bluuebunny/crossref_metadata_2025 huggingface dataset:
The Pancreatic β-Cell Recognition of Insulin Secretagogues: Does Cyclic AMP Mediate the Effect of Glucose?
Authors: Hellman B. , Idahl L.-Å. , Lernmark Å. , Täljedal I.-B.
Published: May 2006

Insulin release and the content of cAMP were studied in microdissected pancreatic islets of noninbred
ob/ob
(obese) mice. In the absence of 3-isobutyl-1-methylxanthine, a phosphodiesterase inhibitor, 20 mM glucose had no effect on cAMP save a very small initial rise detectable by a freeze-stop perifusion technique only. However, combined with this methylxanthine, 20 mM glucose produced significant increases of cAMP both in perifused islets and in islets conventionally incubated in closed vials. Glucose shared this capacity to raise the cAMP level with D-glyceraldehyde and 1,3-dihydroxyacetone. Isobutylmethylxanthine (0.05-1.0 mM) or 5 μg/ml of cholera toxin, an activator of adenylate cyclase, also increased the islet cAMP level; the effects of the methylxanthine, whether or not combined with cholera toxin, were potentiated by glucose. Isobutylmethylxanthine (0.05-1.0 mM) or 5 μg/ml of cholera toxin potentiated insulin release in response to 20 mM glucose. However, only 0.5-1.0 mM isobutylmethylxanthine stimulated insulin release in the presence of 3 mM glucose, whereas 0.05-0.1 mM isobutylmethylxanthine or 5 μg/ml of cholera toxin had no effect on secretion at the low glucose concentration. These discrepancies between cAMP-promoting and insulin-releasing activities suggest that glucose does not initiate insulin release by activating the β-cell adenylate cyclase. By being metabolized in the β-cells, glucose may both create a release-initiating signal not identical with cAMP and enhance cAMP formation, leading to potentiation of the effect of the initiator signal.
Multivariate Approximation by Locally Blended Univariate Interpolants
Authors: Birkhoff Garrett , Cavendish James C. , Gordon William J.
Published: May 2006

A method is given for constructing simple new “finite elements” that seem well-suited to approximating smooth functions in rectangular polygons decomposed into rectangular cells. Some of the key properties of the elements are derived, and analogous three-dimensional “bricks” are constructed.
Different Blocks in the Differentiation of Myeloid Leukemic Cells
Authors: Lotem Joseph , Sachs Leo
Published: May 2006

Some clones of mouse myeloid leukemic cells (D
+
) can be induced to undergo cell differentiation to mature macrophages and granulocytes, and other clones (D
-
) could not be induced to differentiate to mature cells. Normal mature macrophages and granulocytes have surface receptors that form rosettes with erythrocytes coated with specific immunoglobulin or immunoglobulin-complement. The D
+
clones were induced to form receptors by prednisolone, cytosine-arabinoside, 5-iododeoxyuridine, actinomycin D, or serum from mice injected with endotoxin. All these compounds thus induced a common change in the cell surface membrane. The induction of receptors required protein synthesis, and receptors were formed before the appearance of mature cells. There were two types of D
-
clones. One type was induced by these compounds to form receptors, although with a lower inducibility than D
+
clones; in the other type there was no induction of receptors. The results indicate that there are different blocks in the differentiation of myeloid leukemic cells. Some leukemic cells (IR
+
D
+
) can be induced to form receptors and to differentiate to mature cells; others (IR
+
D
-
) can form receptors but not mature cells; and a third type (IR
-
D
-
) could not be induced to form receptors or mature cells.
Excision of Damaged Thymine Residues from Gamma-Irradiated Poly(dA-dT) by Crude Extracts of
Escherichia coli
Authors: Hariharan P. V. , Cerutti P. A.
Published: May 2006

Crude extracts of
E. coli
endo I
-
and
E. coli
endo I
-
uvrA6
-
possess the ability to remove thymine products of the 5,6-dihydroxy-dihydrothymine type from γ-irradiated or osmium tetroxide-oxidized poly(dA-dT). It is shown that the uvrA-gene product, which is responsible for incision close to photodimers in prereplication ultraviolet repair in
E. coli
, is not required for, but may aid in, the excision of γ-ray products of the 5,6-dihydroxy-dihydrothymine type. Ring-damaged thymine products are also removed by
E. coli
extracts from osmium tetroxide-oxidized poly(dA-dT), which contains only 5,6-dihydroxy-dihydrothymine but no strand breakage, indicating that product excision occurs in the absence of radiation-induced breaks. On the average, 8 to 16 nucleotides are removed from the polymer per ring-damaged thymine residue excised by extracts from both strains and for γ-irradiated and osmium tetroxide-oxidized polymer.
Use of the Avidin-Biotin Complex for Specific Staining of Biological Membranes in Electron Microscopy
Authors: Heitzmann H. , Richards Frederic M.
Published: May 2006

To expand the electron microscopist's options in localization and visualization, a new and general staining technique has been tested. The avidin-biotin complex serves as a coupling between the electron-dense marker, ferritin, and points of interest in biological samples. When specific cellular components are tagged with biotin, those components may be visualized with ferritin-linked avidin. Because of the remarkably strong affinity of avidin and biotin (characterized by an association constant of 10
15
M
-1
), the staining is rapid and stable.

The preparation of ferritin-avidin conjugate is described, and examples are presented of the application of this complex to biotin-tagged membranes. The ghosts of
Acholeplasma laidlawii
have been treated with biotinyl-
N
-hydroxysuccinimide ester to label protein amino groups. Erythrocyte membrane oligosaccharides have been oxidized by periodate or by galactose oxidase, and the resulting aldehydes labeled with biotin hydrazide.

The avidin-biotin complex in electron microscopy seems especially appropriate for seqential staining procedures, as well as for visualization of reaction sites of biotin-labeled, low-molecular-weight reagents.
Evidence for Cholinergic Synapses Between Dissociated Rat Sympathetic Neurons in Cell Culture
Authors: O'Lague P. H. , Obata K. , Claude P. , Furshpan E. J. , Potter D. D.
Published: May 2006

Sympathetic principal neurons were dissociated from superior cervical ganglia of new-born rats, and grown in cell culture. In electrophysiological experiments two types of excitatory synapses were found. One, which was relatively rare, was shown to operate by electrical transmission. The other, the predominant type, had several characteristics of chemical transmission, and pharmacological evidence indicated it was cholinergic.
Interaction of
Escherichia coli
30S Ribosomal Subunits with MS2 Phage RNA in the Absence of Initiation Factors
Authors: Szer W. , Leffler S.
Published: May 2006

MS2 RNA binds at 0° to 30S subunits from
E. coli
and, to a smaller extent, to those of a
Pseudomonas
species, as judged by filtration on nitrocellulose membranes; this mRNA does not bind to 30S subunits from
Bacillus brevis
or
Caulobacter crescentus
. Binding does not depend on the presence of initiation factors; it is sensitive to aurintricarboxylic acid but insensitive to edeine and is competitive with such synthetic polynucleotides as poly(U) and poly(AUG). Complex formation can also be detected by electrophoresis on polyacrylamide-agarose gels. By this procedure,
E. coli
30S subunits are separated into two major components. Only the more slowly moving component, which contains the ribosomal protein S1, interacts with the RNA.
Metamorphosis of
Aplysia californica
in Laboratory Culture
Authors: Kriegstein Arnold R. , Castellucci Vincent , Kandel Eric R.
Published: May 2006

To utilize the advantages offered by the large identified nerve cells of the marine mollusc
Aplysia californica
for cellular biological studies of development, we have devised simple techniques for growing this species in the laboratory in large number with a generation time as short as nineteen weeks. We have used the cultured animals to study the life cycle from fertilized egg to reproductive adult. The major developmental and behavioral changes occur at metamorphosis, when the larvae settle on the seaweed
Laurencia pacifica
and the locomotor and feeding behaviors are transformed into their adult forms. We have examined the timetable for the abandonment of larval behaviors and the emergence of adult ones and found that the transition from swimming to crawling occurs first and marks the onset of metamorphosis. The change from ciliary feeding to radular feeding occurs later and signals the end of metamorphosis. Other adult behaviors, such as the reflex responses and fixed-action patterns of the mantle organs, appear after metamorphosis.
Organization of Immunoglobulin Genes: Reiteration Frequency of the Mouse κ Chain Constant Region Gene
Authors: Honjo T. , Packman S. , Swan D. , Nau M. , Leder P.
Published: May 2006

Hybridization kinetic analyses with synthetic DNA indicate that there are only two to three copies of the κ constant region gene per haploid genome. This result lends weight to the argument that the immunoglobulin light chain is encoded by more than one continuous gene sequence.
Host Control of Endogenous Murine Leukemia Virus Gene Expression: Concentrations of Viral Proteins in High and Low Leukemia Mouse Strains
Authors: Strand Mette , Lilly Frank , August J. T.
Published: May 2006

Two of the major molecular components of murine leukemia virus particles, the internal protein (p30) and the envelope glycoprotein (gp69/71) have been measured in the spleens of normal, 6- to 10-week-old mice of various inbred strains and crosses. Both proteins were detected in virtually all mice. Extracts of high leukemia, high murine leukemia virus strains (AKR, C58, PL) showed high levels of both proteins; extracts of other strains usually showed lower levels. Of particular interest, however, were the exceptions to these general observations: (
1
) Very little gp69/71 could be detected in spleens of BALB mice, and this trait was dominant in crosses with AKR and DBA/2, both of which express a high level of gp69/71. Thymus-deficient BALB/c-
nu
/
nu
(nude) mice, in contrast, showed a higher concentration of gp69/71 typical of other low leukemia strains, suggesting that the virtual absence of the protein in normal BALB/c mice may result from immunologic suppression. (
2
) With C57L, C57BL/10Sn, and C57BL/6 strains the concentration of p30 was lower, in some cases much lower, than would be expected from the concentration of gp69/71. (
3
) DBA/2 mice showed high levels of gp69/71, 10-fold greater than that of p30, whereas congenic DBA/2-

Fv-1
b

mice showed the opposite pattern. (
4
) Mice of 129-G
IX
-
strain showed no detectable levels of either p30 or gp69/71 proteins, although the congenic 129 (G
IX
+
) showed appreciable levels of both. The absence of these proteins in 129-G
IX
-
mice is a recessive trait, as F
1
hybrids with AKR and DBA/2 showed appropriate levels of both proteins. It is concluded that expression of viral p30 and gp69/71 proteins in mice is not coordinately linked and that expression is complex, being influenced by several genes, including
Gv-1, H-2, Fv-1
, and probably still others.
13
C Nuclear Magnetic Resonance Spectroscopic Evidence for Hydrophobic Lipid-Protein Interactions in Human High Density Lipoproteins
Authors: Stoffel Wilhelm , Zierenberg Ottfried , Tunggal Budi , Schreiber Ekkehard
Published: May 2006

Phosphatidylcholines, sphingomyelins, cholesterol, and cholesterol esters were enriched with
13
C by chemical synthesis in specific positions of their hydrophilic groups and aliphatic chains. Their spin-lattice relaxation times were determined in organic solvents. The substances were organized as liposomes and recombined with total human high density apolipoproteins and the two separated main components, apolipoprotein A-I (apoLp-Gln-I) and apolipoprotein A-II (apoLp-Gln-II).

These
13
C nuclear magnetic resonance data established that in reassembled high density lipoproteins the phospholipid molecules bind to the apoprotein moieties with their hydrophobic fatty acid chains and not with their hydrophilic zwitterionic groups. Apolipoprotein A-I preferentially binds phosphatidylcholine, although its lipid-binding capacity is smaller than that of apolipoprotein A-II. Apolipoprotein A-II avidly reassembles with sphingomyelin by hydrophobic interactions. A model of the molecular organization of the high density lipoportein particle has been derived.
13
C Nuclear Magnetic Resonance Spectroscopy of Native and Recombined Lipoproteins
Authors: Assmann Gerd , Highet Robert J. , Sokoloski Edward A. , Brewer H. Bryan
Published: May 2006

13
C nuclear magnetic resonance data on native and recombined lipoproteins are reported. [
Methyl
-
13
C]phosphatidylcholine, [
methyl
-
13
C]sphingomyelin, 1,2-[
dioleoyl-1
-
13
C]-
sn
-phosphatidylcholine and cholesteryl-[
1
-
13
C]oleate were enriched with 90% carbon-13 in respective molecules by chemical synthesis and used for recombination experiments with high density lipoprotein apoproteins. Relaxation times for these specifically enriched lipids in organic solvents,
2
H
2
O, and lipid-protein complexes isolated by ultracentrifugal flotation, were measured. The results show that both the phosphatidylcholine and sphingomyelin polar headgroups have the same hydrophilic environment in either sonicated lipid particles or reassembled lipoproteins, and suggest that ionic interaction of lipid and apolipoproteins is of minor importance in the formation of plasma lipoprotein complexes. Our experiments indicate that
13
C nuclear magnetic resonance spectroscopy will contribute to the understanding of lipidprotein interaction in lipoproteins and membranes.
Type C Viral Expression in Primate Tissues
Authors: Sherr Charles J. , Benveniste Raoul E. , Todaro George J.
Published: May 2006

Nucleic acid sequences homologous to a [
3
H]DNA transcript prepared from an endogenous baboon type C virus are readily detected in both the cellular DNA and RNA of several different Old World monkeys. Competitive radioimmunoassays for the major viral protein (p30) of baboon type C viruses detect viral antigen in a normal stumptail spleen and a rhesus ovarian carcinoma. The p30 antigen from these tissues is closely related by several immunologic criteria to the p30 protein of baboon type C viruses. The results indicate that normal primate tissues transcribe endogenous viral-specific RNA and translate at least one viral structural protein.
Analysis of the C-Value Paradox by Molecular Hybridization
Authors: Rosbash Michael , Ford Peter J. , Bishop John O.
Published: May 2006

Poly(A)-containing RNA was isolated from ovaries of
Xenopus laevis laevis
and
Triturus cristatus carnifex
and used as a template for the synthesis of radioactive complementary DNA with RNA-dependent DNA polymerase. When annealed with an excess of homologous DNA, the complementary DNA is rendered double-stranded with kinetics that suggest that the coding sequences are single-copy in both these organisms. In
Triturus
, these sequences are distinct from the majority of the genome, which consists of repeated sequences, and distinct from the ribosomal cistrons, which are present in proportion to the increase in C-value relative to the
Xenopus
genome. Moreover, the number of different poly(A)-containing molecules in the ovary (sequence complexity) is the same in
Xenopus
and in
Triturus
.
Interferon action II. Membrane-bound alkaline ribonuclease activity in chick embryo cells manifesting interferon-mediated interference.
Authors: Marcus P I , Terry T M , Levine S
Published: May 2006

Membrane fractions from chick embryo cells manifesting viral interference mediated by interferon or poly(I)-poly(C) contain high levels of an alkaline ribonuclease. Enhanced RNase activity is not observed when inhibitors of cell protein or RNA synthesis are present during interferon treatment, or when heterologous interferon is used. The RNase associated with comparable membrane fractions from cells treated with mock-interferon is about 1/10 as active, and shows qualitative differences. In principle, divergent views of interferon action may be reconciled to a common mode of action by postulating that viral interference results from a newly induced or activated RNase of cellular origin and proper specificity that acts to reduce the accumulation and functional capacity of newly synthesized viral RNAs, particularly mRNA. Previous data in support of interferon's acting to inhibit virion-derived transcription in vivo are now interpreted as demonstrating enhanced degradation of viral transcripts (mRNA).
Tay-Sachs' and Sandhoff's diseases: the assignment of genes for hexosaminidase A and B to individual human chromosomes.
Authors: Gilbert F , Kucherlapati R , Creagan R P , Murnane M J , Darlington G J , Ruddle F H
Published: May 2006

The techniques of somatic cell genetics have been used to establish the linkage relationships of loci coding for two forms (A and B) of hexosaminidase (EC 3.2.1.30; 2-acetamido-2-deoxy-beta-D-glucoside acetamidodeoxyglucohydrolase) and to determine whether a structural relationship exists between these forms. In a series of human-mouse hybrid cell lines, hexosaminidase A and B segregated independently. Our results and those reported by other investigators are used to analyze the proposed structural models for hexosaminidase. We have also been able to establish a syntenic relationship between the gene locus responsible for the expression of hexosaminidase A and those responsible for mannosephosphate isomerase and pyruvate kinase-3 and to assign the gene for hexosaminidase B to chromosome 5 in man. There is thus a linkage between specific human autosomes and enzymes implicated in the production of lipid storage diseases.
Short-lived messenger RNA in HeLa cells and its impace on the kinetics of accumulation of cytoplasmic polyadenylate.
Authors: Puckett L , Chambers S , Darnell J E
Published: May 2006

Accumulation of [3H]adenine in the acid-soluble pool and in nuclear and cytoplasmic poly(A) of HeLa cells shows that the nuclear poly(A) rises along a curve similar to that of the acid-soluble pool. By use of a [3H]guanosine pulse-chase experiment in adenine-grown cells, at least 35-50% of the pulse-labeled mRNA was found to have a half-life of about 1-2 hr. A mathematical model involving nuclear poly(A) synthesis and conservative transport to the cytoplasm has been derived from the new information about mRNA with a short half-life. This model predicts curves similar to those found for nuclear and cytoplasmic accumulation of poly(A). Thus, there is no necessity on kinetic grounds to invoke either nuclear turnover or cytoplasmic synthesis of poly(A).
Cobalt(III), a probe of metal binding sites of Escherichia coli alkaline phosphatase.
Authors: Anderson R A , Vallee B L
Published: May 2006

To facilitate the study of individual metal binding sites of polymeric metalloproteins, conversion of exchange-labile Co(II) in E. coli alkaline phosphatase (EC 3.1.3.1) to exchange-inert Co(III) was examined. Oxidation of Co(II) alkaline phosphatase with hydrogen peroxide results in a single absorption maximum at 530 nm and loss both of the characteristic electron paramagnetic signal and of enzymatic activity. Zinc neither reactivates this enzyme nor displaces the oxidized cobalt atoms. Metal and amino-acid analyses demonstrate that oxidation alters neither cobalt binding nor amino-acid composition of the enzyme. Al data are consistent with the conclusion that hydrogen peroxide oxidizes Co(II) in alkaline phosphatase to Co(III). Polymeric metalloenzymes can contain different categories of metal atoms serving in catalysis, structure stabilization, and/or control and exerting their effects independently or interdependently. The in situ conversion of exchange-labile Co(II) to exchange-stable (Co(III) offers a method to selectively and differentially "freeze" cobalt atoms at their respective binding sites. The accompanying spectral changes and concomitant retardation in ligand exchange reactions may be used to differentiate between specific metal binding sites that serve different roles in polymeric metalloenzymes.
Mobility of cholera toxin receptors on rat lymphocyte membranes.
Authors: Craig S W , Cuatrecasas P
Published: May 2006

Fluorescein-labeled cholera toxin binds detectably to 40-60% of rat mesenteric lymph node cells and induces a temperature-dependent redistribution (patch and cap formation) of cell surface toxin receptors. The redistribution is inhibited by several "metabolic," "microtubule," and "microfilament" inhibitors, by concanavalin A, and by anticholera toxin IgG. Various studies indicate that cholera toxin is at least bivalent, and that this property may be related to both the induction of receptor redistribution and to the activation of adenylate cyclase. Membrane components which are probably identical to the sialo-glycolipid, GM1 ganglioside, appear to be mobile in the plane of the membrane. The possible role of toxin multivalency and receptor mobility in the mechanism of toxin action is considered.
phiX174 DNA-dependent DNA synthesis in vitro: requirement for P1 ban protein in dnaB mutant extracts of Escherichia coli.
Authors: Schuster H , Mikolajczyk M , Rohrschneider J , Geschke B
Published: May 2006

Ammonium sulfate fractionation of crude extracts of E. coli yields a soluble enzyme fraction (about 25-fold purification) that catalyzes the conversion of phiX174 single-stranded DNA to duplex DNA. The reaction is rifampicin-resistant, requires single-stranded DNA, Mg++, deoxynucleoside triphosphates, and ATP, and is stimulated by KCl. Such soluble enzyme fractions were prepared from E. coli strains carrying the prophage mutant P1bac, in which the viral dnaB analog (ban) protein is expressed constitutively, or P1bacban, in which the expression of ban protein is prevented. DNA-synthesizing activity of ban protein containing fractions from wild-type or dnaB(P1bac) lysogens was more temperature-resistant than that from E. coli containing only wild-type dnaB protein, whereas that from dnaB(P1bacban) lysogens of dnaB cells was extremely thermolabile. It is suggested that the temperature-resistant DNA synthesis with fractions from P1bac lysogens is mediated by the P1 ban protein.
Translation of hormone-induced messenger RNA in amphibian oocytes: I. Induction by estrogen of messenger RNA encoded for vitellogenic protein in the liver of the male African clawed toad (Xenopus laevis).
Authors: Lanclos K D , Hamilton T H
Published: May 2006

Induction of the synthesis of the vitellogenic proteins, lipovitellin and phosvitin, in the liver of the male African clawed toad (Xenopus laevis) was investigated as a function of time after treatment with estradiol-17beta [1,3,5(10)-estratriene-3,17beta-diol]. The appearance of mRNAs encoded for lipovitellin and phosvitin in the cytoplasmic fraction of the liver was assayed by microinjections of hepatic mRNA preparation [either polyribosomes or poly(A)-rich RNA] into oocytes obtained from mature female toads. Oocytes were then incubated in the presence of radioactive amino acid(s) at 19 degrees for periods of time varying from 4 to 18 hr after microinjection. The results show that at 2 hr after hormone treatment more mRNA was present in the cytoplasm, and that from 2 to 72 hr after treatment the level of induced mRNA increased almost linearly to 110% above the control values. Experiments employing specific lipovitellin antiserum indicated no radioactive lipovitellin among the proteins synthesized in oocytes microinjected with hepatic mRNAs isolated from 3 to 9 hr after hormone treatment. However, a marked synthesis of immunoprecipitable, radioactive lipovitellin and an enhanced incorporation of [3H]serine occurred in the oocytes microinjected with hepatic mRNA preparations obtained from toads treated with hormone for 12 or more hr. The identities of the proteins encoded by the mRNAs induced early in estrogen action (2-9 hr) in the male amphibian liver are unknown. It is surmised that some of these proteins may function in the regulation of the subsequent synthesis of the vitellogenic proteins.
Evidence for the translocation of 5'-nucleotidase across hepatic membranes in vivo.
Authors: Little J S , Widnell C C
Published: May 2006

Hepatic 5'-nucleotidase (EC 3.1.3.5; 5'-ribonucleotide phosphohydrolase) activity has been studied in cisternal elements of the Golgi complex and in secretion vacuoles, both isolated after ethanol administration to rats in vivo. The enzyme in secretion vacuoles was latent, so that a 5-fold increase in activity was observed when incubations were carried out in the presence of detergent; evidence is presented that the latency is caused by the impermeability of the membrane to substrate. Essentially no latency was observed in Golgi cisternae. Confirming the results of Farquhar et al. [(1974) J. Cell Biol. 60, 8-25], reaction product from 5'-nucleotidase was localized by cytochemical procedures on the inside of secretion vacuoles and on the cytoplasmic side of Golgi cisternae. After solubilization in detergent, the enzyme from both fractions reacted almost identically with both antibody to the purified enzyme and concanavalin A. In contrast, when intact fractions were incubated with an excess of antibody or concanavalin A, only 22-23% of the enzyme was inhibited in secretion vacuoles whereas 51-84% was inhibited in Golgi cisternae. Sonication of secretion vacuoles in the presence of antibody or concanavalin A increased the inhibition 2- to 3-fold. It is suggested that during the formation of secretion vacuoles from the Golgi cisternae, 5'-nucleotidase is translocated from the cytoplasmic side of the membrane to the inside.
Specific endocrine and hormonal receptor changes associated with liver regeneration in adult rats.
Authors: Leffert H , Alexander N M , Faloona G , Rubalcava B , Unger R
Published: May 2006

Partial removal of the adult rat liver--a procedure that stimulates liver regeneration--causes arterial levels of insulin and thyroxin to fall and glucagon levels to rise. These changes are correlated with the quantity of liver tissue excised and not solely with nonspecific operative manipulations or with blood glucose levels. During regeneration (about 24 hr postoperatively), hepatic plasma membranes become partially resistant to binding glucagon; this change is specific because it is not detected in laparotomized controls and insulin binding is not significantly altered. It remains to be proven whether any one or more of these specific endocrine changes are casually related to the initiation of hepatic proliferation.
Hypoxanthine phosphoribosyltransferase deficiency: association of reduced catalytic activity with reduced levels of immunologically detectable enzyme protein.
Authors: Upchurch K S , Leyva A , Arnold W J , Holmes E W , Kelley W N
Published: May 2006

In the present study hemolysates from fourteen patients with a genetically determined deficiency of hypoxanthine phosphoribosyltransferase (EC 2.4.2.8; IMP:pyrophosphate phosphoribosyltransferase) activity were examined immunologically for the presence of material that crossreacts with the normal enzyme. A quantitative assay for crossreacting material in enzyme-deficient hemolysates was based on the inhibition of the immunoprecipitation of the normal enzyme. As little as 3% of normal crossreacting material could be detected. One patient in this series was found to have a normal amount of crossreacting material, whereas the remainder had no detectable crossreacting protein. The lack of detectable crossreacting material in these patients raises the possibility that a defect in synthesis or degradation of enzyme protein may be present in many patients deficient in hypoxanthine phosphoribosyltransferase.
Radioimmune determination of hypoxanthine phosphoribosyltransferase crossreacting material in erythrocytes of Lesch-Nyhan patients.
Authors: Ghangas G S , Milman G
Published: May 2006

We have developed a sensitive radioimmunoassay capable of detecting and quantitating 20 ng of hypoxanthine phosphoribosyltransferase (EC 2.4.2.8; IMP:pyrophosphate phosphoribosyltransferase) protein. For this assay, hypoxanthine phosphoribosyltransferase from human erythrocytes was iodinated with 125I under mild conditions using hydrogen peroxide and lactoperoxidase attached to Sepharose-4B. Antisera prepared against homogeneous human hypoxanthine phosphoribosyltransferase precipitates the iodinated enzyme as effectively as the unlabeled enzyme. The radioimmunoassay has been used to look for hypoxanthine phosphoribosyltransferase crossreacting material in hemolysates from sixteen different patients with a marked genetic deficiency of this enzyme characteristic of the Lesch-Nyhan syndrome. Fifteen hemolysates contained no detectable (less than 1% of normal) crossreacting material. One hemolysate contained a normal amount of crossreacting material. Hypoxanthine phosphoribosyltransferase from this patient (E.S.) has been shown to be a Km mutant enzyme.
Bis-(N-maleimidomethyl) ether: an antisickling reagent.
Authors: Zak S J , Geller G R , Finkel B , Tukey D P , McCormack M K , Krivit W
Published: May 2006

The interaction of bis-(N-maleimidomethyl) ether with oxyhemoglobin results in covalent linkages of both maleimide groups, converting them to succinyl derivatives of beta93 Cys and beta97 His at their sulfhydryl and imidazolyl side chains, respectively. The resultant hemoglobin is stable, and reveals a left-shifted oxyhemoglobin equilibrium curve in which cooperativity is abolished. This reagent readily traverses the red cell membrane and prevents the sickling reaction upon deoxygenation. It appears to affect none of the activities of the red cell enzymes adversely, nor does it appear to affect the red cell membrane. Since there are several defined effects on the stereochemical status of the molecule conferred by interaction with bis-(N-maleimidomethyl) ether, the precise mechanism of the anitsickling effect remains to be elucidated. A more subtle perturberant will be required to specify a precise antisickling effect. By use of bis-(N-maleimidomethyl) ether a precise locus on the beta chain of human hemoglobin S can be perturbed to produce the desired effect.
Complete translation of poliovirus RNA in a eukaryotic cell-free system.
Authors: Villa-Komaroff L , Guttman N , Baltimore D , Lodishi H F
Published: May 2006

Poliovirus RNA stimulates imcorporation of 35S from both [35S]methionine and formyl-[35S]methionyl-tRNAfMet in cell-free systems derived from HeLa cells or from poliovirus-infected HeLa cells. The largest product formed under the direction of the viral RNA is the same size as the polyprotein thought to represent translation of the entire RNA. Synthesis of this polyprotein and other large products was stimulated greatly by increasing the salt concentration during the reaction from the optimum for initiation (90 mM) to the optimum for elongation (155 mM). Only one initiation peptide could be identified, and a tryptic digest of the product contained mainly peptides that cochromatographed with peptides from authentic viral proteins. The RNA from a deletion mutant of poliovirus initiated protein synthesis at the same site used by standard RNA and programmed synthesis of an appropriately deleted set of polypeptides. The results strongly support the model of translation of poliovirus RNA from a single initiation site into a continuous polyprotein that is cleaved to form the functional proteins. It is suggested that uninfected HeLa cell extracts can carry out the cleavages of nascent polyprotein.
Temporal changes in tectal cell surface specificity induced by nerve growth factor.
Authors: Merrell R , Pulliam M W , Randono L , Boyd L F , Bradshaw R A , Glaser L
Published: May 2006

The change in cell surface adhesive specificity previously shown to occur between day 7 and 8 of development in the chick optic tectum ]Gottlieb et al. (1974), Proc. Nat. Acad. Sci. USA 71, 1800-1802] can be induced in rotating cultures of tectal cells by the addition of 10(-7) M mouse submaxillary gland nerve growth factor. Insulin, proinsulin, dexamethasone, and performic-acid-oxidized nerve growth factor are individually inactive in this system, but nerve growth factor in which the three tryptophan residues/subunit have been oxidized with N-bromosuccinimide is active. Thus, the specificity of this system for nerve growth factor is different than that observed with embryonic dorsal root or sympathetic ganglia, where the oxidized tryptophan derivative is inactive in stimulating neurite production. It is possible that in this system nerve growth factor serves as an analog of another specific trophic factor, presumably structurally related to nerve growth factor, may be active at much lower concentrations.
Conformational fluctuations of DNA helix.
Authors: Depew D E , Wang J C
Published: May 2006

Treatment of a homogeneous sample of circular DNA molecules that contain at least one single-strand scission per molecule with ligase yields a population of covalently closed circles heterogeneous in the topological winding number alpha. Under proper conditions, the product molecules with different values of alpha can all be resolved by gel electrophoresis. The distributions in alpha for several DNAs, covalently closed at different temperatures, have been determined and the results have been interpreted in terms of thermal fluctuations of the DNA helix.
Functional level of rat liver tryptophan 2,3-dixoygenase messenger RNA during superinduction of enzyme with actinomycin D.
Authors: Killewich L , Schutz G , Feigelson P
Published: May 2006

Tryptophan 2,3-dioxygenase [EC 1.13.11.11; L-tryptophan:oxygen 2,3-oxidoreductase (decyclizing)] activity is induced by glucocorticoid hormones and superinduced by actinomycin D. Previous experiments had shown that hormonal induction of the enzyme activity is accompanied by parallel increases in tryptophan 2,3-dioxygenase mRNA level. In this study, we measured the tryptophan 2,3-dioxygenase mRNA levels during superinduction as well as hormonal induction, to determine whether superinduction of the enzyme activity is also mediated through changes in mRNA concentration. Tryptophan 2,3-dioxygenase mRNA was measured in a Krebs ascites cell-free protein synthesizing system supplemented with rabbit reticulocyte initiation factors. We found that during superinduction of the enzyme activity by actinomycin D, the mRNA level is identical to that of the actinomycin D-free controls. Our results do not, therefore, support the hypothesis that hormonal induction and/or superinduction of tryptophan 2,3-dioxygenase mRNA are regulated by a rapidly turning over repressor.
Androgen- and estrogen-binding macromolecules in developing mouse brain: biochemical and genetic evidence.
Authors: Fox T O
Published: May 2006

Androgen- and estrogen-binding macromolecules from the hypothalamus plus preoptic area of 3- to 4-week-old mice have been detected and partially characterized. These components bind the respective hormones with high affinity (saturating at 4-8 nM) and sediment with rates typical of presumed steroid receptors (4.0-4.5 S in 0.15 M NaCl, 5.0-7.5 S without salt). A 90-95% reduction in androgen binding found in the androgen-insensitivity mutant mouse, testicular feminization (Tfm), provides a genetic control for the specificity of binding. This reduced androgen binding with Tfm/Y mutants and blocking experiments with non-radioactive estradiol [estra-1,3,5(10)-triene-3,17beta-diol] and testosterone (17beta-hydroxy-4-androsten-3-one) indicate the existence of at least two binding components: one with high affinity only for estradiol, the other with affinity for both androgens and estrogen. Based on these properties, a receptor mechanism that detects relative concentrations of androgens and estrogens is proposed.
Identification of a region in 23S rRNA located at the peptidyl transferase center.
Authors: Sonenberg N , Wilchek M , Zamir A
Published: May 2006

A photolyzable derivative of dipeptidyl-tRNA, p-azido-N-tBoc-Phe-[3H]Phe-tRNA, bound reversibly to 70S ribosomes in the presence of poly(U), becomes, when irradiated, covalently attached to components of the 50S ribosomal subunit. Most of the reaction occurs within the 23S RNA, while ribosomal proteins are only weakly labeled. Reversible binding as well as the covalent reaction are reduced in the absence of poly(U) or in the presence of several antibiotics specific for the 50S ribosomal subunit. There are apparently few sites (or perhaps a single site) of reaction on the 23S RNA that are exclusively located within the 2000 nucleotides from the 3 terminus of the molecule. Part of the 23S RNA within this region must therefore be closely associated with the peptidyl transferase center of the ribosome.
Evidence of homologous relationship between thermolysin and neutral protease A of Bacillus subtilis.
Authors: Levy P L , Pangburn M K , Burstein Y , Ericsson L H , Neurath H , Walsh K A
Published: May 2006

A comparison of the partial amino-acid sequence of neutral protease A from Bacillus subtilis with the structure of thermolysin (EC 3.4.24.4) from Bacillus thermoproteolyticus reveals that these two proteins are homologous. Of 171 residues placed in neutral protease (54% of the sequence), 83 residues (49%) occur in identical positions in thermolysin, and include nine of the 13 residues previously identified as components of the active site of thermolysin. This similarity provides support for the hypothesis that the two enzymes have similar three-dimensional structures and a common mechanism of action. Since these enzymes differ markedly in their resistance to heat inactivation, a comparison of their structures may eventually provide a chemical basis for explaining the differences in their thermal stability.
Thermolabile T (tumor) antigen from cells transformed by a temperature-sensitive mutant of simian virus 40.
Authors: Tenen D G , Baygell P , Livingston D M
Published: May 2006

Partially purified tumor (T) antigen from a strain of Chinese hamster lung cells transformed by wild-type simian virus 40 (SV 40) and either of two temperature-sensitive SV 40 mutants has been studied as a DNA binding protein. The DNA binding activity present in the T-antigen-containing fractions is inhibited by purified hamster anti-T IgG but not by equivalent amounts of nonimmune hamster IgG. T from either wild-type- or tsC219-transformed cells is relatively stable during heating at 44 degrees compared to T prepared from tsA239-transformed cells. These results strongly suggest that T is a product of the SV 40 A gene.
Adenovirus-2 mRNA is transcribed as part of a high-molecular-weight precursor RNA.
Authors: Bachenheimer S , Darnell J E
Published: May 2006

The order of transcription and the length of nascent RNA transcripts from adenovirus-2 (Ad-2) DNA in the nucleus of infected cells has been deduced by labeling the growing RNA chains in vivo for a very brief period, separating the RNA on the basis of size, and hybridizine to the ordered EcoRI restriction endonuclease fragments derived from Ad-2 DNA. The majority of the virus-specific RNA molecules are synthesized as very high-molecular-weight units beginning at a common point at least 25-30,000 base pairs from one end of the Ad-2 DNA. These molecules can be reduced in size without further RNA synthesis. The experiments indicate the obligatory origin of Ad-2 mRNA from a high-molecular-weight precursor molecule.
Induction of an immune response through interaction of helper cells with an immune complex bound to the surface of B cells.
Authors: Romano T J , Lerman S P , Bangasser S , Thorbecke G J , Nisonoff A
Published: May 2006

Adoptive anti-trinitrophenyl (Tnp) responses were elicited from Tnp-hemocyanin (Tnp-KLH)-primed cells by challenge with an immune complex of KLH and a Tnp conjugate of the Fab fragment of rabbit anti-KLH. Removal of T cells by treatment with anti-Thy 1.2 (phi C3H) + complement abolished this effect. Tnp-KLH primed cells that had been incubated with a Tnp conjugate of pneumococcal polysaccharide type III (Tnp-SIII), and which were then unable to respond to Tnp-KLH, made an anti-Tnp response upon incubation with rabbit anti-Dnp when transferred together with cells primed with rabbit gamma globulin. Since Tnp-KLH was not added in these experiments, it would appear that the cells were triggered by immune complexes on their surfaces consisting of Tnp-SIII and rabbit anti-Dnp with the help of T cells primed with rabbit gamma globulin; the presence of free antigen was evidently not required. Therefore, antigen-antibody complexes, when bound to the surface of B cells, may mediate T-B cell cooperation in the humoral immune response.
Guinea pig transfer factor-like activity detected in vitro.
Authors: Dunnick W , Bach F H
Published: May 2006

"Transfer factor" was prepared by Sephadex G-25 chromatography of lymph node cell lysates from guinea pigs immunized with ovalbumin or bovine gamma globulin. Treatment of nonimmune peritoneal exudate cells with the transfer factor and specific antigen leads to inhibition of migration of the cells, whereas cells treated with the transfer factor alone or specific antigen alone are not inhibited from migrating. An average of 24-28% inhibition is observed in the presence of transfer factor and specific antigen, but only 5-15% inhibition in the presence of transfer factor and nonspecific antigen. The guinea pig transfer factor we have tested in vitro has some physical characteristics in common with human transfer factor.
Role of methionine in bacterial chemotaxis: requirement for tumbling and involvement in information processing.
Authors: Springer M S , Kort E N , Larsen S H , Ordal G W , Reader R W , Adler J
Published: May 2006

Chemotactic responses are mediated by modulation of the frequency of tumbling. Studies with methionine auxotrophs of wild-type Escherichia coli and four mutants which tumble continuously show that methionine or one of its metabolites is involved in the tumbling process. Following removal of methionine, the wild type and two mutants, after various periods of time, became unable to tumble. The presence of constant levels of chemical attractants considerably shortened these periods in the three strains and eliminated tumbling in another mutant. This effect of attractants considerably shortened these periods in the three strains and eliminated tumbling in another mutant. This effect of attractants implies that methionine or some derivative of methionine is also involved in transducing chemical stimuli to bacterial responses.
Separation of sarcoma virus-specific and leukemia virus-specific genetic sequences of Moloney sarcoma virus.
Authors: Scolnick E M , Howk R S , Anisowicz A , Peebles P T , Scher C D , Parks W P
Published: May 2006

We have studied the nucleic acid sequences in nonproducer cells transformed by Moloney sarcoma virus or Abelson leukemia virus (two types of replication-defective, RNA-containing, viruses isolated by passage of Moloney leukemia virus in BALB/c mice). DNA probes from the Moloney leukemia in virus detect RNA in both Abelson virus-transformed nonproducer cells and Moloney sarcoma virus-transformed nonproducer cells. A sarcoma-specific cDNA, prepared from the Moloney sarcoma virus, has extensive homology to RNA found in heterologous nonproducer cells transformed by Moloney sarcoma virus, has little homology to RNA in cells producing Moloney leukemia virus, and no detectable homology to RNA in nonproducer cells transformed by the Abelson virus. By analogy to earlier data on avian and mammalian sarcoma viruses, these results suggest that the Moloney sarcoma virus arose by recombination between a portion of the Moloney leukemia virus genome and additional sarcoma-specific information, and indicate that the expression of this information in not essential for Abelson virus-mediated fibroblast transformation.
Studies on memory: inhibitors of protein synthesis also inhibit catecholamine synthesis.
Authors: Flexner L B , Goodman R H
Published: May 2006

The rates of accumulation of newly synthesized catecholamines and endogenous catecholamine levels in mice were determined after treatment with cycloheximide, acetoxycycloheximde, puromycin, and anisomycin. The rates of accumulation were found to be decreased by all antibiotics tested, weakening the assumption that their amnestic effects are due solely to inhibition of protein synthesis.
Promoter-dependent transcription of tRNAITyr genes using DNA fragments produced by restriction enzymes.
Authors: Küpper H , Contreras R , Landy A , Khorana H G
Published: May 2006

Two DNA fragments prepared from the transducing bacteriophage strains ø80psuIII+ and ø80hpsuIII+,- by digestion with restriction enzymes contain one tyrosine tRNA gene (suIII+) and two tyrosine tRNA genes (suIII+, su-) in tandem, respectively, a single promoter in both cases, and some additional DNA regions at the two ends of both. Using these fragments, we have studied characteristics of the promoter-dependent transcription of the tyrosine tRNA genes. The promoter-dependent transcripts were shown to correspond to the expected tRNA precursors. Exposure of the transcript from the single gene fragment to an S100 extract from Escherichia coli gave, via intermediates, 4S material which was active in enzymatically accepting tyrosine and contained some modified bases.
Mapping of biological functions on RNA of avian tumor viruses: location of regions required for transformation and determination of host range.
Authors: Joho R H , Billeter M A , Weissmann C
Published: May 2006

A map of the large T1 oligonucleotides of the RNA of Prague Rous sarcoma virus, strain B (Pr RSVb) has recently been established (Coffin and Billeter, submitted for publication). Since the RNA of Rous associated virus, type 1 (RAV-1) lacks many of the large 1 oligonucleotides of Pr RSV-B and contains others not present in the latter, the RNA of recombinants between RAV-1 and Pr RSV-B could be analyzed with regard to the origin of its sequences. Recombinants were selected for transforming capacity (characteristic for Pr RSV-B) and ability to grow on C/B chicken fibroblasts (characteristic for RAV-1). Four out of five recombinants examined had undergone at least two crossovers. The set of Pr RSV-B-specific oligonucleotides present in all recombinants defined an RNA region near the poly(A) segment; this must contain genetic information required for transformation required for transformation (the onc function). All recombinants lost a set of contiguous Pr RSV-B-specific oligonucleotides and concomitantly acquired a set of RAV-1-specific oligonucleotides. These define a region in the middle section of the oligonucleotide map, all or some of which must be required for determining growth capacity on C/B cells (the env function).
Membrane-microtubule interactions: concanavalin A capping induced redistribution of cytoplasmic microtubules and colchicine binding proteins.
Authors: Albertini D F , Clark J I
Published: May 2006

The relationship between microtubules and concanavalin A surface receptors during concanavalin A capping in primary cultures of rabbit ovarian granulosa cells was examined by electron microscopic and fluorescence labeling techniques. Cells treated with concanavalin A and hemocyanin at 4 degree or 37 degree and then incubated at 37 degree for 1 hr formed large juxtanuclear caps that were observed with shadow cast replicas of the cell surface. Thin section analysis of capped cells revealed an abundance of microtubules immediately beneath the cap which were arranged approximately perpendicular to the plane of the membrane. The capping process was unaffected by the antimicrotubule agents colchicine or vinblastine. Further, vinblastine treatment of capped calls resulted in the formation of numerous paracrystals that were confined to the cytoplasm underlying the capped region of the membrane; uncapped cells displayed paracrystals that were randomly dispersed in the cytoplasm. Exposure of fixed cells to fluorescein thiocarbamyl colchicine, which localizes colchicine binding proteins, revealed an intensely fluorescent region that corresponded to the cap; this staining pattern was absent in uncapped cells. These findings indicate that concanavalin A mediated capping modifies the cytoplasmic disposition of microtubules and colchicine binding proteins. Further, it is suggested that the capped region of the plasma membrane is a preferred site of microtubule polymerization.