bluuebunny/crossref_metadata_2025 huggingface dataset:
Structural geology of the Earth's exterior
Authors: Burchfiel B. C.
Published: May 2006

Plate tectonics offers an explanation for the present motions and heterogeneity of the rocks that form the external part of the Earth. It explains the origin of the first-order heterogeneity of oceanic and continental lithospheres. Furthermore, it explains the youth and simplicity of the oceanic lithosphere and offers the potential to explain the antiquity, complexity, and evolution of the continental lithosphere. The framework of plate tectonics must be used carefully, because there are geological features within continents, particularly in the more ancient rocks, that may require alternative explanations. The task of understanding lithospheric motions through geologic time must be focused on the continents, where the major evidence for 95% of Earth history resides.

In interpreting earth motions from the geologic record, three needs seem paramount: (
i
) to develop a three-dimensional understanding of the kinematics, dynamics, and thermal structure of modern plate boundary systems and at the same time to recognize those geological and geophysical features that are unrelated to plate interaction; (
ii
) to use this understanding to reconstruct the extent and evolution of ancient systems that form the major elements of continental crust; and (
iii
) to determine the dynamics and evolution of systems that have no modern analogs. Decoupling along subhorizontal zones within the lithosphere may be widespread in all types of plate boundary systems. Thus, in order to interpret the motion and dynamics of the mantle correctly, it is important to know if upper lithospheric motion within boundary systems is controlled directly or indirectly by or is independent of deeper mantle motions.
Organelle DNA variation and systematic relationships in the genus
Zea
: Teosinte
Authors: Timothy D. H. , Levings C. S. , Pring D. R. , Conde M. F. , Kermicle J. L.
Published: May 2006

Chloroplast and mitochondrial DNAs from six races of annual teosinte (Guatemala, Huehuetenango, Balsas, Central Plateau, Chalco, and Nobogame), perennial teosinte, and maize were compared and grouped by restriction endonuclease fragment analyses. Three groups of chloroplast DNAs were detected: (
i
) perennial teosinte and Guatemala; (
ii
) Balsas and Huehuetenango; and (
iii
) all other teosintes. Four groups of mitochondrial DNAs were separated: (
i
) perennial teosinte; (
ii
) Guatemala; (
iii
) Nobogame; and (
iv
) all other teosintes. Separation of the teosinte and maize organelle DNAs into five groups (Guatemala; perennial teosinte; Balsas and Huehuetenango; Central Plateau and Chalco; Nobogame and maize) approximated the biosystematic relationships of the taxa. It was suggested that the evolutions of the chloroplast and mitochondrial DNAs may be independent of each other, that variation of organelle DNA within a species complex of an organism may be the common condition, and that the DNAs of the organelle and nuclear systems evolve in reasonable harmony.
Enzymatic methyl esterification of specific glutamyl residue in corticotropin.
Authors: Kim S , Li C H
Published: May 2006

Ovine corticotropin (alpha s-ACTH) was enzymatically methylated with purified calf brain protein methylase II (protein O-methyltransferase; S-adenosyl-L-methionine: protein-carboxyl O-methyltransferase, EC 2.1.1.24) and S-adenosyl-L-[methyl-14C]methionine. After incubation for 60 min at 37 degrees C, 30 mol % of the hormone was methylated on the basis of the [14C]methyl incorporation. In order to assess the location of methylation, the modified peptide was digested with pepsin. Analytical results derived from studies on the peptic digest led to the suggestion that the alpha s-ACTH-(6--28) peptide fragment was esterified. Because there is only one available methylation site at Glu-28, these results indicate that Glu-28 of alpha s-ACTH was specifically methyl esterified to yield [Glu(OMe)28]-alpha s-ACTH.
A general priming system employing only dnaB protein and primase for DNA replication.
Authors: Arai K , Kornberg A
Published: May 2006

Priming of phage phi X174 DNA synthesis is effected simply by dnaB protein and primase when the DNA is not coated by single-strand binding protein (SSB). The five prepriming proteins (n,n',n'',i, and dnaC protein) required for priming a SSB-coated phi X174 DNA circle are dispensable. The dnaB protein-primase priming system is also active on uncoated phage G4 and M13 DNAs and on poly(dT). Multiple RNA primers, 10--60 nucleotides long, are transcribed with patterns distinctive for each DNA template. Formation of a stable dnaB protein.DNA complex in the presence of primase and ATP supports the hypothesis that dnaB protein provides a mobile replication promoter signal for primase.
Photooxidation of antenna bacteriochlorophyll in chromatophores from carotenoidless mutant
Rhodopseudomonas sphaeroides
and the attendant loss of dimeric exciton interaction
Authors: Rafferty C. N. , Bolt J. , Sauer K. , Clayton R. K.
Published: May 2006

Intense continuous illumination of purified chromatophores from carotenoidless mutant
Rhodopseudomonas sphaeroides
results in progressive photooxidative loss of the near infrared absorption band near 860 nm assigned to antenna bacteriochlorophyll. The quantum yield of this reaction is low, approximately 1.7 × 10
-5
. The loss in near infrared absorption is accompanied by a proportional shift in the absorption maximum to shorter wavelengths. The double circular dichroism feature in the near infrared decreases at a faster rate than does the absorbance. These results are explained by a model in which the antenna bacteriochlorophyll, initially associated as dimers (λ
max
= 860.2 nm), is progressively converted to the monomeric state (λ
max
= 851.9 nm). The wavelength shift is attributed to disruption of exciton coupling in the dimer. Acetone/methanol extraction indicates that the maximum molar extinction coefficients of the dimer and monomer do not differ by more than 4%. The occurrence of an absorption maximum at 852 nm for monomeric bacteriochlorophyll in a protein complex demonstrates that it is not necessary to invoke aggregation of the chromophores as the origin of the shift from 770 nm in typical organic solvents.
Requirement for membrane potential in injection of phage T4 DNA.
Authors: Labedan B , Goldberg E B
Published: May 2006

The first stages of infection by phage T4 may be divided into energy-dependent and energy-independent processes. Irreversible adsorption, unplugging, and initial exposure of the DNA terminus may occur at 4 degrees C, or at 37 degrees C in bacteria whose energy-yielding metabolism has been poisoned. DNA injection into the cytoplasm needs higher temperatures and energy from the host cell. The nature of this energy requirements was deduced from the use of metabolic inhibitors. Our results show that T4 DNA injection specifically requires the presence of a protonmotive force across the cytoplasmic membrane of the host. Moreover, the chemical gradient (delta pH) does not appear to be essential, but the membrane potential (delta psi) is required.
Interaction of phencyclidine ("angel dust") with a specific receptor in rat brain membranes.
Authors: Vincent J P , Kartalovski B , Geneste P , Kamenka J M , Lazdunski M
Published: May 2006

[3H]Phencyclidine binds to synaptic membranes from rat brain in a saturable, reversible, and selective fashion, with a dissociation constant Kd of 0.25 microM and a maximal binding capacity of 2.4 pmol/mg of membrane protein--i.e., 250 pmol/g of brain. The binding activity is concentrated in synaptosomal fractions, is higher in cerebral cortex and corpus striatum than in other parts of the rat brain, and is not detectable in the spinal cord. Only molecules of the phencyclidine series and ketamine are able to bind to the phencyclidine receptor. [3H]Phencyclidine bound to its receptor is not displaced by the classical neurotransmitters or neuromodulators. There is a good correlation between the apparent affinities of a series of phencyclidine analogs for the phencyclidine receptor and the pharacological activities of these analogs as measured by the rotarod assay.
Selective loss of noradrenergic phenotypic characters in neuroblasts of the rat embryo.
Authors: Jonakait G M , Wolf J , Cochard P , Goldstein M , Black I B
Published: May 2006

To define the fate of embryonic neuroblasts in rat gut, which transiently express several noradrenergic traits, we investigated the high-affinity uptake of norepinephrine. At 12.5 days of gestation, these cells exhibited immunoreactivity to tyrosine hydroxylase [tyrosine 3-monoxygenase; L-tyrosine, tetrahydropteridine:oxygen oxidoreductase (3-hydroxylating), EC 1.14.16.2] and endogenous catecholamine fluorescence. However, by 13.5 days these noradrenergic neurotransmitter phenotypic characters essentially disappeared. In contrast, norepinephrine uptake, which was also apparent at 12.5 days, persisted at least through 17.5 days. These observations indicate that norepinephrine uptake develops as an additional noradrenergic characteristic in these cells and persists after the disappearance of other noradrenergic traits. Consequently, neurotransmitter phenotypic characters may be transiently displayed during normal development in vivo.
Double-stranded RNA inhibits a phosphoprotein phosphatase present in interferon-treated cells.
Authors: Epstein D A , Torrence P F , Friedman R M
Published: May 2006

In accord with previous studies, (I)n . (C)n, a potent inhibitor of the cell-free protein-synthesizing system of interferon-treated L cells, stimulates incorporation of 32P from [gamma-32P]ATP into the 67,000-dalton protein, P1. The double-stranded RNA (I)n . (br5C)n, which is inactive as an inhibitory of protein synthesis, does not stimulate phosphorylation of P1 under conditions approximating those of protein synthesis. However, we have found conditions under which (I)n . (br5C)n is approximately as effective as (I)n . (C)n in stimulating incorporation of label from [gamma-32P]ATP into 67,000-dalton protein. Upon transfer of labeled P1 from these conditions to those compatible with protein synthesis, there is a time-dependent decrease in label in the 67,000-dalton protein. This decrease is more rapid in the presence of (I)n . (br5C)n than in the presence of (I)n . (C)n. This differential decrease is also observed when 32P-labeled extracts are diluted into buffer containing 10 mM ATP, hexokinase and 1 and M glucose, or Escherichia coli alkaline phosphatase. A partial proteolytic digest of P1 labeled in the absence of double-stranded RNA or in the presence of (I)n . (C)n or (I)n . (br5C)n gives rise to similar peptide patterns. These results suggest that dephosphorylation as well as phosphorylation determines the net incorporation of 32P into P1. Moreover, these results suggest the existence of a phosphatase activity that may be inhibited more strongly by (I)n . (C)n than by (I)n . (br5C)n.
Significant structure theory applied to liquid
4
He
Authors: Ryoo Ryong , Jhon Mu Shik , Eyring Henry
Published: May 2006

The large kinetic zero-point motion of
4
He molecules has been properly taken into account in the significant structure theory of liquids in order to describe the liquidity of liquid
4
He. The Debye and the Bose-Einstein partition functions are used for the solid-like molecules of the normal fluid component and for the whole gas-like molecules, respectively. The solid-like molecules of the superfluid component are considered as ground state molecules without having positional degeneracies. The Bragg-Williams approximation of an order/disorder phase transition has been applied to the λ transition of liquid
4
He. Thermodynamic and surface properties of liquid
4
He have been calculated up to the critical point. The results are satisfactory when compared with the observed values.
Complete nucleotide sequence of a 23S ribosomal RNA gene from Escherichia coli.
Authors: Brosius J , Dull T J , Noller H F
Published: May 2006

The complete nucleotide sequence of the 23S RNA gene from the rrnB operon of Escherichia coli has been determined. The sequences of both strands of the entire gene were determined, most of the sequence was independently confirmed by use of alternate restriction fragments, and all restriction cuts overlapped. The DNA region corresponding to mature 23S rRNA contains 2904 nucleotides. Kethoxal-reactive sites protected by 30S subunits are found between positions 2300 and 28000, placing the subunit interface in this region of the molecule. The functional importance of this region is further supported by studies by other investigators, including homology with chloroplast and mitochondrial rRNA.
In vitro synthesis of the full-length complement of the negative-strand genome RNA of vesicular stomatitis virus.
Authors: Testa D , Chanda P K , Banerjee A K
Published: May 2006

Under the normal conditions of in vitro RNA synthesis, the virion-associated RNA polymerase of vesicular stomatitis virus synthesizes five monocristronic mRNAs and a 48-nucleotide-long leader RNA that represents the exact 3'-terminal region of the genome RNA [Colonno, R. J. & Banerjee, A. K. (1978) Cell, 15, 93-101]. When the transcribing core was preincubated with ATP and CTP, reisolated, and then incubated in the presence of the beta, gamma imido analogue of ATP (AdoPP[NH]P) and the three normal ribonucleoside triphosphates, the full-length complementary strand of the genome RNA was synthesized in vitro. The results suggest that specific phosphorylated states of regulatory proteins may control transcription in vitro to generate the full-length plus strands.
Carboxybiotin translocation mechanisms suggested by diffraction studies of biotin and its vitamers.
Authors: DeTitta G T , Parthasarathy R , Blessing R H , Stallings W
Published: May 2006

Biotin is a coenzyme that fixes CO2 for transfer in a family of carboxylase, decarboxylase, and transcarboxylase enzymes. Their enzyme reactions involve two basic steps during which a carboxybiotinyl intermediate forms at one site and translocates to a second (distinct) site for CO2 transfer. Our diffraction studies of biotin and its vitamers suggest that translocation involves rotation about one, or at most two, bonds in biotin's valeryl chain. The rotations are energetically economical gauche in equilibrium trans rotations about the two valeryl bonds nearest the biotin bicyclic ring. They move a carbon atom of a CO2 moiety bound at N-1' approximately 7 A, a distance in accord with spectroscopic measurements of one of the biotin enzymes. From our studies we infer that sulfur in biotin imparts to the valeryl chain a conformational variability necessary for bond rotation and, hence, translocation between catalytic sites.
Suspension culture of separated follicles consisting of differentiated thyroid epithelial cells
Authors: Nitsch Lucio , Wollman Seymour H.
Published: May 2006

We have prepared thyroid follicles in suspension culture to use as a model system
in vitro
for investigation of some properties of the thyroid gland. The follicles were free of endothelial cells, fibroblasts, and other nonepithelial cells. They were prepared by collagenase treatment of minced rat thyroid glands followed by differential filtration of the suspension through nylon meshes. Small clusters of principal thyroid epithelial cells were separated from large fragments and single cells. They were cultured in dilute suspension in Coon's modified F-12 medium in dishes coated with agarose to avoid having the cells attach to the dishes. By culture day 3, most of the clusters formed closed follicles containing a periodic acid-Schiff-positive colloid but without a basal lamina. Follicle walls contained an occasional C cell. The epithelium resembled that in the thyroid of a recently hypophysectomized rat, with normal polarity and organelle complement normal with respect to position and abundance, with basally located lysosomes, no pseudopods, and no colloid droplets. The cells were responsive to thyroid-stimulating hormone (thyrotropin) and to dibutyryl cyclic AMP. Thyroid-stimulating hormone at 10 munits/ml resulted in apical migration of lysosomes and formation of pseudopods and colloid droplets within 30 min; longer exposure resulted in depletion of luminal colloid. The results indicate that the suspended follicles resemble follicles
in vivo
with respect to morphology and responsiveness to thyroid-stimulating hormone in the absence of other cell types.
“Sexual” behavior in parthenogenetic lizards (
Cnemidophorus
)
Authors: Crews David , Fitzgerald Kevin T.
Published: May 2006

All-female, parthenogenetic species afford a unique test of hypotheses regarding the nature and evolution of sexuality. Basic data on the behavior of parthenogens are lacking, however. We have discovered, from observations of captive
Cnemidophorus uniparens, C. velox
, and
C. tesselatus
, behavior patterns remarkably similar to the courtship and copulatory behavior of closely related sexual species. Briefly, in separately housed pairs, one lizard was repeatedly seen to mount and ride its cagemate and appose the cloacal regions. Dissection or palpation revealed that, in each instance, the courted animal was reproductively active, having ovaries containing large, preovulatory follicles, while the courting animal was either reproductively inactive or postovulatory, having ovaries containing only small, undeveloped follicles. These observations are significant for the questions they raise. For example, is this behavior a nonfunctional vestige of the species' ancestry, or is this behavior necessary for successful reproduction in the species (e.g., by priming reproductive neuroendocrine mechanisms as has been demonstrated in sexual species)?
The RAD52 gene is required for homothallic interconversion of mating types and spontaneous mitotic recombination in yeast.
Authors: Malone R E , Esposito R E
Published: May 2006

The rad52-1 mutation prevents homothallic mating type interconversion and reduces mitotic recombination in yeast. It has been previously reported that rad52-1 abolishes meiotic recombination. These data suggest either that a generalized recombination function(s) is required for mating type switching or that generalized recombination and specific homothallic functions are jointly controlled by the RAD52 gene. The rad52-1 mutation affects the interconversion of the two yeast mating types (a and alpha) differently, suggesting that the interconversion process is not equivalent for both mating types. This type of asymmetry is not predicted by current models of homothallic switching.
Regulation of simian virus 40 early transcription in vitro by a purified tumor antigen.
Authors: Rio D , Robbins A , Myers R , Tjian R
Published: May 2006

Cloned DNA templates were used to direct the transcription of early and late simian virus 40 (SV40) genes by a cell-free RNA-synthesizing system. Transcription by RNA polymerase II was sensitive to low levels of alpha-amanitin and completely dependent on exogenously added DNA template. RNA products of discrete lengths were efficiently synthesized when transcription was directed by DNA restriction fragments containing promoter sequences for either early or late genes of SV40. Addition of the D2 tumor antigen to the template DNA inhibited transcription originating from the SV40 early promoter. In contrast, the D2 protein had little or no effect on the transcription from SV40 or adenovirus 2 (Ad2) late promoter sequences. When a mixture of cloned DNA containing SV40 early promoter and Ad2 late promoter was used to direct RNA synthesis, the D2 protein specifically inhibited the synthesis of SV40 early genes but not that of Ad2 late sequences. The D2 DNA binding protein also had no effect on the transcription directed by SV40 mutant templates that contain an intact early promoter sequence but lack specific tumor-antigen binding sites. We have confirmed that, under the conditions of the transcription assay, the D2 protein binds and interacts specifically with its recognition sites on wild-type template DNAs but fails to bind to mutant or Ad2 DNA templates that lack sequences containing SV40 tumor-antigen binding sites. These findings provide evidence that a direct interaction between tumor antigen and its specific binding sites on DNA is the mechanism by which the SV40 A gene autoregulates its transcription.
beta-Endorphin: characteristics of binding sites in rabbit spinal cord.
Authors: Ferrara P , Li C H
Published: May 2006

The interaction of human beta-endorphin with binding sites in rabbit spinal cord has been characterized. The stereospecific high-affinity binding sites are concentrated in the dorsal half of the spinal cord. Scatchard analysis of the binding data shows heterogeneity of the binding sites that can be resolved into two populations with apparent dissociation constants of 3.0 (+/-2.0) X 10(-10) and 3.3 (+/- 0.5) X 10(-9) M. Sodium ions decrease the binding of human beta-endorphin to spinal cord to the same extent as found in rat brain. The ability of several opiates and opioid peptides to inhibit the binding of human beta-endorphin is also presented.
Variations in properties of virus released from morphologically different cell lines transformed in vitro by Friend leukemia virus.
Authors: Tsuei D , Pogo B G , Friend C
Published: May 2006

The properties of the virus synthesized by each of three morphologically different cell lines originating from DBA/2J fetal liver cells transformed by the anemic strain of Friend leukemia virus in vitro were analyzed. The cells of line G-1 are malignant in syngeneic DBA/2 mice, grow in suspension, and are erythroid in origin. Cells of lines G-2 and G-3 are adherent, are epithelial in appearance, and produce no tumors in DBA/2J mice. Higher reverse transcriptase activity was detected in the culture fluid of lines G-2 and G-3, although virus from G-1 cells was more leukemogenic. Differences were also found in the virion density and size of the viral genome. RNA from the virions produced by G-2 and G-3 cells sedimented at 75 S in a sucrose gradient; virion RNA from G-1 cells sedmiented at 60 S. However, when subjected to polyacrylamide gel electrophoresis, all three virus strains showed identical RNA subunits with an estimated molecular weight of 2.6 X 10(6). Analysis of virion proteins by slab gel electrophoresis showed differences in envelope protein (gp71) components but not in the major core protein (p30). The properties of these viruses are stable and remain unchanged after passage in 3T3 cells.
Purification of myosin light chain kinase from bovine cardiac muscle.
Authors: Wolf H , Hofmann F
Published: May 2006

Myosin light chain kinase was purified > 100,000-fold to apparent homogeneity with a yield of 10% from bovine cardiac muscle. Sodium dodecyl sulfate gels of the purified kinase showed one stained band corresponding to a Mr of 94,000. The enzyme was activated > 10-fold in the presence of Ca2+ (apparent Ka = 0.6-1.2 microM) and calmodulin (apparent Ka = 3-5 nM). The purified enzyme had a specific activity of 20-30 mumol of phosphate transferred per min per mg from ATP to cardiac myosin light chain 2. One mole of phosphate was incorporated per 94,000 g of the kinase in the presence of Ca2+ and calmodulin or of cyclic AMP-dependent protein kinase or of both additions. In addition to myosin light chain kinase, a calmodulin-binding protein of unknown function was purified from bovine cardiac muscle. This protein had a Mr of 85,000, was composed of two dissimilar subunits (Mr of 61,000 and 15,000), and competed with myosin light chain kinase for calmodulin. The protein appears to be closely related to the calmodulin-binding protein I purified from brain.
Altered enolase in aged Turbatrix aceti results from conformational changes in the enzyme.
Authors: Sharma H K , Rothstein M
Published: May 2006

Young- and old-type enolases (2-phospho-D-glycerate hydrolyase, EC 4.2.1.11) from the free-living nematode Turbatrix aceti can be unfolded in 1.25 M guanidine hydrochloride and subsequently refolded with essentially a quantitative recovery. After refolding, both enolases form an identical or near-identical third type of the enzyme as determined by spectral criteria, sensitivity to heat, immunotitration, and rate of inactivation by bacterial protease. By the same criteria, the refolded enolase is closer in conformation to the native old form of the enzyme than to the young form. The results prove that young and old enolases are conformational isomers and that an in vivo transformation from young to old enzyme takes place by conformational changes without covalent modification. The process may be related to the previously demonstrated slowing of enolase turnover in T. aceti. Errors in sequence cannot be involved in the age-related alteration of the enzyme.
Bacteriorhodopsin is an inside-out protein.
Authors: Engelman D M , Zaccai G
Published: May 2006

Neutron scattering is particulary useful when parts of a structure can be deuterated. From Halobacterium halobium we have obtained, by biosynthetic incorporation, purple membranes in which all of the valines or all of the phenylalanines are present in deuterated form. Difference Fourier techniques permit a general assessment of the distribution of valine and phenylalanine in projections of the purple membrane structure. These show that valine is distributed toward the periphery of a single bacteriorhodopsin molecule, whereas phenylalanine is distributed toward its center. We use the facts that the amino acid sequence is known and that much of it can be assigned to the alpha helices of the bacteriorhodopsin structure to interpret our results. Comparison of our maps with the distribution of valine and phenylalanine around alpha-helical perimetrs establishes the distribution of other amino acids and leads to the conclusion that the charged and polar groups of the bacteriorhodopsin molecule tend to lie at the molecular interior, away from contact with lipid, while the nonpolar surfaces are directed outward, making contact with the lipid regions. Thus, the protein is "inside-out" compared with the organization of soluble proteins.
Inhibiting the onset of hormone-induced desentiziation of viable thymocytes by N alpha-tosyl-L-lysine chloromethyl ketone.
Authors: Zick Y , Cesla R , Shaltiel S
Published: May 2006

N alpha-Toysl-L-lysine chloromethyl ketone (Tos-LysCH2Cl) was found to inhibit irreversibly the onset of the hormone-induced refractory state in intact thymocytes. When thymocytes (approximately 2 X 10(7) cells per ml) are treated with Tos-LysCH2Cl(10(-4) M, for 90 min at pH 7 and 37 degrees C) the cells retain their viability, including a full capacity to recognize and respond to hormonal stimuli, yet they selectively lose their ability to become desensitized to persistent triggering by a hormone, as reflected in the state of activation of intracellular cyclic AMP-dependent protein kinase (ATP:protein phosphotransferase, EC 2.7.1.37). Whereas upon hormonal stimulation of untreated cells the immediate rise in the state of activation of this enzyme (up to an activity ratio of > 0.85) is followed by an exponential decline to basal values within approximately 60 min, in TosLysCH2Cl-treated cells the hormone-triggered elevation in the state of activation of the enzyme is maintained for > 60 min. Evidence is presented to suggest that in thymocytes TosLysCH2Cl inhibits the regulatory process that normally uncouples the adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] system without interfering with previous or subsequent molecular events connected with the transfer of hormonal signals across the cell membrane. This technique allows, therefore, the preparation of viable thymocytes with a limited and distinct regulatory defect introduced by chemical (covalent) means. As such, it is most useful for studies aimed at the elucidation of the mechanism of cell desensitization and for further characterization and localization of key components responsible for cellular refractoriness.
Regulation of insulin receptors: evidence for involvement of an endocytotic internalization pathway.
Authors: Baldwin D , Prince M , Marshall S , Davies P , Olefsky J M
Published: May 2006

Cultured human fibroblasts degrade insulin by a receptor-mediated process. When intracellular hormone degradation is inhibited by chloroquine, 125I-labeled insulin internalizes and accumulates intracellularly. In contrast, cultured IM-9 lymphocytes do not degrade receptor-bound insulin or accumulate 125I-labeled insulin in the presence of chloroquine. Insulin-induced receptor loss occurs in both cell types, and chloroquine inhibits this process in fibroblasts but not in IM-9 lymphocytes. Transglutaminase is a membrane-associated enzyme thought to mediate the process of ligand-induced receptor aggregation and internalization; levels of this enzyme were high in fibroblasts but barely detectable in IM-9 lymphocytes. Furthermore, dansylcadaverine--a potent inhibitor of transglutaminase--blocked insulin-induced receptor loss in fibroblasts but was without effect in IM-9 lymphocytes. These results support the concept that insulin receptor regulation is mediated via an endocytotic internalization pathway in human fibroblasts and that the mechanisms of this process differ among cell types.
Switches in macromolecular synthesis during induction of competence for transformation of Streptococcus sanguis.
Authors: Raina J L , Ravin A W
Published: May 2006

The induction of synchronous development of competence for genetic transformation in Streptococcus sanguis, by either endogenous or exogenous competence factor (CF), is manifested in the transient synthesis of a new set of at least 10 polypeptides, ranging from 14,000 to 51,000 in molecular weight. Eight polypeptides (E14, E16, E24, E28, E32, E37, E44, E51) appear early, and two polypeptides (L34, L42) appear 5-10 min later. One of the newly synthesized early polypeptides, E16, is shown to be a component of the presynaptic complex containing single-stranded DNA that is produced in vivo upon uptake of native donor DNA. Concomitant with this induced synthesis of competence-specific polypeptides there is a net decrease in RNA and protein synthesis but no change in DNA synthesis; donor DNA-binding ability and transformability reach maxima during the phase of diminishing macromolecular synthesis. Subsequently, donor DNA-binding ability and transformability decay at disproportionate rates as cells return to the normal state of macromolecular synthesis within one generation. Coincident with the induction of competence, the synthesis of a new RNA transcript of high molecular weight appears to be induced which continues during the restricted phase of total cellular RNA synthesis.
Postreplication repair and the susceptibility of Chinese hamster cells to cytotoxic and mutagenic effects of alkylating agents.
Authors: Friedman J , Huberman E
Published: May 2006

A cell variant (VR-43) resistant to the cytotoxic effect of N-methyl-N'-nitro-N-nitrosoguanidine and N-methyl-N'-nitro-N-nitrosourea was isolated from Chinese hamster V79 cells for use in studies of the relationship among cell survival, mutagenesis, and DNA repair by alkylating agents. Resistance to ouabain or 6-thioguanine was used as the genetic marker. After treatment with N-methyl-N'-nitro-N-nitrosoguanidine, the VR-43 cells exhibited mutation frequencies that were lower, on a dose basis, than those of the wild-type V79 cells. However, when analyzed at equicytotoxic doses, the VR-43 cells were more mutable than the V79 cells. No difference in cell survival or mutagenicity could be observed after treatment with other mutagens such as N-ethyl-N'-nitro-N-nitrosoguanidine, ethyl methanesulfonate, or x-rays. Postreplication repair was analyzed by determination of the molecular weight of the newly synthesized DNA by alkaline sucrose gradients. After treatment with N-methyl-N'-nitro-N-nitrosoguanidine, the VR-43 cells exhibited an enhanced postreplication repair relative to the V79 cells. No such enhancement was found after N-ethyl-N'-nitro-N-nitrosoguanidine or ethyl methanesulfonate treatment. Based on these results we propose that, after treatment of these and presumably other mammalian cells with some methylating mutagens, postreplication repair can cope with DNA lesions responsible for cytotoxicity and, to a lesser degree, with lesions responsible for mutagenicity.
A human soluble suppressor factor affecting lymphocyte responses in vitro.
Authors: Shou L , Schwartz S A , Good R A , Peng R , Chen C L
Published: May 2006

A soluble suppressor factor (SSF) has been demonstrated in the supernatant of normal human peripheral blood lymphocyte cultures that exhibits suppressive activity toward the proliferative response of normal lymphocytes to concanavalin A or alloantigens in mixed lymphocyte culture (MLC) or toward pokeweed mitogen-stimulated immunoglobulin synthesis and secretion in vitro. Suppression of the proliferative response in MLC reached maximal levels when added SSF-containing supernatant approximated 20% by volume of the culture medium. Suppression in the MLC was found to act at the proliferative stage. SSF acts independently of cytotoxicity and is stable at 56 degrees C for 30 min but is inactivated at higher temperatures. Addition of SSF to the MLC as late as day 4 after initiation of the culture results in suppression of transformation. This factor(s) may regulate the magnitude of several immune responses in humans.
Immunosuppression by human plasma alpha 1-acid glycoprotein: importance of the carbohydrate moiety.
Authors: Bennett M , Schmid K
Published: May 2006

Human plasma alpha 1-acid glycoprotein (orosomucoid) and derivatives of this protein produced by sequential enzymatic cleavage of the glycosidic residue (sialic acid, galactose, N-acetylglucosamine, mannose) were tested for the ability suppress a number of immune functions of mouse spleen cells in vitro at physiological concentrations. alpha 1-Acid glycoprotein and especially the agalacto/asialo derivative suppressed (i) the mitogenic responses to concanavalin A, lipopolysaccharide, and alloantigens, (ii) antibody responses to sheep erythrocytes, and (iii) induction of cell-mediated lympholysis against allogeneic target cells. The native protein and its derivative did not inhibit the proliferation of EL-4 lymphoma cells and did not suppress the lysis of YAC-1 lymphoma cells by natural killer cells. The enhanced potency of the agalacto/asialo derivative indicates that the nature of the carbohydrate moiety exposed on the protein determines the effectiveness of the glycoprotein and its deglycosylated derivatives may function to regulate immune responses in various physiological and pathological conditions.
Insulin-dependent diabetes mellitus induced by subdiabetogenic doses of streptozotocin: obligatory role of cell-mediated autoimmune processes.
Authors: Paik S G , Fleischer N , Shin S I
Published: May 2006

The role of thymic functions in the development of insulin-dependent diabetes was investigated in athymic nude (nu/nu) mice and euthymic heterozygous (+/nu) littermates of BALB/c origin treated with streptozotocin. The injection of a single high dose of streptozotocin (200 mg/kg body weight) induced rapid and permanent hyperglycemia in both nu/nu and +/nu mice. In contrast, the injection of the same total dose divided into multiple "subdiabetogenic" doses (40 mg/kg per day for 5 consecutive days) caused the development of delayed but progressive hyperglycemia only in the thymus-competent +/nu mice. Female mice of either genotype were significantly less susceptible to streptozotocin at both doses. Restoration of thymic immunity in nu/nu mice by thymus grafts also restored the susceptibility to the hyperglycemic effects of multiple low doses of streptozotocin. Moreover, splenic lymphocytes from +/nu mice previously made diabetic with the multiple low-dose injections of streptozotocin induced transient glucose intolerance in nu/nu mice. The ability of the diabetic spleen cells to transfer the diabetic state was abolished when the splenic lymphocytes were depleted of the T cells but not when they were depleted of B cells. These results provide direct proof that thymus-dependent functions play an obligatory etiologic role in the development of diabetes in mice treated with repeated subdiabetogenic doses of streptozotocin. These observations also add to the growing evidence that autoimmune amplification mechanisms may be critically involved in the etiology of juvenile-onset diabetes mellitus in humans.
Erythrocyte choline concentrations are elevated in manic patients.
Authors: Jope R S , Jenden D J , Ehrlich B E , Diamond J M , Gosenfeld L F
Published: May 2006

The intracellular choline concentration in erythrocytes of manic patients who never had lithium therapy was 3 times the value for normal human subjects. Lithium therapy increased this value to 11 times normal, and the increase began within the first 2 days of lithium therapy. Plasma choline concentration was normal in untreated manic patients and did not change during lithium therapy.
Characterization of Y73, an avian sarcoma virus: a unique transforming gene and its product, a phosphopolyprotein with protein kinase activity.
Authors: Kawai S , Yoshida M , Segawa K , Sugiyama H , Ishizaki R , Toyoshima K
Published: May 2006

The Y73 strain of avian sarcoma virus recently isolated in Japan is defective in replication and is associated with subgroup A leukosis virus (YAV). The virus caused sarcoma but not acute leukosis when inoculated into chickens. Studies on the viral RNA showed that a 26S RNA, etimated to be 4.8 kilobases long, was Y73 viral RNA carrying a transforming gene. The 26S RNA has sequences in common with the RNA of an avian leukosis virus but no homology with the src gene sequence of avian sarcoma virus (ASV). Thus, Y73 has a unique sarcoma-inducing gene. A phosphorylated polyprotein of 90,000 daltons (p90) was immunoprecipitated from extracts of Y73-transformed chicken embryo cells by a variety of antisera reacting with gag gene products. When a bacteria-bound immunocomplex containing the p90 protein was incubated with [gamma-32P]ATP, the Y73-specific p90 and the IgG heavy chain were phosphorylated by a p90-associated protein kinase. The amino acid phosphorylated in vitro was exclusively tyrosine in both cases, whereas p90 phosphorylated in vivo contained phosphoserine as a major phospho amino acid with traces of phosphotyrosine and phosphothreoine.
Specific labeling of the lac carrier protein in membrane vesicles of Escherichia coli by a photoaffinity reagent.
Authors: Kaczorowski G J , LeBlanc G , Kaback H R
Published: May 2006

4-Nitrophenyl-alpha-D-galactopyranoside (NPG) was used as a photoaffinity reagent to specifically inactivate the beta-galactoside transport system in Escherichia coli ML 308-225 membrane vesicles. Photolysis of NPG produced time-dependent, irreversible loss of transport activity with corresponding incorporation of [3H]NPG into the membrane. Both processes were blocked by beta-D-galactopyranosyl 1-thio-beta-D-galactopyranoside, a high-affinity substrate of the lac carrier protein and inactivation of lactose transport was specific because NPG photolysis did not affect proline uptake or the ability of the vesicles to generate an electrochemical proton gradient. Arylation of the lac carrier protein was stoichiometric and resulted in the formation of 0.25 nmol of NPG adduct per mg of membrane protein. All attempts to regenerate transport activity by reillumination in the presence of externally added nucleophiles failed, indicating that arylation is functionally irreversible. When vesicles labeled with [3H]NPG under defined experimental conditions were solubilized and analyzed by gel electrophoresis, only one radioactive peak with an apparent molecular weight of 30,000 was observed, confirming that the reaction is highly specific. The results demonstrate that NPG is an active-site-directed photoaffinity label for the lac carrier protein.
A nondenaturing zwitterionic detergent for membrane biochemistry: design and synthesis.
Authors: Hjelmeland L M
Published: May 2006

The synthesis and evaluation of a new detergent that is a zwitterionic derivative of cholic acid is presented. This detergent combines the useful properties of both the sulfobetaine-type detergents and the bile salt anions. The new detergent proved to be effective at solubilizing membrane proteins in a nondenatured state.
In vivo regulation of rat liver 3-hydroxy-3-methylglutaryl-coenzyme A reductase: enzyme phosphorylation as an early regulatory response after intragastric administration of mevalonolactone.
Authors: Arebalo R E , Hardgrave J E , Noland B J , Scallen T J
Published: May 2006

Although substantial evidence supports the conclusion that 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase [mevalonate:NADP+ oxidoreductase (CoA-acylating), EC 1.1.1.34] is the major regulatory enzyme in cholesterol biosynthesis, the molecular events involved in the in vivo regulation of this enzyme have remained obscure. To study this problem, rats were given a single 100-mg dose of mevalonolactone by intragastric tube. The rats were sacrificed 20 or 60 min later, and liver microsomes were prepared by ultracentrifugation. Two phases of inhibition of microsomal HMG-CoA reductase were observed. The first phase of inhibition, observed 20 min after mevalonolactone administration, was completely reversed by preincubation of the microsomes with purified phosphoprotein phosphatase. The second phase of inhibition, observed 60 min after mevalonolactone administration, was not reversed by phosphoprotein phosphatase. The reactivation of liver microsomal HMG-CoA reductase by phosphoprotein phosphatase was blocked by potassium fluoride or by phosphoprotein phosphatase inhibitor. Results obtained by immunotitration also showed that microsomal HMG-CoA reductase obtained from animals killed 20 min after mevalonolactone administration was significantly activated by phosphoprotein phosphatase treatment of the microsomes. These findings demonstrate that phosphorylation of rat liver HMG-CoA reductase is an early in vivo regulatory response after intragastric administration of mevalonolactone.
Increased levels of (2'-5')oligo(A) polymerase activity in human lymphoblastoid cells treated with glucocorticoids.
Authors: Krishnan I , Baglioni C
Published: May 2006

An enzymatic activity that synthesizes (2'-5')-oligo(A) from ATP is induced in animal cells treated with interferon. This activity, designated (2'-5')A polymerase, is also elevated in human lymphoblastoid Daudi and Raji cells treated with hydrocortisone. The polymerase activity increases significantly after 24 hr of treatment and declines when hydrocortisone is removed from the culture medium. The product of the enzyme prepared from hydrocortisone-treated cells is indistinguishable from (2'-5')oligo(A) synthesized with polymerase of interferon-treated cells either by an endonuclease activation assay or by chromatographic analysis. The increase in (2'-5')A polymerase is not mediated by secretion of interferon by hydrocortisone-treated cells; less than 1 unit of interferon per ml is present in the culture medium during treatment with this glucocorticoid hormone. Moreover, this increase is related to the concentration of hydrocortisone in the culture medium and is inhibited by the addition of cortexolone. This steroid interferes with the interaction between glucocorticoid hormones and their receptor. Cortexolone has no effect, however, on the induction of (2'-5')A polymerase by interferon. The synthetic glucocorticoid dexamethasone also increases the polymerase activity. Experiments with inhibitors show that such an increase requires RNA and protein synthesis.
Identification of two Escherichia coli factor Y effector sites near the origins of replication of the plasmids (ColE1 and pBR322.
Authors: Zipursky S L , Marians K J
Published: May 2006

The Escherichia coli replication factor Y has been characterized as a phi X174 (+) strand specific DNA-dependent phosphohydrolase. In conjunction with other E. coli replication proteins, factor Y is involved in the formation of heterogeneous primers that are elongated by the E. coli DNA polymerase III elongation machinery. We report here that the heat-denatured DNAs of plasmids pBR322 and ColE1 serve as effectors for the hydrolysis of ATP by factor Y. The DNA sequences of pBR322 responsible for factor Y effector activity have been localized. Two separate regions of the pBR322 chromosome support Y ATPase activity. These sequences are near the replication origin and are located on opposite DNA strands.
Exchange kinetics of the Schiff base proton in bacteriorhodopsin.
Authors: Ehrenberg B , Lewis A , Porta T K , Nagle J F , Stoeckenius W
Published: May 2006

Using rapid mixing techniques and resonance Raman spectroscopy, we have found that the 1H/2H exchange time for the Schiff base proton of bacteriorhodopsin in purple membrane is 4.7 msec, when experiments are carried out at pH 2 or pH 7 at room temperature in the dark. We argue that diffusion of neutral water into the membrane is fast on this time scale. Also, model Schiff bases in solution have a pKa between 6 and 7, and we show that such model Schiff bases have much faster exchange rates. Therefore, we conclude that the Schiff base proton in bacteriorhodopsin is protected from interaction with the medium, probably by interaction with a protein group, and this would account for a pKa considerably higher than 6-7.
Iron-sulfur proteins: spin-coupling model for three-iron clusters.
Authors: Kent T A , Huynh B H , Münck E
Published: May 2006

Recent Mössbauer and EPR studies of two ferredoxins and of aconitase have given evidence for a three-iron cluster, probably of a [3Fe-3S] type. The studies of the oxidized EPR-active centers have shown that the three iron sites are characterized by significantly different magnetic hyperfine coupling constants. For the ferredoxin from Azotobacter vinelandii, for instance, we have observed A1 = -41 MHz, A2 = +18 MHz, and [A3] = 5 MHz. We demonstrate here that the magnetic properties of the clusters can be explained with a simple model of three high-spin ferric ions (S = 5/2) exchange-coupled to a system spin S = 1/2. The model assumes isotropic exchange and different couplings between the iron sites. The results show that the three sites have intrinsic hyperfine interactions similar to those of ferric rubredoxin; the differences in the observed interactions reflect the geometrical features of spin coupling. Furthermore, the three exchange coupling constants are equal within a factor of 2. This implies that the three-iron cluster is a single covalently linked structure and should not be considered as a [2Fe-2S] cluster weakly coupled to a third iron atom.
Active proliferation of Rous sarcoma virus-infected, but not normal, chicken heart mesenchymal cells in culture medium of physiological composition.
Authors: Balk S D
Published: May 2006

Normal as well as Rous sarcoma virus-infected chicken pectoral and chicken embryo fibroblasts proliferate actively in a plasma containing medium of physiological ion concentrations (Ca2+, 1.2 mM; Mg2+, 0.7 mM). Reduction of medium calcium and magnesium concentrations is necessary to achieve selective quiescence of normal fibroblasts in these cell systems. By contrast, normal chicken heart mesenchymal cells proliferate only sluggishly (one doubling or less during a 6-day period) in a plasma containing medium of physiologic ion concentrations, whereas Rous sarcoma virus-infected heart mesenchymal cells proliferate actively (more than four doublings during an initial 2-day phase of exponential growth). The chicken heart mesenchymal cell system therefore has great potential for studies of the mechanism that initiates cell replication and of the failure in cellular regulatory processes that is responsible for the autonomous initiation of replication of neoplastic cells. From comparison of the chicken heart mesenchymal cell system to dialyzed plasma-based systems in which 3T3 cells tend to proliferative quiescence, it is argued that this proliferative quiescence of 3T3 cells is a result of cell starvation and is not physiologically meaningful.