bluuebunny/crossref_metadata_2025 huggingface dataset:
Structural markers on core protein p30 of murine leukemia virus: functional correlation with Fv-1 tropism.
Authors: Gautsch J W , Elder J H , Schindler J , Jensen F C , Lerner R A
Published: May 2006

Tryptic peptide maps from more than 50 isolates of murine leukemia virus (MuLV) have shown that, in general, the structure of core protein p30 is highly conserved. However, a structurally variable region of p30 has been identified that is functionally associated with Fv-1 tropism. On the basis of this structural variability, MuLV strains can be classified as B-tropic, N-tropic, xenotropic, and/or as being derived from wild mice. Certain xenotropic viruses have a p30 like that of B-tropic MuLV and presumably would be subject to restriction in cells containing an Fv-In allele. Other p30 structural markers serve to distinguish the exogenous Friend, Moloney, and Rauscher viruses from endogenous MuLV. Furthermore, some MuLV strains have structural differences in their p30s that are useful as strain-specific markers. Finally, a possible sarcoma-associated alteration in the structure of p30 has been noted in the ml clone of Moloney murine sarcoma virus.
Changes in hepatic messenger RNA for phospho
enol
pyruvate carboxykinase (GTP) during development
Authors: Ruiz Josefa P. Garcia , Ingram Robert , Hanson Richard W.
Published: May 2006

Phospho
enol
pyruvate carboxykinase (GTP) [GTP;oxaloacetate carboxy-lyase(transphosphorylating); EC 4.1.1.32] is absent in rat liver cytosol during fetal life and is synthesized initially at birth.
De novo
synthesis of the enzyme can be induced prematurely by injection of dibutyryl cyclic AMP or glucagon into fetal animals
in utero
. In this study a wheat germ translation assay was used to quantitate the level of total functional mRNA for phospho
enol
pyruvate carboxykinase in the liver of fetal rats at 21 days of pregnancy under different induction situations. The translatable mRNA for the enzyme was marginally detectable in fetal rat liver. Administration of either glucagon or dibutyryl cyclic AMP to fetal rats
in utero
caused a marked induction of functional mRNA for this enzyme. Three hours after administration of dibutyryl cyclic AMP, the level of translatable mRNA increased almost 23-fold, but by 6 hr the level dropped approximately 60%. Administration of actinomycin D prior to dibutyryl cyclic AMP in 21-day fetal rats prevented the appearance of newly synthesized poly(A)-containing RNA in the cytoplasm as well as the induction of translatable mRNA for phospho
enol
pyruvate carboxykinase. In animals delivered prematurely and maintained for varying periods, the translatable mRNA for the enzyme accumulated in the liver at a rate comparable to that observed for enzyme synthesis.
Cosmids: a type of plasmid gene-cloning vector that is packageable in vitro in bacteriophage lambda heads.
Authors: Collins J , Hohn B
Published: May 2006

Evidence is presented that ColE1 hybrid plasmids carrying the cohesive-end site (cos) of lambda can be used as gene cloning vectors in conjunction with the lambda in vitro packaging system of Hohn and Murray [(1977) Proc. Natl. Acad. Sci. USA 74, 3259--3263]. Due to the requirement for a large DNA molecule for efficient packaging, there is a direct selection for hybrids carrying large sections of foreign DNA. The small vector plasmids do not contribute a large background in the transduced population, which is therefore markedly enriched for large hybrid plasmids (over 90%). The efficiency of the in vitro packaging system is on the order of 10(5) hybrid clones per microgram of foreign DNA for hybrids in the 20--30 million dalton range.
Efficient correction of a mutation by use of chemically synthesized DNA.
Authors: Razin A , Hirose T , Itakura K , Riggs A D
Published: May 2006

The mutated base in the am3 lysis-defective mutant of the bacteriophage phiX174 has been corrected by a combined in vitro enzymatic DNA synthesis and in vivo replication of the heteroduplex product. Chemically synthesized oligodeoxyribonucleotides carrying the wild-type sequence have been used to prime DNA synthesis with am3 phiX174 DNA serving as a template. The resultant semisynthetic heteroduplex composed of an am3(+) strand and a wild-type (-) strand, with one mismatched base pair at position 587 on the phiX174 DNA sequence, was used to infect spheroplasts. The progeny phage were analyzed by a parallel plaque assay on wild-type host, Escherichia coli C, to screen for wild-type phenotype, and on E. coli HF4714, an amber suppressor strain, to determine the total progeny phage. When a 23-base-long synthetic primer was used, about one-third of total progeny were found to be wild type. Shorter primers yielded lower percentages of wild type; they also had poorer priming activity.
Nucleotide sequence of the attenuator region of the histidine operon of Escherichia coli K-12.
Authors: Di Nocera P P , Blasi F , Di Lauro R , Frunzio R , Bruni C B
Published: May 2006

The attenuator region of the histidine operon of Escherichia coli K-12 has a potential coding capacity for two peptides, one of 16 amino acids and another of 30 amino acids. This region is followed by a perfect palindrome of 14 base pairs separated by five nucleotides. A G+C-rich region precedes and follows a possible transcription termination sequence. These features are compatible with a model in which active translation of a leader mRNA interferes with transcription termination, thus causing derepression of the histidine operon. The sequence of the region coding for the hypothetical 16-amino acid peptide is of particular relevance because it indicates the site and a possible mechanism of action of histidyl-tRNAhis in regulating histidine gene expression. Seven contiguous histidine codons are present within this sequence: : formula: (see text)
Isolation, characterization, and synthesis of a corticotropin-inhibiting peptide from human pituitary glands.
Authors: Li C H , Chung D , Yamashiro D , Lee C Y
Published: May 2006

A corticotropin-inhibiting peptide (CIP) has been isolated from human pituitary extracts. It consists of 32 amino acids with a proposed sequence identical to residues 7--38 of corticotropin (ACTH). The peptide has been synthesized by the solid-phase method. The melanotropic activity of the peptide is estimated to be 30% of the potency of ACTH. It is devoid of corticosteroidogenic activity but is able to inhibit ACTH-stimulated corticosterone production in isolated rat adrenal cells.
Differences among 100-A filamentilament subunits from different cell types.
Authors: Bennett G S , Fellini S A , Croop J M , Otto J J , Bryan J , Holtzer H
Published: May 2006

The protein subunit of 100-A filaments constitutes approximately 50% of the cytoskeleton protein of chick fibroblasts. In addition to the 43,000-dalton protein (constitutive actin) common to all cell types, fibroblast cytoskeletons contain a 58,000-dalton protein likely to be the 100-A filament subunit, whereas smooth muscle contains, instead, a 55,000-dalton protein. Additional differences among 100-A filaments are shown by immunofluorescence using antibodies angainst chick fibroblast 58,000-dalton component (anti-F58K) and against chick brain 100-A filament subunits (anti-BF). Anti-F58K binds to 100-A filaments in chick fibroblasts, presumptive myoblasts, chondroblasts, pigment cells, and neurons, but not to 100-A filaments in mouse or human fibroblasts. This antibody stains cables of 100-A filaments induced by sequentially treating cells with cytochalasin B and Colcemid. Anti-BF binds only to neurofilaments and not to 100-A filaments of other cell types studied. Absorption or antibodies with purified subunits from gizzard 100-A filaments eliminates binding of anti-F58K to the filaments of all cell types but does not diminish binding of anti-BF to neurofilaments. Various IgGs also bind nonspecifically to induced cables of 100-A filaments. The problem of nonspecific binding of labeled antibodies, as well as the problem of cell and species specificity of the 100-A filaments, is discussed.
Processing of 60,000-dalton sarc gene protein synthesized by cell-free translation.
Authors: Kamine J , Buchanan J M
Published: May 2006

In this report we show that antiserum prepared against the Mr60,000 transformation-specific antigen of Rous sarcoma virus immunoprecipitates both the Mr60,000 and Mr 25,000 transformation-specific proteins that are synthesized by cell-free translation of virion RNA; however, in the cell-free system the Mr 60,000 protein appears to be synthesized as a precursor that is approximately Mr 2000 larger than the [35S]-methionine-labeled protein immunoprecipitated from Rous sarcoma virus-infected cells. Peptide mapping of the cell-free translation product and of this cellular protein has confirmed that they are structurally related to one another. The addition of membrane vesicles to the reticulocyte lysate system during translation specifically cleaves a Mr 2000 segment from the Mr 60,000 protein so that it comigrates with the cellular species. Secretory proteins and probably at least some integral membrane proteins are synthesized with short hydrophobic signal sequences at their NH2 terminus. Two facts suggest that the segment lost from the Mr 60,000 transformation-specific protein is a signal sequence: (i) the membrane vesicles process the Mr 60,000 protein only during translation, and (ii) the processed protein is sequestered by the vesicles.
Activation of endogenous type C virus in BALB/c mouse cells by herpesvirus DNA
Authors: Boyd Ann L. , Derge Jeffery G. , Hampar Berge
Published: May 2006

Several virion and nonvirion DNAs were tested for the ability to activate endogenous type C virus in BALB/c-derived mouse cells using the calcium precipitation technique. The DNAs from all herpesviruses tested activated xenotropic type C virus synthesis. These included DNAs from herpes simplex virus types 1 and 2, Epstein-Barr virus, human cytomegalovirus, SA8 virus, infectious bovine rhinotracheitis virus, pseudorabies virus, and herpes saimiri virus (M-DNA). In contrast, DNAs from vaccinia virus, simian virus 40, primate cells, bacteria, mycoplasma, and salmon sperm showed no ability to activate type C virus when tested under the same conditions. Several herpesviruses and vaccinia virus, which were highly infectious for the BALB/c cells used, were tested for their ability to activate type C virus after UV irradiation. All herpesviruses tested were positive, while vaccinia virus was negative. Unirradiated simian virus 40 also showed no ability to activate type C virus. Activation of type C virus by DNA from herpes simplex virus was observed after shearing or sonication of the DNA to an average size of 3 × 10
6
daltons, but was not observed with DNA sonicated to an average size of 1 × 10
6
daltons. Alkali denaturation of DNA from herpes simplex virus or treatment with DNase, but not RNase, destroyed its ability to activate type C virus, as did crosslinking of the DNA with 4,5′,8-trimethylpsoralen (psoralen) and light.
Fetal phenotypic expression by adult rat hepatocytes on collagen gel/nylon meshes.
Authors: Sirica A E , Richards W , Tsukada Y , Sattler C A , Pitot H C
Published: May 2006

Hepatocytes from adult rats were maintained in primary culture for up to 10-13 days on nylon meshes coated with a thin layer of rat tail collagen gel. Their ultrastructure closely resembled that of the liver parenchymal cell in vivo, but hepatocytes in late culture exhibited a pronounced buildup of microfilaments beneath their apical cell surface. Hepatocytes in early and late cultures secreted albumin, transferrin, and alpha1-acid glycoprotein into the medium; they exhibited a 7- to 10-fold induction of tyrosine aminotransferase activity by dexamethasone; and they expressed an alkaline phosphatase that was similar to that of normal rat liver with respect to its inhibition by the liver enzyme inhibitor L-homoarginine. In addition, the hepatocytes in culture demonstrated phenotypic changes characteristic of fetal liver parenchymal cells. These changes, which paralleled an increase in DNA synthesis, included the expression of and linear increase in the activity of the fetal liver cell enzyme gamma-glutamyl transpeptidase, an increased production of alpha1-fetoprotein, and a change in the substrate specificity of fructose-bisphosphate aldolase to that of the fetal liver isozyme.
Resonance Raman studies of bathorhodopsin: Evidence for a protonated Schiff base linkage
Authors: Eyring Gregory , Mathies Richard
Published: May 2006

A dual beam pump/probe technique has been used with a 585-nm probe wavelength to obtain maximal resonance enhancement of the Raman lines of bathorhodopsin in a photostationary steady-state mixture at -160°C. These studies show that bathorhodopsin has a protonated Schiff base vibration at 1657 cm
-1
which shifts upon deuteration to 1625 cm
-1
. Within our experimental error (±2 cm
-1
) these frequencies are identical to those observed in rhodopsin and isorhodopsin. These effects show that the strength of the C=N bond and the degree of protonation of the Schiff base nitrogen are the same in bathorhodopsin, rhodopsin, and isorhodopsin. The implication of these results for the structure of the retinal chromophore in bathorhodopsin are discussed. The resonance Raman spectrum of pure bathorhodopsin has been generated by accurately subtracting the residual contributions of rhodopsin and isorhodopsin from spectra of the low temperature photostationary mixture. Bathorhodopsin is found to have lines at 853, 875, 920, 1006, 1166, 1210, 1278, 1323, 1536, and 1657 cm
-1
. Also, by using an intensified vidicon detector, we have observed Raman scattering from bathorhodopsin at room temperature by generating a photostationary steady state with pulsed laser excitation. At room temperature the three characteristic lines of bathorhodopsin are found at 858, 873, and 920 cm
-1
. The fact that the frequencies of these bathorhodopsin lines are nearly identical at both temperatures implies that the retinal conformation in bathorhodopsin formed at -160°C is the same as that formed at room temperature.
Incorporation of glucocerebrosidase into Gaucher's disease monocytes in vitro.
Authors: Dale G L , Kuhl W , Beutler E
Published: May 2006

Several carriers were evaluated for use in the delivery of exogenous glucocerebrosidase to monocytes from Gaucher's disease patients. Only gamma globulin-coated, resealed erythrocytes proved to be an effective vehicle for enzyme delivery. Glucocerebrosidase added in this manner normalized intracellular enzyme levels for at least 18 hr. In this model system for the study of enzyme replacement therapy, soluble enzyme, enzyme in uncoated resealed erythrocytes, and enzyme incorporated into liposomes were ineffective.
Prolyl-leucyl-glycinamide, cyclo(leucylglycine), and derivatives block development of physical dependence on morphine in mice.
Authors: Walter R , Ritzmann R F , Bhargava H N , Flexner L B
Published: May 2006

Pro-Leu-Gly-NH2 (MIF) and several structural analogues, all injected in 50-microgram doses daily in mice receiving morphine chronically, were found to prevent development of physical dependence as measured by changes in body temperature associated with naloxone-induced withdrawal. Dose-response studies, using again a protocol of daily injections of peptide at 50, 5, 0.5, 0.05, 0.005 microgram per mouse revealed MIF and cyclo(Leu-Gly) to be the most potent peptides and to be effective in blocking physical dependence to morphine at a dose as low as 0.5 and 0.05 microgram per mouse, respectively. The benzyloxycarbonyl derivative of MIF, Pro-Leu, and Pro- -Leu exhibited significant activities down to a dose of 5 microgram of peptide per mouse.
Characterization of the reverse transcriptase of a type C RNA virus produced by a human lymphoma cell line
Authors: Goodenow Robert S. , Kaplan Henry S.
Published: May 2006

The reverse transcriptase (RNA-dependent DNA nucleotidyltransferase) of the type C RNA virus produced by the human lymphoma cell line SU-DHL-1 was purified by ion-exchange chromatography of SU-DHL-1 culture fluids and repetitive affinity chromatography on poly(rC)·agarose, as were the polymerases of several other type C viruses. The DHL-1 enzyme used template-primers at levels expected of a viral reverse transcriptase, and sodium dodecyl sulfate gel electrophoretic analysis of radioiodinated DHL-1 enzyme revealed a peak at a position corresponding to those of several other type C viral reverse transcriptases (namely, at 72,000-78,000 daltons). The purified enzyme was partially neutralized by antibodies specific for the reverse transcriptase of simian sarcoma virus. Two-dimensional analysis on thin-layer cellulose plates of tryptic hydrolysates of the radioiodinated enzymes of several viruses revealed that six peptides are common to the polymerases of simian sarcoma virus, gibbon ape leukemia virus, baboon endogenous virus, and the DHL-1 virus, and that two to four peptides are unique to each of these enzymes. The DHL-1 viral reverse transcriptase appears to be most closely related structurally to the enzymes of simian sarcoma virus, gibbon ape leukemia virus, and baboon endogenous virus. However, the DHL-1 viral enzyme differed from any one or combination of the other subhuman primate viral enzymes by virtue of its unique peptides. The implications of these findings with respect to the probable origin of the DHL-1 virus are discussed.
Identification of iron-sulfur centers in the iron-molybdenum proteins of nitrogenase.
Authors: Kurtz D M , McMillan R S , Burgess B K , Mortenson L E , Holm R H
Published: May 2006

The core extrusion method has been applied to the determination of the type ([2Fe-2S], [4Fe-4S]) and number of iron-sulfur centers in the FeMo proteins of the nitrogenases from Clostridium pasteurianum and Azotobacter vinelandii. The method involves extrusion with o-xylyl-alpha, alpha'-dithiol, ligand exchange of the extrusion products with p-CF3C6H4SH (RFSH), and identification and quantitation of the resultant [FenSn(SRF)4]2- complexes (n = 2,4) by 19F NMR spectroscopy. In hexamethylphosphoramide/water, 4:1 (vol/vol), 49-56% of the Fe content was extruded as [Fe4S4(SRF)4]2-, corresponding to 3,4-4.0 Fe4S4 cores per alpha 2 beta 2 subunit complex. The extruded iron does not arise from the FeMo cofactor, separate examination of which detected no extrusion products, and corresponds to 90-103% of noncofactor iron. No significant quantity of Fe2S2 cores was extruded. These results indicate the presence of four [4Fe-4S] centers per alpha 2 beta 2 subunit complex in preparations undepleted in iron. There are two main structural populations of iron atoms in these proteins, those in the cubane-type Fe4S4 cores and those in the FeMo cofactor.
Comparison of amino acid sequence of bovine coagulation Factor IX (Christmas Factor) with that of other vitamin K-dependent plasma proteins.
Authors: Katayama K , Ericsson L H , Enfield D L , Walsh K A , Neurath H , Davie E W , Titani K
Published: May 2006

The amino acid sequence of bovine blood coagulation Factor IX (Christmas Factor) is presented and compared with the sequences of other vitamin K-dependent plasma proteins and pancreatic trypsinogen. The 416-residue sequence of Factor IX was determined largely by automated Edman degradation of two large segments, containing 181 and 235 residues, isolated after activating Factor IX with a protease from Russell's viper venom. Subfragments of the two segments were produced by enzymatic digestion and by chemical cleavage of methionyl, tryptophyl, and asparaginyl-glycyl bonds. Comparison of the amino acid sequences of Factor IX, Factor X, and Protein C demonstrates that they are homologous throughout. Their homology with prothrombin, however, is restricted to the amino-terminal region, which is rich in gamma-carboxyglutamic acid, and the carboxyl-terminal region, which represents the catalytic domain of these proteins and corresponds to that of pancreatic serine proteases.
Acidic polypeptides can assemble both histones and chromatin in vitro at physiological ionic strength.
Authors: Stein A , Whitlock J P , Bina M
Published: May 2006

We provide evidence that nucleosomes can assemble in vitro at physiological ionic strength (0.1-0.2 M NaCl/10 mM Tris . HCl, pH 8.0) in the absence of "assembly factors" and that poly(glutamic acid) greatly facilitates chromatin assembly under these conditions. We also show that in the presence of either poly(glutamic acid) or poly(aspartic acid), core histones assemble into octamers at physiological ionic strength. We suggest that it is a property of histones to assemble into octamers upon their interaction with macromolecules containing regions of high negative charge density, and we discuss several implications of this property.
Structure and specific sequences of avian erythroblastosis virus RNA: Evidence for multiple classes of transforming genes among avian tumor viruses
Authors: Bister Klaus , Duesberg Peter H.
Published: May 2006

Two major RNA species were found in several clonal isolates of avian erythroblastosis virus (AEV) and avian erythroblastosis-associated helper virus (AEAV) complexes: one of 8.7 kilobases (kb), the other of 5.5 kb. The 5.5-kb species was identified as AEV RNA because (
i
) it was absent from non-transforming AEAV isolated from the same virus complex, (
ii
) it was present in complexes of AEV and different helper viruses, and (
iii
) its structure is similar to that of avian acute leukemia viruses of the MC29 group. Molecular hybridization indicated that 54% of AEV RNA is specific and 46% is related to other viruses of the avian tumor virus group, particularly to AEAV, therefore termed group-specific. The genetic structure of AEV RNA was deduced by mapping oligonucleotides representing specific and group-specific sequences and by comparing the resulting map to maps of AEAV and of other avian tumor viruses derived previously. AEV RNA contains a
gag
gene-related, 5′ group-specific section of 1 kb, an internal AEV-specific section of 3 kb unrelated to any other viral RNA tested, and a 3′ group-specific section of 1.5 kb. The 5′ section of AEV RNA is closely related to analogous 5′ sections of the MC29 group viruses and is homologous with a 5′ RNA section that is part of the
gag
gene of AEAV. The 3′ section is also shared with AEAV RNA and includes a variant C-oligonucleotide near the 3′ end that is different from the highly conserved counterparts of all other exogenous avian tumor viruses. By analogy with Rous sarcoma virus and the acute leukemia viruses of the MC29 group, the internal specific section of AEV RNA is thought to signal a third class of
onc
genes in avian tumor viruses. Comparisons with AEAV and the MC29 group viruses suggest that both the 5′
gag
-related and the internal specific RNA sections of AEV are necessary for
onc
gene function.
In vitro production of 1,25-dihydroxyvitamin D3 by rat placental tissue.
Authors: Tanaka Y , Halloran B , Schnoes H K , DeLuca H F
Published: May 2006

Weanling female rats were fed a vitamin D-deficient diet for 4 months until they reached maturity. They were mated with normal, vitamin D-replete male rats and, at 20 days of pregnancy, the female rats were killed and their placentae were removed, homogenzied, and incubated with 25-hydroxyvitamin D3. The incubation mixtures were extracted and the extracts were subjected to Sephadex LH-20 column chromatography followed by high-pressure liquid chromatography. The 1,25-dihydroxyvitamin D3 region of the high-pressure liquid chromatogram was recycled to purity and the structure of the product was identified as 1,25-dihydroxyvitamin D3 by ultraviolet absorption spectrophotometry and by mass spectrometry. Thus it is now evident that placenta, in addition to renal tissue, is capable of converting 25-hydroxyvitamin D3 to the hormonal form of vitamin D, 1,25-dihydroxyvitamin D3.
Purification of two spectrin-binding proteins: biochemical and electron microscopic evidence for site-specific reassociation between spectrin and bands 2.1 and 4.1.
Authors: Tyler J M , Hargreaves W R , Branton D
Published: May 2006

Two peripheral proteins of the human erythrocyte membrane that are capable of forming a stable complex with spectrin have been purified. The proteins, band 2.1 (Mr 210,000) and band 4.1 (Mr 82,000), are water soluble and exist as monomers in solution. Both exhibit strong, specific binding to purified spectrin molecules as determined by cosedimentation in sucrose gradients and both enhance binding to spectrin-depleted, inside-out vesicles that have been stripped of bands 2.1 and 4.1. Rotary replicas of bound material reveal site-specific associations among native, but not heat-denatured, molecules.
Evidence for a large internal pressure in biological membranes.
Authors: Conrad M J , Singer S J
Published: May 2006

We have developed a method (hygroscopic desorption) for measuring the binding of small molecules to membranes. With this method, we have found that the binding of the amphipathic compounds chlorpromazine, 2,4-dinitrophenol, and 1-decanol to various cell membranes is remarkably low, with partition coefficients, Kp, no larger than about 0.1. On the other hand, with phospholipid vesicles of large or small diameters, Kp values for these compounds were much larger. The results suggest that there exists in membranes a large internal pressure that excludes the amphipaths from the membranes and that does not exist in phospholipid vesicles.
Tumor-enhancing suppressor activator T cells in spleens and thymuses of tumor immune mice.
Authors: Hellström I , Hellström K E , Bernstein I D
Published: May 2006

Cells from the spleens and thymuses of BALB/c mice whose Moloney sarcoma virus (MSV)-induced, primary sarcomas have regressed 2-3 months earlier ("MSV regressors") or are in the process of regressing can, when adoptively transferred to syngeneic mice given MSV at the age of 20 days, prevent the natural regression of the MSV sarcomas in the recipient mice. The cells responsible for this tumor-enhancing effect express the Thy 1 marker. They are not demonstrable in the thymuses of normal untreated mice or in mice that have either been immunized against or are bearing methylcholanthrene-induced sarcomas. The tumor-enhancing cells are not destroyed after administration of 400 rads (1 rad = 1.00 x 10(-2) J/kg) of whole body radiation. However, the effect of the irradiated cells is seen only in the presence of a nonirradiated T-cell population, represented in the thymuses of normal control mice, with which we postulate that they interact. Studies on a transplantable, chemically induced, murine leukemia virus antigen-negative sarcoma, MCA-1460, further support the concept that relatively radioresistant thymus cells from immune mice can enhance tumor outgrowth by interacting with radiosensitive T cells that are present in nonimmune mice.
Mouse mammary tumor virus genome expression in chemical carcinogen-induced mammary tumors in low- and high-tumor-incidence mouse strains
Authors: Dusing-Swartz S. , Medina D. , Butel J. S. , Socher S. H.
Published: May 2006

Involvement of mouse mammary tumor virus (MMTV) in 7,12-dimethylbenz[
a
]anthracene (DMBA)-induced mammary tumorigenesis was investigated in low- (BALB/c) and high- (BALB/cfC3H) mammary-tumor-incidence mouse strains. Both strains contain endogenous MMTV integrated into the cellular genome. Additionally, BALB/cfC3H mice are infected with exogenous MMTV-S which is responsible for a higher incidence of mammary tumors in breeding females. Administration of DMBA to virgin mice of both strains resulted in a moderate frequency of mammary tumors within 40 wk after treatment. No differences were found in DMBA-induced tumor incidences at 18 wk (6% and 7%) or at 38 wk (29% and 36%) after treatment of BALB/c and BALB/cfC3H mice, respectively. Expression of MMTV in these tumors was examined by assaying for the presence of MMTV RNA by hybridization using MMTV-specific cDNA and by immunohistochemical staining utilizing antibodies against MMTV 52,000-dalton glycoprotein, gp52, and 28,000-dalton internal protein, p28. Of 16 BALB/c tumors assayed, 11 did not contain detectable levels of MMTV RNA and the remaining 5 tumors contained only low levels (0.0005-0.0010%) of viral RNA. Importantly, MMTV RNA was not detected in 5 of 27 BALB/cfC3H tumors. The other BALB/cfC3H tumors contained quantities of MMTV RNA ranging from 0.0006 to 0.4170%. Most BALB/cfC3H tumors with detectable levels of MMTV RNA also synthesized viral proteins gp52 and p28. Thus, expression of the complete MMTV genome is not requisite for maintenance of the tumor phenotype in DMBA-induced mammary tumors in either BALB/c or BALB/cfC3H virgin mice under 1 year of age.
In vitro maturation of mesencephalic dopaminergic neurons from mouse embryos is enhanced in presence of their striatal target cells.
Authors: Prochiantz A , di Porzio U , Kato A , Berger B , Glowinski J
Published: May 2006

Long-term survival of mesencephalic and striatal cells from mouse embryos in dissociated primary cultures is described. Catecholaminergic neurons in mesencephalic culutres were identified histochemically and by measuring [3H]dopamine uptake and synthesis from [3H]tyrosine. According to experiments using specific inhibitors of catecholamine uptake, at least two-thirds of the catecholaminergic neurons are dopaminergic. These neurons differentiated whether or not striatal target cells were present, but striatal cells stimulated the development of the dopaminergic neurons. [3H]Dopamine uptake was increased by at least 2-fold regardless of the age of the cocultures (4-15 days). Enhanced [3H]dopamine synthesis was also observed (at least 2-fold) at later times (12-15 days).
Induction of acetylcholine receptors on cultured skeletal muscle by a factor extracted from brain and spinal cord.
Authors: Jessell T M , Siegel R E , Fischbach G D
Published: May 2006

Extracts of chicken brain and spinal cord increase the total number of acetylcholine receptors and the number of acetylcholine receptors and the number of receptor clusters on uninnervated skeletal myotubes in culture. The active component in these extracts may be unique to neural tissue. Spinal cord cells grown in culture contain the active factor and they secrete it into the medium. Most of the activity is associated with a small molecule possibly a peptide. Such a factor may be responsible for the clustering of receptors at newly formed nerve-muscle synapses.
Multiple phosphorylation sites in protein I and their differential regulation by cyclic AMP and calcium.
Authors: Huttner W B , Greengard P
Published: May 2006

The phosphorylation of protein I, a specific neuronal protein, has been found to be regulated both by cyclic AMP (cAMP) and by calcium, in intact as well as in lysed synaptosome preparations from rat brain. In order to determine the phosphorylation site(s) of protein I that were regulated by cAMP and calcium, protein I was purified after it was phosphorylated under various conditions. This purified protein I was then subjected either to peptide mapping after limited proteolysis in sodium dodecyl sulfate/polyacrylamide gels or to tryptic fingerprinting. 8-Br-cAMP selectively increased the phosphorylation of the same protein I peptide fragment in both intact and lysed synaptosomes. Depolarization-induced calcium influx into intact synaptosomes, or the addition of calcium to lysed synaptosomes, caused a stimulation of the phosphorylation not only of this peptide but also of other distinct peptides. Differential regulation by cAMP and calcium of the phosphorylation of multiple sites on the same neuronal protein may provide a molecular basis for interactions between these two second-messenger systems in certain nerve terminal functions.
Synthesis of simian virus 40 t antigen in Escherichia coli.
Authors: Roberts T M , Bikel I , Yocum R R , Livingston D M , Ptashne M
Published: May 2006

Plasmids are constructed by using recombination in vitro according to Roberts, T.M., Kacich, R. & Ptashne, M. (1979) Proc. Natl. Acad. Sci. USA 76, 760-764 in which the t antigen gene of simian virus 40 is fused to a promoter of the Escherichia coli lac operon. In the fusions, transcription commences at the lac promoter, and, in some of the fusions, translation begins at the ATG initiator codon of the t gene. This translation is directed most efficiently by those plasmids in which the lac sequences abut the t gene such that a hybrid ribosome binding is encoded. In this case, the Shine-Dalgarno sequence is of lac origin but the ATG derives from the t gene. translation from this initiator codon is greatly decreased if the lac sequences are separated from the ATG by 17 base pairs and is abolished if the AT of this triplet is deleted. Cells bearing the productive fusions synthesize a 20,000-dalton protein with t antigenic determinants. This protein has an isoelectric point(s) indistinguishable from that of t antigen isolated from simian virus 40-transformed cells. Moreover, a partial sequence of the amino-terminal region of the bacterial product is that predicted for authentic t antigen. We conclude that these bacteria are for authentic t antigen. We conclude that these bacteria are producing a protein, the sequence of which is identical to that of authentic t antigen unfused to other polypeptides.
Coupling of opiate receptors to adenylate cyclase: requirement for Na+ and GTP.
Authors: Blume A J , Lichtshtein D , Boone G
Published: May 2006

Inhibition of the adenylate cyclase activity in homogenates of mouse neuroblastoma-glioma hybrid cells (NG108-15) by the opioid peptide [D-Ala2,Met5]enkephalin amide (AMEA) requires the presence of Na+ and GTP. In this process, the selectivity for monovalent cations is Na+ greater than or equal Li+ greater than K+ greater than choline+; ITP will replace GTP but ATP, UTP, or CTP will not. The apparent Km for Na+ is 20 mM and for GTP it is 1 microM. Under saturating Na+ and GTP conditions, the apparent Ki for AMEA-directed inhibition is 20 nM for basal and 100 nM for prostaglandin E1-activated adenylate cyclase activity. For both cyclase activities, maximal inhibition is only partial (i.e., approximately 55% of control in each case). In intact viable NG108-15 cells, the decrease in basal and prostaglandin E1-stimulated intracellular cyclic AMP concentrations by AMEA is also dependent upon extracellular Na+. The enkephalin-directed reductions in cyclic AMP concentrations are at least 75%. The specificity of the monovalent cation requirement for enkephalin action on intact cells is the same as for enkephalin regulation of homogenate adenylate cyclase activity. Based on these data, a model is presented in which the transfer of information from opiate receptors to adenylate cyclase requires active separate membrane components, which correspond to the sites of action of Na+ and GTP in this process.
Phosphoethanolamine as a growth factor of a mammary carcinoma cell line of rat.
Authors: Kano-Sueoka T , Cohen D M , Yamaizumi Z , Nishimura S , Mori M , Fujiki H
Published: May 2006

We have identified phosphoethanolamine as the pituitary-derived growth-promoting material that specifically stimulates the rat mammary carcinoma cell line 64-24. We have been studying the growth characteristics of the 64-24 cell line, which was isolated from a highly hormone-dependent tumor and which retains in culture many characteristics of the original tumor. Previously, crude bovine pituitary extract was shown to contain a significant amount of growth-stimulating activity for these cells, and a growth factor from this extract was purified to homogeneity. This report describes the identification of the growth factor as phosphoethanolamine. Further, the biological activity of phosphoethanolamine was found to be virtually identical to that of the purified growth factor. A possible role of phosphoethanolamine in the growth of mammary tumor cells as well as of normal mammary epithelial cells and other tissues is discussed.
Pyrolysis products from amino acids and protein: Highest mutagenicity requires cytochrome
P
1
-450
Authors: Nebert Daniel W. , Bigelow Sanford W. , Okey Allan B. , Yahagi Takie , Mori Yuko , Nagao Minako , Sugimura Takashi
Published: May 2006

Pyrolysis products of proteins and amino acids are highly mutagenic, but metabolism of these chemicals by rat liver subcellular fractions is known to be required for production of the mutagenic intermediates. We examined the mutagenesis of seven purified pyrolysis products from tryptophan, lysine, glutamic acid, and soybean globulin with
Salmonella typhimurium
strain TA98 in the presence of liver fractions from genetically “responsive” C57BL/6N and
Ah
b
/
Ah
d
or “nonresponsive” DBA/2N and
Ah
d
/
Ah
d
mice that had been pretreated
in vivo
with benzo[
a
]pyrene. For all pyrolysis products tested, mutagenesis is 2-fold to more than 1000-fold greater with C57BL/6N and
Ah
b
/
Ah
d
than with DBA/2N or
Ah
d
/
Ah
d
liver fractions. A sucrose density gradient assay for detecting the
Ah
regulatory gene product, the receptor, was studied with C57BL/6N hepatic cytosol. At levels 100 times in excess of [1,6-
3
H]2,3,7,8-tetrachlorodibenzo-
p
-dioxin, nonlabeled 2,3,7,8-tetrachlorodibenzo-
p
-dioxin, 3-methylcholanthrene, and β-naphthoflavone (inducers of cytochrome
P
1
-450) are able to displace the radioligand from its hepatic cytosolic receptor; four pyrolysates from tryptophan, glutamic acid, and soybean globulin did not have this capacity.

These data indicate that the pyrolysis products tested, although not effective as inducers of cytochrome
P
1
-450, are most mutagenic when metabolized by
P
1
-450. Potent
P
1
-450 inducers—present in pyrolysates during the combustion process—might be present in quantities insufficient to initiate mutagenesis or carcinogenesis but might have a synergistic action, or act as “comutagens” or “cocarcinogens,” with the N-containing heterocyclic pyrolysis products. A
quantitative
relationship between mutagenic and carcinogenic potency of these pyrolysis products remains, however, to be demonstrated.
Widespread distribution of protein I in the central and peripheral nervous systems.
Authors: De Camilli P , Ueda T , Bloom F E , Battenberg E , Greengard P
Published: May 2006

Protein I, a naturally occurring substrate for cyclic AMP-dependent and calcium-dependent protein kinases, previously has been found only in mammalian brain, where it has been demonstrated to be located in neurons. Various tissues and organs outside the brain have now been examined for the possible occurrence of protein I, by using both an immunohistochemical approach and a chemical procedure involving radioimmunolabeling of polyacrylamide gels. Protein I has been found in the inner plexiform layer of the retina, in the posterior pituitary, and in the autonomic nervous system. In tissue composed predominantly of cells other than nerve cells, immunoreactivity was present only where innervation was present. Protein I appeared to be localized in some, but not all, nerve terminals and synapses.
Trypsin liberates an arginine vasopressin-like peptide and neurophysin from a Mr 20,000 putative common precursor.
Authors: Russell J T , Brownstein M J , Gainer H
Published: May 2006

Although the hypothesis that vasopressin and its associated neurophysin are synthesized together in one macromolecular common precursor was put forward more than a decade ago, direct conformation of this hypothesis has been lacking. A [35S]cysteine-labeled putative precursor for vasopressin-related neurophysin (Mr 20,000, pI 6.1) has been isolated from the supraoptic nuclei of rats. This precursor was subjected to limited proteolysis with trypsin which produced a Mr 10,000 protein and peptide products. The former was identified as neurophysin on the basis of its pH-dependent affinity for vasopressin and its behavior in isoelectric focusing systems (pI 4.6-4.8). The tryptic peptides proved to be vasopressin-like because they: (i) were rich in cysteine, (ii) comigrated with vasopressin on gel filtration columns in 6 M guanidine HCl, (iii) bound to a neurophysin-Sepharose affinity column at pH 5.7, and (iv) were recognized by antibodies against vasopressin. These data on the Mr 20,000, pI 6.1 protein represent direct experimental evidence for a candidate for the common precursor of vasopressin and neurophysin. We propose that this common precursor be called "propressophysin."
Electron-nuclear double resonance of the hydrogen peroxide compound of cytochrome c peroxidase: identification of the free radical site with a methionyl cluster.
Authors: Hoffman B M , Roberts J E , Brown T G , Kang C H , Margoliash E
Published: May 2006

The results of electron-nuclear double resonance and electron paramagnetic resonance (EPR) studies on the hydrogen peroxide compound of yeast cytochrome c peroxidase are inconsistent with previous proposals for the source of the EPR signal in this compound, in particular with its identification with an aromatic amino acid radical such as would arise by oxidation of a tryptophanyl side chain. The present observations lead us to propose that the EPR signal is associated with a cluster containing at least one methionine and in which proximate side chains share the charge created by loss of one electron.
Isolation and structure of pro-methionine-enkephalin: Potential enkephalin precursor from porcine hypothalamus
Authors: Huang Wei-Yong , Chang Robert C. C. , Kastin Abba J. , Coy David H. , Schally Andrew V.
Published: May 2006

A hexapeptide that we have named pro-methionine-enkephalin has been isolated from acid extracts of porcine hypothalami and found to have the amino acid sequence H-Tyr-Gly-Gly-Phe-Met(O)-Arg-OH. This peptide is not a fragment of either porcine β-lipotropin or β-endorphin, which suggests that it could be a precursor of [Met]enkephalin in the brain by a pathway differing from the one usually postulated. The existence of still larger precursors of [Met]enkephalin, perhaps related to the recently reported α-neo-endorphin, is strongly implied by these studies. Synthetic H-Tyr-Gly-Gly-Phe-Met-Arg-OH exhibited low, but significant, opiate activity
in vitro
.
Internal motions in DNA.
Authors: Hogan M E , Jardetzky O
Published: May 2006

We have measured the 31P and 1H NMR parameters of fractionated double-stranded DNA fragments 600, 300, and 150 base pairs long. The results indicate that, inside an intact double helix, both the deoxyribose and the sugar-phosphate backbone of the helix fluctuate substantially from their equilibrium geometry. The time constant for those coupled motions is of the order of 1 nsec.
Quantitative relationship between β-adrenergic receptor number and physiologic responses as studied with a long-lasting β-adrenergic antagonist
Authors: Terasaki Wesley L. , Linden Joel , Brooker Gary
Published: May 2006

The aminobenzyl analog of propranolol, 1- (
p
-amino-α,α-dimethylphenethylamino)-3-(1-naphthoxy)-2- propanol, was synthesized and found to be a potent β-adrenergic blocking agent. The β-adrenergic receptors of cultured rat C6 glioma cells (2B clone) as assessed by [
125
I]iodohydroxybenzylpindolol binding were decreased 50 and >95% after pretreatment with 8 nM and 1 μM aminobenzylpropranolol, respectively. Unlike propranolol, aminobenzylpropranolol displayed a prolonged blockade of receptors that was maintained during several hours of washing. [
125
I]Iodohydroxybenzylpindolol saturation binding experiments in cells exposed to aminobenzylpropranolol and subsequently washed indicated that the compound effectively diminished receptor number with no change in the affinity of the remaining receptors for iodohydroxybenzylpindolol. Aminobenzylpropranolol inhibited catecholamine-stimulated intracellular cyclic AMP accumulation; with increasing blockade, isoproterenol dose—response curves became progressively shifted to the right but the maximal response was unaltered. Aminobenzylpropranolol inhibited the β-adrenergic contractile response in atria isolated from rats and guinea pigs. Treatment with 0.1 and 10 μM aminobenzylpropranolol produced decreases of 0.5 and 2 orders of magnitude in the contractile potency of isoproterenol. As in glioma cells, aminobenzylpropranolol failed to decrease the maximal response to isoproterenol. The effects of aminobenzylpropranolol persisted during extensive washing of atria (up to 17 hr). Repeated exposures to isoproterenol at concentrations sufficient to produce maximal tension development also failed to alleviate the blockade. The inotropic potency of histamine in guinea pig atria was not affected by aminobenzylpropranolol. These data suggest that catecholamines are capable of eliciting full biological responses in glioma cells and isolated atria even though the great majority of β-adrenergic receptors are persistently blocked.
Structure of a bacterial photosynthetic membrane
Authors: Miller Kenneth R.
Published: May 2006

The internal photosynthetic membranes of a photosynthetic bacterium,
Rhodopseudomonas viridis
, have been studied with a variety of electron microscope techniques. The membranes are composed of a sheet of apparently identical subunits arranged in a hexagonal fashion. The individual subunits repeat at a distance of 110 Å. Optical transforms have been used to enhance micrographs of this ordered membrane, and the images synthesized in this way show details of each subunit. The individual subunits are asymmetric, differing slightly in appearance at the outer and inner surfaces of the membrane, and these surface patterns seem to be combined in the image of the thylakoid membrane in negative stain. These studies fix a maximum size for the photosynthetic unit of
R. viridis
and suggest the suitability of this membrane for further diffraction analysis.
Kinetics of the cycloheximide-induced phase changes in the biological clock in
Gonyaulax
Authors: Walz Brigitte , Sweeney Beatrice M.
Published: May 2006

Cycloheximide, an inhibitor of protein synthesis on cytoplasmic ribosomes in eukaryotes, is shown to shift the phase of the circadian rhythm in stimulated bioluminescence in the marine dinoflagellate
Gonyaulax polyedra
. Kinetic analysis of the phase changes shows that this effect may be subdivided into two distinctly different and well-separated parts. The first (early) phase change occurs with 15-min exposure to cycloheximide and is saturated at low concentrations (≈10 nM). The second (late) phase changes requires about 150 min of exposure to cycloheximide and is saturated at 0.36 μM cycloheximide. Twenty-times-higher concentrations cause no further phase changes. The magnitudes of both early and late phase changes depend on the time of day when the cells are exposed to cycloheximide. Early phase changes vary from 5 hr advance at circadian time (CT) 20 to 1 hr delay at CT 12; late phase changes are larger, the maximal advance being 12 hr at CT 16 and the greatest delay, 10 hr at CT 14. It is proposed that the early phase changes are caused by alterations in the ion distribution across membranes as a consequence of the permeation of cycloheximide. Late phase changes may be the result of inhibition of protein synthesis.

The phase response curve for the late phase change is identical to that obtained with saturating light pulses in otherwise constant darkness in
Gonyaulax
. Maximal phase changes drive the clock into the part of the circadian cycle between CTs 4 and 9. Perturbations in this part of the circadian cycle are without effect on phase. Incubation of
Gonyaulax
with cycloheximide for a critical duration at a critical time induces arhythmicity, but longer exposures to the inhibitor at the same time do not. This observation suggests the existence of a singularity in the circadian clock of
Gonyaulax
.