in the gp70s had occurred. The presence of the high-mannose oligosaccharides in the envelope glycoprotein of the dual-tropic viruses results from altered carbohydrate processing. The conservation of this altered carbohydrate pattern in a number of hosts and under various conditions of growth suggests that the viral protein structure is the primary factor in determining the different mode of glycosylation of the MCF gp70s. Thus, these viral glycoproteins provide an important model system for studying the relationship between protein structure and patterns of glycosylation.
Hypothetical strain-free oligoradicals
Authors: Hoffmann Roald , Eisenstein Odile , Balaban Alexandru T.
Published: May 2006

Several new classes of oligoradicals free of angle strain are suggested and examined by means of molecular orbital calculations. The collapse products of these hypothetical radicals are highly strained molecules. Various electronic strategies for the stabilization of these oligoradicals have been explored.
On the asymptotic eigenvalue distribution of a pseudo-differential operator
Authors: Fefferman C. , Phong D. H.
Published: May 2006

A description of the number
N(K)
of eigenvalues less than
K
for a pseudo-differential operator with positive symbol is given in terms of the number of unit cubes canonically imbedded in the subset of phase space where the symbol is less than
CK
. This gives back in particular the order of magnitude of
N(K)
for elliptic symbols.
Modulation of cyclic AMP metabolism by S-adenosylhomocysteine and S-3-deazaadenosylhomocysteine in mouse lymphocytes.
Authors: Zimmerman T P , Schmitges C J , Wolberg G , Deeprose R D , Duncan G S , Cuatrecasas P , Elion G B
Published: May 2006

Mouse lymphocytes incubated with micromolar concentrations of adenosine or 3-deazaadenosine, in medium supplemented with L-homocysteine, rapidly accumulated supramillimolar concentrations of S-adenosylhomocysteine (AdoHcy) or S-3-deazaadenosylhomocysteine (c3AdoHcy), respectively. Lymphocytes thus preloaded with high levels of AdoHcy or c3AdoHcy exhibited markedly enhanced (5- to 40-fold) cyclic AMP responses to prostaglandin E1, adenosine, 2-chloroadenosine, isoproterenol, and cholera toxin. This enhancement of cyclic AMP response by intracellular AdoHcy or c3AdoHcy was attributable both to amplification of the activity of adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] and to inhibition of cyclic AMP phosphodiesterase (3',5'-cyclic-nucleotide 5'-nucleotidohydrolase, EC 3.1.4.17). Basal and prostaglandin E1- and isoproterenol-stimulated activities of adenylate cyclase, assayed in lymphocyte homogenates, were increased 1.3- to 2.0-fold after treatment of the cells with homocysteine plus either adenosine or 3-deazaadenosine. AdoHcy and c3AdoHcy were found to be competitive inhibitors (with Ki values of 1.7 and 4.8 mM, respectively) of the high-affinity cyclic AMP phosphodiesterase present in lymphocyte homogenates. It is evident, therefore, that increased cellular levels of AdoHcy or c3AdoHcy can affect cellular physiology via perturbation of cyclic AMP metabolism as well as via inhibition of S-adenosylmethionine-dependent methylation reactions.
Identification of proteolytically resistant domains of human erythrocyte spectrin.
Authors: Speicher D W , Morrow J S , Knowles W J , Marchesi V T
Published: May 2006

Digestion of purified human erthrocyte spectrin with proteolytic enzymes at 0 degrees C results in the production of intermediate-size peptides that resist further cleavage at 0 degrees C. By two-dimensional peptide analysis of these intermediate peptides it has been determined that five unique peptides are produced by tryptic cleavage of the alpha subunit of spectrin (band 1); these have apparent molecular weights of 80,000, 46,000, 46,000, 41,000, and 30,000 and account for 97% of the alpha subunit. Similarly, four unique peptides having apparent molecular weights of 74,000, 65,000, 33,000, and 38,000 account for 90% of the beta subunit (band 2). By examining larger peptide fragments, the linear alignment of the unique peptides along each of the spectrin subunits has been established. These results indicate that spectrin is composed of two nonidentical subunits, each containing multiple proteolytically resistant domains. These domains, which may be largely alpha-helical, seem to be connected by small protease-sensitive segments. The proteolytic resistance of these domains is not influenced by the multimeric state of the spectrin molecule.
Relation between enzymic catalysis and energy coupling.
Authors: Fry M , Blondin G A , Green D E
Published: May 2006

The principles that underlie enzyme catalysis also apply to energy coupling processes. A comparison is made between a kinase system that mediates the phosphorylation of glucose by ATP (hexokinase), as the prototype for enzymic catalysis, and the mitochondrial electron-transfer complexes, as the prototypes for energy coupling systems. Induced polarization of chemical bonds and charge separation and elimination are common component events of both enzyme catalysis and energy coupling. Thus, definite limits can be imposed on models of energy coupling; they must comply with the basic principles of enzymic catalysis.
Cytoplasmic processing of myosin heavy chain messenger RNA: evidence provided by using a recombinant DNA plasmid.
Authors: Medford R M , Wydro R M , Nguyen H T , Nadal-Ginard B
Published: May 2006

A recombinant DNA plasmid, designated pMHC25, has been constructed that contains structural gene sequences for rat skeletal muscle myosin heavy chain (MHC). The identity of the MHC sequence insert in pMHC25 was determined by muscle-tissue specificity, inhibition of MHC protein synthesis in vitro by hybrid-arrested translation, purification of mRNA that directs the synthesis of MHC protein in vitro, and hybridization to a 33S cytoplasmic mRNA found only in differentiated muscle cells. pMHC25-DNA-excess filter hybridizations were used to show that more than 90% of the newly synthesized MHC mRNA that appears in the cytoplasm of differentiated L6E9 myotubes contains a long 3' poly(A) tail. In contrast, 90% of the MHC mRNA that accumulates in the cytoplasm of these same cells during myogenic differentiation lacks this long 3' poly(A) tail. These results suggest the occurrence of a posttranscriptional event in differentiated L6E9 myotubes that involves the cytoplasmic processing of poly(A)+ MHC mRNA to poly(A)- or poly(A)-short MHC mRNA.
Structure of endogenous murine leukemia virus DNA in mouse genomes.
Authors: Chattopadhyay S K , Lander M R , Rands E , Lowy D R
Published: May 2006

By using molecularly cloned ecotropic AKR murine leukemia virus (MuLV) DNA, a 400-base-pair ecotropic type-specific segment in the env region has been identified. This DNA segment and other defined viral subgenomic fragments have been used as 32P-labeled probes to identify and analyze the structure of integrated ecotropic viral DNA sequences in uninfected mouse genomes. Those mice from which endogenous ecotropic MuLV of the AKR type have been isolated contained at least one virtually complete linear copy of the viral genome. Strains from which ecotropic MuLV has not been isolated lacked ecotropic-specific sequences. All inbred mouse strains tested also contained MuLV DNAs of genomic length whose restriction endonuclease digestion pattern was characteristic of xenotropic viruses.
Catecholamine biosynthetic enzymes are expressed in replicating cells of the peripheral but not the central nervous system.
Authors: Rothman T P , Specht L A , Gershon M D , Joh T H , Teitelman G , Pickel V M , Reis D J
Published: May 2006

We sought to determine whether the precursors of catecholamine-containing neurons in the developing peripheral and central nervous systems of chickens and rats express the biosynthetic enzymes tyrosine hydroxylase [THase; tyrosine 3-monooxygenase; L-tyrosine, tetrahydropteridine: oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2] or dopamine beta-hydroxylase [DBHase; 3,4-dihydroxyphenylethylamine, ascorbate:oxygen oxidoreductase (beta-hydroxylating), EC 1.14.17.1], prior to the time they withdraw from the cell cycle. Chicken embryos (stages 26-27) were injected with [3H-thymidine and 4 hr later were prepared for the simultaneous demonstration of radioautographically labeled nuclei in immunoreactive THase cells. The brains and sympathetic chains of rat fetuses (days E12-E14), exposed for 2 hr to [3H]thymidine, were treated similarly except that peripheral tissues were stained with a specific antibody to DBHase as well as anti-THase. In the peripheral nervous system of both chicken and rat, nuclei of THase-containing cells were radioautographically labeled. DBHase-containing cells in the peripheral nervous system of rats were also labeled and thus are noradrenergic. THase was localized in cells of the brain of the same rat fetuses beginning on day E12 (no THase was detected on day E11 or E11.5) in the mantle layer of the ventral mesencephalic and rostrolateral rhombocephalic cellular groups; however. THase-containing cells in the central nervous system did not incorporate [3H]thymidine. We conclude that, during development, the adrenergic neuronal precursors of the peripheral nervous system but not of the central, have the capacity to synthesize catecholamines before they withdraw from the cell cycle. Differences in the maturation of peripheral and central neurons may be related to differences in their embryological origin.
Ligand substitution reactions of metallothioneins with EDTA and apo-carbonic anhydrase.
Authors: Li T Y , Kraker A J , Shaw C F , Petering D H
Published: May 2006

The reactions of Zn-, Zn,Cd-, and Cd-thioneins with EDTA and apo-carbonic anhydrase have been studied. The ligand substitution reaction of zinc with EDTA is multiphasic, having both associative and dissociative components in the rate expression. The cadmium sites are about 2 orders of magnitude less reactive. In contrast, apo-carbonic anhydrase abstracts zinc from Zn-thionein and Zn,Cd-thionein in second-order processes that are 2-3 orders of magnitude more rapid than those involving EDTA and approach the rate for unligated Zn2+ with the apo-protein. In comparison with other zinc proteins, Zn-thionein contains unusually reactive metal sites, suggesting that this protein may be a physiological zinc transfer protein, able to donate zinc to zinc-requiring apo macromolecules.