bluuebunny/crossref_metadata_2025 huggingface dataset:
Seesaw signal processing in pineal cells: homologous sensitization of adrenergic stimulation of cyclic GMP accompanies homologous desensitization of beta-adrenergic stimulation of cyclic AMP.
Authors: Klein D C , Auerbach D A , Weller J L
Published: May 2006

Studies of the adrenergic regulation of cyclic GMP in the pineal gland show that (-)-norepinephrine stimulates cyclic GMP primarily in pineal cells, rather than in nerve endings as previously thought. The response exhibits the interesting and unusual characteristic of homologous sensitization: It is maintained by neural stimulation and disappears gradually as a consequence of depressed neural stimulation, due to denervation or decentralization of the superior cervical ganglia or to constant light. The response is restored in intact animals that had been in a constant-light environment when they are returned to a normal light cycle and in ganglionectomized animals by norepinephrine treatment. These findings are especially interesting because the pineal adrenergic--cyclic AMP stimulus--response system exhibits homologous desensitization. The occurrence of homologous sensitization of a cyclic GMP response and desensitization of a cyclic AMP response, which we term seesaw signal processing, in the same tissue or cell has intriguing implications. It provides a mechanism through which the qualitative nature of a multicomponent response can be modified. Such a mechanism could play a role in signal processing by neural or neuroendocrine tissues that release two or more extracellular messages.
Regulation of DNA replication: "target" determinant of the replication control elements of plasmid R6-5 lies within a control element gene.
Authors: Danbara H , Brady G , Timmis J K , Timmis K N
Published: May 2006

The replication control system of plasmid R6-5 has been investigated by characterization of high-copy-number mutant miniplasmids, development of an in vivo assay for the site of action or "target" of the replication control elements, and sequence analysis of the replication control regions of the wild-type plasmid and two copy-number mutant derivatives. These and other experiments have shown that three plasmid determinants--copA/incA, copB, and copT--are involved in DNA replication control. The products of the copB and copA/incA genes, a 9500-dalton basic polypeptide and either a 7200-dalton basic polypeptide or a short untranslated RNA molecule, respectively, are negative-acting elements that interact with the third element, their target, the copT DNA sequence, or its product to regulate the frequency of initiation of plasmid replication. The location of copT within the copA/incA gene and 1600 base pairs upstream from the origin of replication indicates that regulation is effected at a preinitiation stage of replication, such as the production of a primer or other initiation factor.
Regulation of acetylcholine receptor phosphorylation by calcium and calmodulin.
Authors: Smilowitz H , Hadjian R A , Dwyer J , Feinstein M B
Published: May 2006

Acetylcholine receptor-enriched membranes prepared from frozen electric organ of Torpedo californica by differential centrifugation and density step gradient centrifugation were assayed for endogenous phosphorylation in the absence and presence of calmodulin and calcium. Each of the membrane fractions exhibited a 3- to 6-fold stimulation of endogenous phosphorylation by calcium and calmodulin. Both calcium and calmodulin were needed for maximal stimulation although calcium alone afforded a small, reproducible stimulation of endogenous phosphorylation. In the presence of fluoride, a phosphatase inhibitor, the calmodulin plus calcium stimulation was increased an additional 3-fold. The phosphorylation reaction was rapid, and maximal phosphorylation was achieved in 2 min. Stimulation of phosphorylation by calcium and calmodulin was completely inhibited by 25 microM trifluoperazine; at 50 microM it inhibited basal phosphorylation by 60%, suggesting that most of the basal phosphorylation may be due to the endogenous calmodulin present in our membrane preparation. NaDodSO4/polyacrylamide gel electrophoresis revealed that at least three of the phosphorylated species (both in the presence and in the absence of calcium and calmodulin) correspond to subunits of the purified acetylcholine receptor from T. californica (i.e., 65,000, 58,000, and 50,000 daltons) which are the beta, gamma, and delta subunits of the receptor.
DNA flexibility studied by covalent closure of short fragments into circles.
Authors: Shore D , Langowski J , Baldwin R L
Published: May 2006

The ring closure probability, or j factor, has been measured for DNA restriction fragments of defined sequence bearing EcoRI cohesive ends and ranging in size from 126 to 4361 base pairs (bp). The j factor is defined as the ratio of the equilibrium constants for cyclization and for bimolecular association via the cohesive ends. The end-joining reactions are fast compared to covalent closure of the cohesive ends by T4 DNA ligase. The rate of ligase closure is shown to be proportional to the equilibrium fraction of DNA molecules with joined cohesive ends, both in cyclization and in bimolecular association reactions. The j factor changes by less than 10-fold between 242 and 4361 bp, whereas it decreases by more than 100-fold between 242 and 126 bp as the DNA reaches the size range of the persistence length (150 bp). As regards ring closure, short DNA fragments are surprisingly flexible. These data are in good agreement with predictions by others for the ring closure probability of a wormlike chain.
Structure of the fibrin protofibril.
Authors: Fowler W E , Hantgan R R , Hermans J , Erickson H P
Published: May 2006

We identified the two-stranded fibrin protofibril and studied its structure in electron micrographs of negatively stained specimens. Based on these images and on considerations of symmetry, we constructed a model of the protofibril in which the two strands of trinodular fibrin molecules are related by a two-fold screw axis between the strands and two-fold axes perpendicular to them. The two strands are held together by staggered lateral contacts between the central nodules of one strand and outer nodules of the other. The molecules within a strand are joined by longitudinal contacts between outer nodules. This interpretation of the structure of protofibrils is supported by images of trimer complexes whose preparation and structure are described here, in which the central nodule of a fibrin monomer is attached to the crosslinked outer nodules of two other molecules. We conclude that the association of protofibrils to form thicker fibers must involve a second type of lateral contact, probably between outer nodules of adjacent, in-register strands. In total, we identify three intermolecular contacts involved in the polymerization of fibrin.
Cloning and analysis of strong promoters is made possible by the downstream placement of a RNA termination signal.
Authors: Gentz R , Langner A , Chang A C , Cohen S N , Bujard H
Published: May 2006

Downstream placement of a strong transcriptional termination signal has made possible the cloning of bacteriophage T5 promoters known to exhibit high signal strength. The cloning system constructed contains two easily assayable indicator functions whose expression is controlled by the integration of promoters and terminators, respectively. By assessing transcription within the indicator regions, the efficiency of promoters as well as termination signals can be determined in vitro and in vivo.
Are viroids escaped introns?
Authors: Diener T. O.
Published: May 2006

A complex of considerable stability is possible between the 5′ end of U1 RNA and a specific nucleotide sequence of the potato spindle tuber viroid complement. Because base-pairing between the 5′ end of U1 RNA and the ends of introns is believed by some to be responsible for the precise alignment and correct excision of introns, the U1-related sequence may represent the two ends of a presumed intron ancestor of the viroid complement after circularization.
Niche packing and coevolution in competition communities
Authors: Case Ted J.
Published: May 2006

A coevolutionary model of species packing is developed that allows evolutionary adjustment in both niche position and within-phenotype niche width of one-three competing species. The environment is specified as a single resource dimension
x
and availability of resources along
x
is given by a gaussian curve that has parameters
x
[unk]

R

and σ

R

. The model predicts that, for
S
species, the ratio of optimal niche width
w
[unk] to σ

R

is roughly independent of σ

R

and can be approximated by 1/
S
when the competitors are completely resource limited. Niche separation (
d
[unk]/
w
[unk]) increases only moderately with increases in resource diversity σ

R

and is greater for two than for three competing species. To the extent that the competitors are not completely resource limited, both coevolutionary niche separation and niche width decrease. Many of the general trends in niche width and niche separation predicted by this coevolutionary model parallel those from optimal foraging theory and limiting similarity models of community structure. The coevolutionary model stands out, however, in the singularly high values predicted for niche separation, making coevolved communities highly invasible. Hence, the theory suggests, as some empirical evidence indicates, that coevolved competition communities can only eixst as such on remote islands or in other habitats that might be free from invasion by outside species.
Mutation in an intervening sequence splice junction in man.
Authors: Orkin S H , Goff S C , Hechtman R L
Published: May 2006

The alpha 2-globin gene of an individual with alpha-thalassemia associated with the absence of alpha 2 mRNA was cloned in bacteriophage. This mutant globin gene was normally active in transcription in vitro. The DNA sequence of the gene, however, revealed a pentanucleotide deletion within the 5' splice junction of the first intervening sequence. Following the G of the invariant G-T dinucleotide normally located within such junctions, a deletion of T-G-A-G-G was found. No other sequence abnormalities within the mutant gene were present. We speculate therefore that this deletion within the splice junction is the primary genetic defect in this individual with thalassemia and that loss of a functional splice junction results in failure of stable mRNA formation.
Characterization, by immunoprecipitation, of myeloid- and monocyte-specific antigens present on the human promyelocytic cell line (HL-60) in three stages of differentiation.
Authors: Mulder A , Alexander S , Engelfriet C P , von dem Borne A E , Strominger J L
Published: May 2006

The human promyelocytic leukemia cell line HL-60 is reactive with an antiserum raised against normal human granulocytes (AGS). Immunoprecipitation with AGS on [35S]methionine-labeled HL-60 cell lysates with subsequent analysis by NaDodSO4/polyacrylamide slab gel electrophoresis shows a major antigenic doublet with molecular weights of 88,000 and 86,000, together with some minor antigens of lower molecular weight. Upon stimulation with dimethyl sulfoxide or 12-O-tetradecanoylphorbol 13-acetate, which induces HL-60 to differentiate to mature granulocytes or monocytes/macrophages, respectively, this antigenic doublet disappears. 12-O-Tetradecanoylphorbol 13-acetate induces the synthesis of an antigen, molecular weight 83,000, reactive with an antimonocyte serum. Neutrophil-specific alloantigens were not detected on HL-60 or its differentiated derivatives.
Homologous species restriction in lysis of erythrocytes by terminal complement proteins.
Authors: Hänsch G M , Hammer C H , Vanguri P , Shin M L
Published: May 2006

The cytolytic efficiency of the terminal complement protein complex, C5b-9, varies with the species of origin of C8 and C9. In the present study, we explored the susceptibility of erythrocytes from various species to lysis by C5b6,7 plus C8 and C9 from different species. EC5b6,7 intermediates were prepared on human, guinea pig, rabbit, mouse, and rat erythrocytes with human C5b6 and guinea pig C7. The degree of lysis of these intermediates by C8 and C9 was found to vary widely depending on the species of the proteins and the target cells. In all cases, lysis was least efficient when C8 and C9 were homologous with respect to the target cell species. This effect was mostly attributable to C9. The inefficient lysis in a homologous system is not due to a failure of C9 binding. Rather, the poor lysis in the homologous system may be attributable to inefficient insertion or channel formation.
Human trophoblast cell-surface antigens defined by monoclonal antibodies.
Authors: Lipinski M , Parks D R , Rouse R V , Herzenberg L A
Published: May 2006

A series of monoclonal antibodies has been raised against the human choriocarcinoma cell-line, BeWo. Four antigens, Trop-1, -2, -3, and -4, are defined on normal and malignant trophoblast cells. Trop-1 and Trop-2 appear to be specifically expressed on syncytio- and cytotrophoblasts, whereas Trop-3 and Trop-4 are also detected on various tumor cell lines, normal lymphocytes, and monocytes. Anti-Trop-1 and anti-Trop-2 antibodies might prove useful for detection and isolation of fetal trophoblast cells circulating in pregnant women's blood and for diagnosis and therapy in patients having choriocarcinomas and other germ-cell neoplasms.
Vasoactive intestinal polypeptide and cholinergic mechanisms in cat nasal mucosa: studies on choline acetyltransferase and release of vasoactive intestinal polypeptide.
Authors: Lundberg J M , Anggård A , Emson P , Fahrenkrug J , Hökfelt T
Published: May 2006

Quantitative immunohistochemical analysis of the sphenopalatine ganglion in normal cats revealed that virtually all ganglion cells (98.5%) were immunoreactive to vasoactive intestinal polypeptide (VIP). After surgical removal of this ganglion, the content of both VIP and choline acetyltransferase (acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6), a specific marker for cholinergic neurons, was decreased to about 70-80% in the nasal mucosa. In contrast, in animals subjected to sympathectomy combined with sensory (trigeminal) denervation, no significant change in VIP or choline acetyltransferase was found in the nasal mucosa. The present findings strongly suggest that a large proportion of the VIP neurons in the sphenopalatine ganglion contain choline acetyltransferase. This further supports the view that VIP is present in a population of autonomic cholinergic neurons innervating exocrine glands in the cat. In contrast, the ciliary ganglion contained high levels of choline acetyltransferase but no VIP. Parasympathetic nerve stimulation (15 Hz) caused a maximal vasodilation and a 500-fold increase in VIP output from the nasal mucosa. The plasma VIP immunoreactivity eluted at the same position as porcine VIP on gel permeation chromatography. Atropine pretreatment, which is known to abolish nasal secretion, caused a further 3-fold increase in VIP output during the nerve stimulation (15 Hz). Simultaneously, a markedly prolonged duration of the vasodilatory response was observed. The increased output of VIP during parasympathetic nerve stimulation by atropine pretreatment suggests that the transmitter acetylcholine may inhibit the release of the coexisting peptide--i.e., VIP--via muscarinic autoreceptors.
The hard-hexagon model and Rogers—Ramanujan type identities
Authors: Andrews George E.
Published: May 2006

In regime II of Baxter's solution of the hard-hexagon model [Baxter, R. J. (1980)
J. Phys. A
13, L61-L70], he presents six conjectures identifying certain one-dimensional partition functions with infinite products. An outline of the proof of these conjectures is given here.
Specific gangliosides function as host cell receptors for Sendai virus.
Authors: Markwell M A , Svennerholm L , Paulson J C
Published: May 2006

The ability of specific gangliosides to function as host cell receptors for Sendai virus was investigated by using Madin-Darby bovine kidney cells which become resistant to infection upon treatment with Vibrio cholerae sialidase. Sialidase-treated cells were incubated for 20 min at 37 degrees C with individual, highly purified gangliosides containing homogeneous carbohydrate moieties and then inoculated with virus for 10 min. Susceptibility of the cells to infection was monitored by hemagglutination titer of the virus produced 48 hr after inoculation. Incubation of the cells with gangliosides containing the sequence NeuAc alpha 2,3Gal beta 1,3GalNAc (i.e., GD1a, GT1b, and GQ1b) fully restored susceptibility to infection to the cells. However, the ganglioside GQ1b in which the sequence ends with two sialic acids in a NeuAc alpha 2,8NeuAc linkage instead of a single sialic acid as in GD1a and GT1b, was effective as a receptor at a concentration 1/100th that of any of the other gangliosides tested. Incubation with gangliosides similar in structure to GD1a, GT1b, and GQ1b but lacking the sialic acid attached to the terminal galactose (i.e., GM1 and GD1b) had no effect. The results from control experiments in which gangliosides were incubated at 0 degrees C with cells or in which trypsin was used to remove gangliosides adsorbed to cells were consistent with the premise that the gangliosides must actually insert into the cellular membrane to function as Sendai virus receptors. Addition of 4 X 10(6) molecules of 14C-labeled GD1a per cell made the cells fully susceptible to infection. Analysis of the ganglioside content of cell membranes showed that gangliosides GD1a, GT1b, and GQ1b are natural components of these cells and are present in quantities sufficient to act as receptors. These results demonstrate that gangliosides with the proper carbohydrate sequence, such as GD1a, GT1b, and GQ1b, function as natural receptors for Sendai virus in host cells.
Exchange of 1,N6-etheno-ATP with actin-bound nucleotides as a tool for studying the steady-state exchange of subunits in F-actin solutions.
Authors: Wang Y L , Taylor D L
Published: May 2006

The fluorescent analog of ATP 1-N6-ethenoadenosine 5'-triphosphate (epsilon-ATP) exchanges readily with nucleotides bound to G-actin. The exchange can be observed by measuring the fluorescence intensity, which increases significantly when epsilon-ATP binds to actin. When excess epsilon-ATP is added to a solution of F-actin, a continuous increase in fluorescence intensity is observed, indicating that the nucleotides bound to F-actin are directly or indirectly exchangeable. The kinetics of exchange consist of a fast phase and a slow phase. Both phases are stimulated by shearing and are inhibited by phalloidin treatment, suggesting that the exchange of nucleotides is coupled to the exchange of subunits. Therefore, the exchange reaction can be used as a convenient, nonperturbing tool to study the exchange of free actin subunits with subunits in actin filaments. The exchange of actin subunits was characterized by a pulse-chase experiment. The results suggest that actin subunits assemble and disassemble through the same end of the filament during the fast phase of exchange but through opposite ends of the filament during the slow phase. In addition, the slow phase of exchange is inhibited in the absence of millimolar magnesium ions, but is not significantly affected by cytochalasin B at concentrations between 0.1 and 10 microM. These observations are discussed in relation to possible mechanisms of subunit exchange in steady-state F-actin solutions.
B850 pigment-protein complex of
Rhodopseudomonas sphaeroides
: Extinction coefficients, circular dichroism, and the reversible binding of bacteriochlorophyll
Authors: Clayton Roderick K. , Clayton Betty J.
Published: May 2006

Chromatophores of
Rhodopseudomonas sphaeroides
yield the antenna complex B850 in either of two states, depending on the method of isolation. Methods using dodecyl (= lauryl) dimethylamine oxide yield B850 with an absorption spectrum like that
in vivo
: the bands at 800 and 850 nm, due to the bacteriochlorophyll (Bchl) components Bchl-800 and Bchl-850, are in ratio
A
800
/
A
850
= 0.65 ± 0.05. When B850 is isolated by methods using dodecyl sulfate, the Bchl-800 is attenuated or absent. Bchl assays of these materials and of the isolated antenna complex B875 yielded the following extinction coefficients, ±SD, on the basis of the molarity of Bchl: For B875, ε
875
= 126 ± 8 mM
-1
cm
-1
. For B850 in the normal (high-Bchl-800) state, ε
850
= 132 ± 10 mM
-1
cm
-1
. For the individual components of Bchl in B850, ε
850
of Bchl-850 = 184 ± 13 mM
-1
cm
-1
and ε
800
of Bchl-800 = 213 ± 28 mM
-1
cm
-1
. With these coefficients the molecular ratio of Bchl-850 to Bchl-800 equals 1.8 ± 0.4 for B850 in the high-Bchl-800 state. Starting with B850 depleted of Bchl-800, the addition of dodecyldimethylamine oxide restored the 800-nm absorption band. The 850-nm band became shifted toward the blue, narrowed, and slightly attenuated, and its associated circular dichroism became 20% more intense. Free Bchl added with dodecyldimethylamine oxide accelerated the restoration of Bchl-800 and retarded the attenuation of Bchl-850. We conclude that free Bchl can interact reversibly with a binding site for Bchl-800 in the B850 complex, with dodecyl sulfate favoring dissociation and dodecyldimethylamine oxide promoting association. Thus the reversible dissociation of a native chlorophyll-protein complex has now been demonstrated.
Polymorphic DNA region adjacent to the 5' end of the human insulin gene.
Authors: Bell G I , Karam J H , Rutter W J
Published: May 2006

The length of a segment of DNA associated with the human insulin gene, which has been localized to the short arm of chromosome 11, is heterozygous in 63% of 52 individuals analyzed. This polymorphic region is approximately 500 base pairs from the nucleotide encoding the 5' end of insulin mRNA. The polymorphism appears to be due to an insertion or deletion of DNA sequences so that DNA fragments of different length are generated when DNA from a heterozygous individual is digested with selected restriction endonucleases.
Nucleotide sequence of constant and 3' untranslated regions of a kappa immunoglobulin light chain mRNA of a homozygous b4 rabbit.
Authors: Heidmann O , Auffray C , Cazenave P A , Rougeon F
Published: May 2006

A homozygous a2/a2 and b4/b4 rabbit has been hyperimmunized with Micrococcus lysodeikticus. Poly(A)-containing RNA has been isolated from the spleen and translated in vitro, and translation products have been analyzed by NaDodSO4/polyacrylamide gel electrophoresis. Double-stranded cDNA has been synthesized from poly(A)-containing RNA template and inserted in the Pst I endonuclease site of plasmid pBR322 by using the oligo(dC).oligo(dG) tailing procedure. Tetracycline-resistant ampicillin-sensitive clones containing cDNA complementary to a kappa light chain mRNA have been selected by differential screening and their ability to hybridize to a spleen mRNA having the same size as a mouse kappa light chain mRNA. Two clones, pRk-15 and pRk-32, have been selected to determine the nucleotide sequence of the constant and 3' untranslated regions of kappa light chain mRNA, by the Maxam and Gilbert partial degradation method. Comparison of homologous regions of mouse kappa chain mRNA and b4 rabbit kappa chain mRNA reveals 61% homology in the constant region and 59% homology in the 3' untranslated region.
Restricted reassociation of heavy and light chains from hapten-specific monoclonal antibodies.
Authors: Kranz D M , Voss E W
Published: May 2006

Six murine monoclonal antifluorescyl antibody clones encompassing a defined range of affinities and containing kappa light chains with IgG1 or IgG2 heavy chains were examined. As the fluorescence of the ligand is quenched greater than 90% when fluorescein is bound by antifluorescyl antibodies, fluorescence quenching was assayed to monitor polypeptide reconstitution and active site formation on mixing of resolved heavy (H) and light (L) chains. Of 36 possible experimental combinations of H- and L-chain reaction mixtures, only homologous H and L chains (derived from the same parental immunoglobulin molecule) bound fluorescein. Results from fluorescence polarization studies, conducted independently of fluorescence quenching, confirmed the findings. Competitive inhibition and molecular sieve experiments showed that, despite preferential association of homologous H and L chains, several heterologous H and L chains associated to form intact 7S molecules, although no active site was constituted. Thus, polypeptide recombination and formation of functional antigen binding sites are two processes that immunocytes must regulate during cell differentiation and generation of diversity. A mechanism and underlying the observed preferential reassociation of specific H and L chains and a means of generating affinity maturation, as exhibited by the antifluorescein system, is proposed.
Decrease in macrophage antigen catabolism caused by ammonia and chloroquine is associated with inhibition of antigen presentation to T cells.
Authors: Ziegler H K , Unanue E R
Published: May 2006

This paper describes the effects of two lysosomotropic compounds, ammonia and chloroquine, on the interaction of mononuclear phagocytes with immune T cells. The uptake and ingestion of Listeria monocytogenes by macrophages were not affected by the drugs; however, the macrophage catabolism of 125I-labeled Listeria was reduced in a dose-dependent way. The macrophage presentation of Listeria to T cells, an I-region-dependent phenomenon, was also inhibited. The degree of inhibition of catabolism paralleled that of antigen presentation. The inhibition of antigen presentation took place if the macrophages were treated before and during Listeria uptake; minimal inhibition took place if the macrophages were treated 30 min after Listeria ingestion, at which time a significant amount of bacteria was already catabolized. Our previous studies had shown that the macrophage-associated antigen recognized by T cells became apparent 30-60 min after uptake of Listeria. We conclude that ammonia and chloroquine affected an intracellular handling step required for the expression of the immunogen relevant for T-cell recognition.
Nucleotide sequence and the encoded amino acids of human serum albumin mRNA.
Authors: Dugaiczyk A , Law S W , Dennison O E
Published: May 2006

The complete nucleotide sequence of human serum albumin mRNA has been determined from recombinant cDNA clones and from a primer-extended cDNA synthesis on the mRNA template. The sequence is composed of 2078 nucleotides, starting upstream from a potential ribosome binding site in the 5' untranslated region. It contains all the translated codons and extends into the poly(A) at the 3' terminus. Part of the translated sequence codes for a hydrophobic prepeptide, Met-Lys-Trp-Val-Thr-Phe-Ile-Ser-Leu-Leu-Phe-Leu-Phe-Ser-Ser-Ala-Tyr-Ser, followed by a basic propeptide, Arg-Gly-Val-Phe-Arg-Arg. These signal peptides are absent from mature normal serum albumin and, so far, have not been identified in their nascent state in humans. A remaining 1755 nucleotides of the translated mRNA sequence code for 585 amino acids, which are in agreement, with few exceptions, with the published amino acid sequence for human serum albumin. The mRNA sequence verifies and refines the repeating homology in the triple-domain structure of the serum albumin molecule.
Human ribosomal RNA gene: nucleotide sequence of the transcription initiation region and comparison of three mammalian genes.
Authors: Financsek I , Mizumoto K , Mishima Y , Muramatsu M
Published: May 2006

The transcription initiation site of the human ribosomal RNA gene (rDNA) was located by using the single-strand specific nuclease protection method and by determining the first nucleotide of the in vitro capped 45S preribosomal RNA. The sequence of 1,211 nucleotides surrounding the initiation site was determined. The sequenced region was found to consist of 75% G and C and to contain a number of short direct and inverted repeats and palindromes. By comparison of the corresponding initiation regions of three mammalian species, several conserved sequences were found upstream and downstream from the transcription starting point. Two short A + T-rich sequences are present on human, mouse, and rat ribosomal RNA genes between the initiation site and 40 nucleotides upstream, and a C + T cluster is located at a position around -60. At and downstream from the initiation site, a common sequence, T-AG-C-T-G-A-C-A-C-G-C-T-G-T-C-C-T-CT-T, was found in the three genes from position -1 through +18. The strong conservation of these sequences suggests their functional significance in rDNA. The S1 nuclease protection experiments with cloned rDNA fragments indicated the presence in human 45S RNA of molecules several hundred nucleotides shorter than the supposed primary transcript. The first 19 nucleotides of these molecules appear identical--except for one mismatch--to the nucleotide sequence of the 5' end of a supposed early processing product of the mouse 45S RNA.
Ferritin: a zinc detoxicant and a zinc ion donor.
Authors: Price D , Joshi J G
Published: May 2006

Rats were injected with 1 mg of Zn2+ as zinc sulfate or 2 mg of Cd2+ as cadmium sulfate per kg of body weight on a daily basis. After seven injections, ferritin and metallothionein were isolated from the livers of the rats. Significant amounts of zinc were associated with ferritin. Incubation of such ferritin with apoenzymes of calf intestinal alkaline phosphatase, yeast phosphoglucomutase, and yeast aldolase restored their enzymic activity. The amount of zinc injected was insufficient to stimulate significant synthesis of metallothionein, but similar experiments with injection of cadmium did stimulate the synthesis of metallothionein. The amount of Zn2+ in ferritin of Cd-injected rats was greater than that in ferritin in Zn-injected rats, which was greater than that in ferritin of normal rats. Thus at comparable protein concentration ferritin from Cd-injected rats was a better Zn2+ donor than was ferritin from Zn-injected or normal animals. Ferritin is a normal constituent of several tissues, whereas metallothionein is synthesized under metabolic stress. Thus ferritin may function as a "metal storage and transferring agent" for iron and for zinc. It is suggested that ferritin probably serves as the initial chelator for Zn2+ and perhaps other metal ions as well and that under very high toxic levels of metal ions the synthesis of metallothionein is initiated as the second line of defense.
Trp aporepressor production is controlled by autogenous regulation and inefficient translation.
Authors: Kelley R L , Yanofsky C
Published: May 2006

We constructed a trpR-lacZ gene fusion that specifies a hybrid protein that has full beta-galactosidase activity. The gene fusion was associated with the unaltered trpR transcription and translation control region; thus, hybrid beta-galactosidase production was an indicator of expression of the trp aporepressor (trpR) operon. To facilitate in vivo expression studies, a DNA segment containing the trpR-lacZ gene fusion and the trpR controlling region was transferred to bacteriophage lambda and subsequently inserted into the bacterial chromosome. Analyses of hybrid beta-galactosidase production showed that the trpR operon is regulated autogenously but that the rate of synthesis of aporepressor varies only 4- to 5-fold in response to changes in the intracellular concentration of tryptophan. Under comparable conditions, the trp operon is regulated by trp repressor approximately 70-fold. Therefore, the operators of the trp operon and the trpR operon must have very different affinities for trp repressor in vivo. The promoter controlling trpR expression was found to be moderately active. Nevertheless, there are only about 50-300 molecules of trp aporepressor per cell. The low aporepressor level appears to be due to inefficient translation of trpR mRNA.
Identification of an antigen associated with transforming genes of human and mouse mammary carcinomas.
Authors: Becker D , Lane M A , Cooper G M
Published: May 2006

Sera from tumor-bearing mice immunoprecipitated a 86,000-dalton glycoprotein from extracts of NIH cells transformed by human mammary carcinoma DNA. This antigen was not immunoprecipitated from extracts of NIH 3T3 cells, spontaneously transformed NIH cells, NIH cells transformed by normal human DNA, NIH cells transformed by human bladder carcinoma DNA, or NIH cells transformed by Rous sarcoma virus DNA. In addition, sera from mice bearing tumors induced by NIH cells transformed by either normal human DNA or human bladder carcinoma DNA did not immunoprecipitate this antigen from extracts of NIH cells transformed by human mammary carcinoma DNA. However, this antigen was immunoprecipitated by sera from mice bearing tumors induced by NIH cells transformed by mouse mammary carcinoma DNAs and from mice bearing primary mammary carcinomas. These results indicate that this glycoprotein represents an antigen that is specifically associated with expression of the transmissible transforming genes of human and mouse mammary carcinomas.
Formation of covalent adducts between cortisol and 16 alpha-hydroxyestrone and protein: possible role in the pathogenesis of cortisol toxicity and systemic lupus erythematosus.
Authors: Bucala R , Fishman J , Cerami A
Published: May 2006

The incubation of albumin with cortisol or 16 alpha-hydroxyestrone results in the formation of covalent steroid-protein adducts. The rate of adduct formation increases in the presence of sodium cyanoborohydride (NaCNBH3), indicating that the reaction proceeds nonenzymatically through a Schiff base intermediate. Under nonreducing conditions, a stable adduct forms with cortisol and 16 alpha-hydroxyestrone but not with estrone, which lacks a hydroxyl group adjacent to the reactive carbonyl. It is hypothesized that a Heyns rearrangement involving the adjacent hydroxyl group traps the Schiff base and produces a stable ketoamine adduct. The binding of 16 alpha-hydroxyestrone and cortisol to albumin is significantly inhibited by acetylsalicylic acid, which has been shown to acetylate an epsilon-amino group of a lysine residue in albumin. High-pressure liquid chromatography analysis of an acid hydrolysate of 16 alpha-hydroxyestrone-albumin shows that a product containing 16 alpha-hydroxyestrone coelutes with a standard prepared by reacting 16 alpha-hydroxyestrone with the epsilon-amino group of lysine. We propose that the formation of covalent steroid-protein adducts is a generalized phenomenon which may contribute to the pathological effects produced by elevated levels of certain endogenous steroids.
1,25-Dihydroxyvitamin D3: short- and long-term effects on bone and calcium metabolism in patients with postmenopausal osteoporosis.
Authors: Gallagher J C , Jerpbak C M , Jee W S , Johnson K A , DeLuca H F , Riggs B L
Published: May 2006

We evaluated the effects of therapy with a small dose (0.5 microgram/day) of 1,25-dihydroxyvitamin D3 in 12 previously untreated patients with postmenopausal osteoporosis. Combined radiocalcium kinetic and balance studies showed that at base line, net calcium absorption (mean +/- SEM) was low (7 +/- 3%), calcium balance was negative (-59 +/- 22 mg/day), and bone resorption rate (297 +/- 40 mg/day) exceeded bone formation rate (239 +/- 36 mg/day). After short-term therapy (6-8 months), calcium absorption was normal (27 +/- 3%, P less than 0.001 for difference from base line), calcium balance had improved (+2 +/- 26 mg/day, P less than 0.05), and bone resorption rate (195 +/- 35 mg/day, P less than 0.01) had decreased more than bone formation rate (197 +/- 26 mg/day, NS). Compared with base-line values, after long-term therapy (2 years) the increased level of calcium absorption was maintained (27 +/- 3%, P less than 0.001), but calcium retention declined to a level intermediate (-27 +/- 24 mg/day, NS) between base-line and short-term treatment values, and both bone resorption rate (294 +/- 33 mg/day, NS) and bone formation rate (267 +/- 34 mg/day, NS) increased. Urinary hydroxyproline excretion was lower than before treatment (26.3 +/-2.0 mg/day) after both short-term (21.1 +/- 1.6 mg/day, P less than 0.001) and long-term (22.0 +/- 1.8 mg/day, P less than 0.01) treatments. Trabecular bone volume was 11.3 +/- 1.0% at base line and increased to 16.0 +/- 1.2% (P less than 0.01) after long-term treatment. By contrast, the eight patients studied before and after 6-8 months of placebo treatment had no significant change in any variable.
Molecular motion of DNA as measured by triplet anisotropy decay.
Authors: Hogan M , Wang J , Austin R H , Monitto C L , Hershkowitz S
Published: May 2006

We have used triplet anisotropy decay techniques to measure the internal flexibility and overall rotational motion of DNA, covering a time range from 15 ns to 200 mus. Nearly monodisperse DNA fragments 65--600 base pairs long were studied by using the intercalating dye methylene blue as a triplet probe. We found that the slow end-over-end tumbling of short DNA fragments (less than or equal to 165 base pairs) is as predicted for a rigid rod. As expected, a longer DNA fragment (600 base pairs) experiences slow segmental motion of its helix axis. We found that, at the earliest times, anisotropy decays more rapidly than expected for a rigid rod, suggesting that, when bound, methylene blue monitors fast internal motion of the helix. Since the rod-like end-over-end tumbling of short fragments rules out fast bending motions, we conclude that the fast components of DNA anisotropy decay are due to twisting motion of the helix, occurring with a time constant near 50 ns.
Functional reconstitution of the purified sodium channel protein from rat sarcolemma.
Authors: Weigele J B , Barchi R L
Published: May 2006

The purified saxitoxin (STX) binding component of the rat sarcolemmal sodium channel (SBC) has been reconstituted into phospholipid vesicles. The reconstituted SBC displays the pharmacological properties and the ability to control sodium fluxes expected of a functional sodium channel. Batrachotoxin (BTX) increases 22Na+ influx into reconstituted SBC vesicles by greater than 100% over control at early time points. The BTX-stimulated 22Na+ influx is specifically and quantitatively blocked by STX. Veratridine and aconitine also stimulate Na+-flux--although less effectively than BTX--in the order: BTX greater than veratridine greater than aconitine. The logarithmic dose--response curves for BTX and veratridine are sigmoidal with a K0.5 of 1.5 microM and 35 microM, respectively. Vesicles containing the reconstituted SBC demonstrate 3H-labeled STX binding to a single class of high affinity sites witha Kd of 5--7 nM at 0 degrees C; the thermal stability of the STX receptor is markedly enhanced by reconstitution. Our results confirm that the purified STX binding component from rat sarcolemma constitutes the sodium channel itself and contains at least those components sufficient for channel activation, transmembrane ion movement, and inhibition by STX.
Complete amino acid sequence of an HLA-DR antigen-like beta chain as predicted from the nucleotide sequence: similarities with immunoglobulins and HLA-A, -B, and -C antigens.
Authors: Larhammar D , Schenning L , Gustafsson K , Wiman K , Claesson L , Rask L , Peterson P A
Published: May 2006

The complete nucleotide sequence of an HLA-DR antigen-like beta-chain cDNA clone was determined. The 1,080 base pairs include the complete coding region and most of the untranslated portion. The predicted amino acid sequence has 229 residues. The beta chain contains two immunoglobulin-like disulfide loops and a 21-amino acid residue membrane-integrated segment. Ten amino acid residues reside on the cytoplasmic side of the plasma membrane. The single asparagine-linked carbohydrate moiety is attached to asparagine-19. The NH2-terminal 91 residues of the beta chain are homologous to the corresponding region of HLA-A, -B, and -C antigen heavy chains. Residues 92-192 of the beta chain display statistically significant homology to members of the immunoglobulin family, beta 2-microglobulin, and the immunoglobulin-like domain of HLA-A, -B, and -C antigen heavy chains. These data establish that the major histocompatibility antigens of class I and class II type and the constant regions of immunoglobulins are evolutionarily related.
Stimulation of Xenopus oocyte protein synthesis by microinjected adenovirus RNA.
Authors: Richter J D , Jones N C , Smith L D
Published: May 2006

The ability of injected heterologous mRNAs to compete with endogenous mRNAs in Xenopus oocytes was assayed. In confirmation of previous reports, globin mRNA translation in oocytes results in a concomitant decrease in endogenous protein synthesis. In contrast, injection of adenovirus 5 mRNA into the oocyte results in a stimulation of endogenous protein synthesis. The stimulation is dose dependent and does not require nuclear transcription in the oocyte. Preliminary mapping data suggest that the stimulatory RNA is a product of one of the viral immediate early genes.
Activation of a suppressor T-cell pathway by interferon.
Authors: Aune T M , Pierce C W
Published: May 2006

In addition to antiviral activities, murine fibroblast (type I) interferon (IFN-beta) suppresses immune responses. The mechanism(s) by which IFN-beta suppresses antibody responses by murine spleen cells to sheep erythrocytes in vitro has been investigated. IFN-beta-mediated suppression is partially or completely prevented by catalase, 2-mercaptoethanol, and certain peroxidase substrates (ascorbic acid, potassium iodide, and tyrosine). These same reagents also block suppression by mediators from concanavalin A-activated murine suppressor T cells, soluble immune response suppressor (SIRS)/macrophage-derived suppressor factor (Mphi-SF), and act by inactivating Mphi-SF or preventing formation of Mphi-SF from SIRS. Therefore, these experiments suggested that IFN-beta may act by inducing production of a molecule that has properties of SIRS. Treatment of spleen cells with IFN-beta leads to generation of a population of Lyt2+ suppressor T cells that acts by elaborating a soluble factor. This IFN-beta-induced suppressor T-cell factor (IFN-TsF) has properties in common with SIRS. First, both SIRS and IFN-TsF suppress antibody responses with the same characteristic kinetic pattern; responses initiate normally but prematurely terminate after day 4 of culture. Second, IFN-TsF and SIRS are of comparable size (45,000-55,000 daltons) and are converted to Mphi-SF by low (1 microM) concentrations of H2O2 or by macrophages. Third, Mphi-SF obtained from IFN-TsF or SIRS is inactivated by similar concentrations of reagents such as ascorbic acid, potassium iodide, and 2-mercaptoethanol. These data show that the immunosuppressive properties of IFN-beta are due, at least in part, to its ability to activate suppressor T cells that produce mediators that appear to be analogous to those in the SIRS/Mphi-SF pathway of immunosuppression.
Enhancing antibody: a novel component of the immune response.
Authors: Nemazee D A , Sato V L
Published: May 2006

Current descriptions of the immune response identify two classes of antigenic stimuli that result in the production of specific antibody: (i) exogenous antigens and (ii) endogenous variable-region determinants of the immune system. We expand this scheme to include a third class of antigenic stimulus--new determinants created by the binding of antibody to antigen. This paper describes a set of monoclonal antibodies which arose after repeated immunization with antigen alone but which bound antibody--antigen complexes. These antibodies recognize determinants on the antibody portion of the complexes that were expressed as a consequence of antigen binding. Antibodies of this general type, "enhancing antibodies," which can strengthen antibody--antigen and idiotypic-anti-idiotypic antibody interactions, may play important regulatory and effector roles in the immune response. We suggest a model that predicts the occurrence and specificity of different classes of such antibodies and provides a conceptual framework that gives a straightforward explanation of the appearance in the immune response of rheumatoid antibodies and of antibodies that bind cooperatively to antigen.
Growth factor activation of an amiloride-sensitive Na+/H+ exchange system in quiescent fibroblasts: coupling to ribosomal protein S6 phosphorylation.
Authors: Pouysségur J , Chambard J C , Franchi A , Paris S , Van Obberghen-Schilling E
Published: May 2006

Chinese hamster lung fibroblast cells (CCl39) enter the G0/G1 nonproliferative state after serum deprivation. In this report, we show that reinitiation of DNA synthesis by serum or the combination of purified human thrombin and insulin (1-10 microgram/ml) is preceded by very early stimulation of ionic fluxes (Na+/Rb+) and protein phosphorylation (27,000 daltons, 62,000 daltons, and the ribosomal S6 proteins). The potentiating action of insulin on thrombin-stimulated DNA synthesis is also observed on thrombin-stimulated Na+ influx, Rb+ influx, and protein S6 phosphorylation. Moreover, we demonstrate that CCl39 cells possess a Na+/H+ exchange system sensitive to amiloride. Half-maximal inhibition of growth factor-activated Na+ influx and Na+-dependent H+ efflux is obtained with 3-10 microM amiloride. Two lines of evidence indicate that the extrusion of H+ via the activation of the Na+/H+ exchanger is coupled to protein S6 phosphorylation: serum-stimulated phosphorylation is blocked by (i) amiloride at a concentration that abolishes serum-stimulated Na+ influx and (ii) protonophores that acidify the cell interior. The present data support the idea that the regulation of intracellular pH is a key event in the mechanism of growth factor action.
Synthesis of thymosin beta 4 by peritoneal macrophages and adherent spleen cells.
Authors: Xu G J , Hannappel E , Morgan J , Hempstead J , Horecker B L
Published: May 2006

Thymosin beta 4, previously identified as a component of thymosin fraction 5 isolated from calf thymus, is synthesized by peritoneal macrophages derived from rats or mice, including cells from nu/nu mice. Identification of thymosin beta 4 containing [35S]methionine was based on its retention time in reverse-phase HPLC and the recovery of radioactivity in peptides generated by mild acid hydrolysis or by digestion with trypsin. Thymosin beta 4 is also synthesized by rat and mouse spleen cells, and evidence is presented for its release from these cells.
Novel class of mouse mammary tumor virus-related DNA sequences found in all species of Mus, including mice lacking the virus proviral genome.
Authors: Callahan R , Drohan W , Gallahan D , D'Hoostelaere L , Potter M
Published: May 2006

Mice in breeding colonies of feral Mus musculus brevirostris (Azrou, Morocco), M. m. musculus (Studenec, Czechoslovakia), and M. m. molossinus (Fukuoka, Japan) were found to lack the mouse mammary tumor virus (MMTV-alpha) proviral genome in their germ line. MMTV-alpha proviral genomes have been found in all inbred strains of M. musculus by using high-stringency nucleic acid hybridization conditions. We conclude that feral mice in these colonies are heterozygous for a limited number of MMTV-alpha proviral genomes and that those lacking them arose as a result of random chromosomal segregation. All mice in another breeding colony of feral M. m. musculus (Sladeckovce, Czechoslovakia) lack MMTV proviral genes. By relaxing the conditions of nucleic acid hybridization, MMTV-related sequences (designated MMTV-beta) were detected in restricted cellular DNA from MMTV-negative mice and all other inbred strains and feral species of the genus Mus. The apparent ubiquity of the MMTV-beta DNA sequences in the genus Mus and the lack of variation in the pattern of restriction fragments containing these sequences within a species distinguishes them from MMTV-alpha. These results suggest that the MMTV-beta DNA sequences either are the evolutionary progenitors of the infectious MMTV genome or represent an accumulation of evolutionarily divergent MMTV-alpha insertions into the mouse germ line.