bluuebunny/crossref_metadata_2025 huggingface dataset:
DNA gyrase on the bacterial chromosome: possibility of two levels of action.
Authors: Drlica K , Engle E C , Manes S H
Published: May 2006

In previous studies we have shown that oxolinic acid, a specific inhibitor of the A subunit of DNA gyrase, induces DNA cleavage at 100,000-base-pair intervals on the Escherichia coli chromosome. At subsaturating drug concentrations, cleavage is induced at a fraction of these sites and DNA synthesis is partially inhibited. This partial inhibition is surprisingly rapid even when few sites have been inactivated. We now report kinetic measurements suggesting that inactivation of 100,000-base-pair gyrase sites by oxolinic acid does not inhibit DNA synthesis by simply producing barriers to replication fork movement. Slowing the rate of fork movement, thus increasing the time for a fork to reach a barrier, fails to proportionately slow inhibition of DNA synthesis. Moreover, the initial, rapid phase of inhibition is followed by a slower decline that is not accelerated by increasing the frequency of barriers by raising drug concentrations. These data, when added to the observation that additional oxolinic acid-induced cleavage occurs in replicating regions of the chromosome, suggest that gyrase may function at replication forks as well as at 100,000-base-pair intervals on the chromosome.
Schwann cells proliferate but fail to differentiate in defined medium.
Authors: Moya F , Bunge M B , Bunge R P
Published: May 2006

Primary cultures of dorsal root ganglia cells from 18- to 21-day rodent embryos were studied for their ability to express Schwann cell function in a defined medium lacking serum and embryo extract. It was confirmed that Schwann cells, but not fibroblasts, are able to proliferate in response to contact with axons when cultured in this defined medium. We here report that in this medium, however, differentiation of Schwann cells was arrested before completion of ensheathment and before initiation of myelin formation. Electron microscopic analysis confirmed this ensheathment failure and showed that the extracellular matrix components (basal lamina and thin collagenous fibrils) normally produced by axon-related Schwann cells had not been formed. This absence of extracellular matrix, as well as the presence in the Schwann cell of an increased cytoplasmic granularity (observed in the light microscope) and numerous distended cisterns of rough endoplasmic reticulum, suggest a failure in Schwann cell secretion. However, within one week after addition of serum and embryo extract to the culture medium, the ensheathment failure was corrected and myelination occurred; electron microscopic observations showed the presence of basal lamina and collagen fibrils in association with Schwann cells. These results suggest the presence in serum or embryo extract (or both) of factors necessary for the full expression of Schwann cell function (although a similar requirement is not present for the expression of oligodendrocyte function in culture). We propose that these observations indicate a linkage between Schwann cell secretion and axonal ensheathment, including myelin formation.
Kinetic studies of the cooperative binding of subfragment 1 to regulated actin.
Authors: Trybus K M , Taylor E W
Published: May 2006

The transient-state kinetics of binding of myosin subfragment 1 (SF-1) to regulated actin in the presence and absence of Ca2+ were investigated. The binding of SF-1 to pure actin, to actin-tropomyosin (actin-TM), or to actin-tropomyosin-troponin (actin-TM-TN) in the presence of Ca2+ was kinetically the same. In each case, the light-scattering transients were biphasic, suggesting a two-step binding of SF-1 to actin. Binding of SF-1 to regulated actin in the absence of Ca2+ was different from binding in its presence and also varied depending on whether SF-1 or regulated actin was in excess. The kinetic results in the absence of CA2+ are explained by a cooperative binding model, in which the initial binding of SF-1 molecules to open (active) actin sites increases the number of open sites. TN-I labeled with the fluorophore 4-(N-iodoacetoxyethyl-N-methyl)-7-nitrobenz-2-oxa-1,3 diazole (TN*) was used to probe the state of the actin-TM-TN complex. Binding of SF-1 or CA2+ to regulated actin (in the absence of Ca2+) decreased the fluorescence of actin-TM-TN* by 30%, suggesting that binding of SF-1 or CA2+ induces a similar change in state. The change in fluorescence of TN* was also used to measure the rate of the transition from the active to the relaxed state in the absence of CA2+, which was 430 sec-1 at 4 degrees C in 0.1 M KCl. The lag prior to association of SF-1 with regulated actin (in the absence of Ca2+) was abolished when three SF-1 molecules were prebound per seven G-actin monomers. Similarly, a titration of actin-TM-TN* (in the absence of Ca2+) with SF-1 or SF-1-ADP showed that most actin sites are open, as measured by the fluorescence change, when the occupancy of actin-TM-TN* by SF-1-ADP or SF-1 is approximately 50%. The evidence shows that partial occupancy of a block of G-actin sites (possibly seven) by SF-1 or SF-1-ADP stabilizes the open (active) conformation.
Probabilistic view of the transformation of cultured C3H/10T1/2 mouse embryo fibroblasts by 3-methylcholanthrene.
Authors: Fernandez A , Mondal S , Heidelberger C
Published: May 2006

When C3H/10T1/2 cells are treated with a given concentration of a chemical carcinogen, the transformation frequency can vary over 4 orders of magnitude, depending primarily upon the number of cells plated. To explain this phenomenon, we have developed a probabilistic theory of the formation of transformed foci in this system. We define p1 as the probability that a cell will be activated by carcinogen treatment, p2 as the probability per cell generation that an activated cell will be transformed, and p3 as the probability per cell generation that an activated cell will be deactivated. The equation we have derived: log (F/N) = log [2p1p2(1 -- p3)/2(1 -- p3) -- 1]+ n log (1 -- p3) describes focus formation; F is mean number of loci per dish after carcinogen treatment, N is number of cells in a dish at confluence, and n is number of cell generations to confluence. This equation has been verified experimentally; p3 = 0.24 and p1p2 = 3.8 X 10(-6) at a single concentration of 3-methylcholanthrene. This relationship explains previously inexplicable effects of cell density on transformation frequency.
Fibronectin is not present in the focal adhesions formed between normal cultured fibroblasts and their substrata.
Authors: Chen W T , Singer S J
Published: May 2006

Fibronectin is an extracellular matrix protein that has been implicated in the spreading and adhesion of cultured fibroblasts to their substrata. In this paper, double immunoelectron microscopic labeling experiments for fibronectin and for concanavalin A-binding proteins on the cell surface were carried out on ultrathin frozen sections of cultures of embryonic chicken heart fibroblasts. On cross sections though the focal adhesions of the cell to the substratum there was substantial labeling for concanavalin A-binding proteins but no detectable labeling for fibronectin, whereas both the binding proteins and fibronectin were extensively labeled elsewhere on the cell surface and substratum. These results demonstrate that fibronectin is not present within the sites of focal adhesions. Therefore, the functions of fibronectin in cell spreading and adhesion are not directly mediated through its binding at focal adhesion sites. An alternative model is presented which can account for such fibronectin functions.
In vivo germinal mutation detection with "monospecific" antibody against lactate dehydrogenase-X.
Authors: Ansari A A , Baig M A , Malling H V
Published: May 2006

This report describes developments toward a cell specific-locus test for measuring point mutations directly in sperm based upon the use of a monospecific antibody against sperm-specific lactate dehydrogenase-X (LDH-X). The antibody recognizes amino acid differences between mouse and rat LDH-X molecules. In general, mouse sperm do not bind the monospecific antibody against rat LDH-X, but a few exceptional mouse sperm do. Such mouse sperm are believed to contain LDH-X molecules in which an amino acid normally present in mouse LDH-X has been exchanged with one present in rat LDH-X at a place where rat and mouse enzymes are antigenically different. Thus, in the fluorescent antibody technique, mouse sperm carrying normal LDH-X do not stain but the sperm containing mutated enzyme do stain. By using this technique, the spontaneous presumptive mutation frequency in DBA/2 mice was found to be 0.43 X 10(-6); it increased to 7.6 X 10(-6) upon procarbazine treatment of the mice.
Density-dependent selection in a random environment: An evolutionary process that can maintain stable population dynamics
Authors: Turelli Michael , Petry Doug
Published: May 2006

A theoretical analysis of natural selection is presented in which fitnesses depend on population density and randomly varying environmental processes. The theory is based on a general, heuristic analysis of a pair of coupled, nonlinear, stochastic difference equations that describe the joint dynamics of allele frequencies and population size. Four main conclusions emerge from the investigation of a particular class of models: (
i
) growth rates at low population densities tend to increase; (
ii
) individual selection, given sufficient genetic flexibility, will mold growth rates at higher densities so that in spite of
i
, stable deterministic population dynamics are maintained; (
iii
) “more fit” genotypes cannot be simply characterized—in particular, the mean population size need not be increased; and (
iv
) genetic polymorphisms can be maintained in both haploid and diploid organisms.
Cellular and subcellular localization of 1,25-(OH)2-vitamin D3 in rat kidney: comparison with localization of parathyroid hormone and estradiol.
Authors: Stumpf W E , Sar M , Narbaitz R , Reid F A , DeLuca H F , Tanaka Y
Published: May 2006

Results of thaw-mount autoradiographic studies of kidney obtained 1--3 hr after a single injection of 1,25-(OH)2-[3H]vitamin D3 to rats showed strong nuclear concentration of radioactivity in podocytes and cells of distal tubules, including the pars convoluta, macula densa, and pars recta. Intermediate nuclear concentration was found in cells of the thin limb of Henle's loops, whereas weak or no nuclear uptake was seen in cells of the proximal tubules. In contrast, after injection of 3H-labeled parathyroid hormone or [3H]estradiol, mainly cells of the proximal tubules were labeled; after 3H-labeled parathyroid hormone, apical cytoplasmic uptake of radioactivity was seen most strongly in the proximal part of the proximal tubules, whereas, after [3H]estradiol, weak nuclear concentration was seen in cells throughout the proximal tubules. The results are consistent with the concept that synthesis of 1,25-(OH)2-vitamin D3 occurs in the proximal tubules under stimulation of parathyroid hormone or estradiol (or other hormones) and that the major site of action of 1,25-(OH)2-vitamin D3 is in the distal tubules.
Large and persistent electrical currents enter the transected lamprey spinal cord.
Authors: Borgens R B , Jaffe L F , Cohen M J
Published: May 2006

The electrical currents at the surface of the proximal portion of an isolated and transected lamprey spinal cord were measured with an extracellular vibrating probe. Soon after transection, currents of about 0.5 mA/cm2 enter the cut surface of the spinal cord. These currents fall to about a quarter of their initial value within an hour; within the next 2 days they gradually decline from about 100 microA/cm2 to about 4 microA/cm2; they then remain constant up to 6 days posttransection, when the measurements were ended. The pattern of current entry included substantial peaks opposite (and presumably into) the cut ends of giant axons. Response to changes in the ionic composition of the medium indicates that about half of the injury current consists of Na+, and that much of the rest may consist of Ca2+. The measured influx of ions, which adds up to several coulombs per cm2 in a few days, should radically alter the ionic composition of the terminal few millimeters of neural tissue. Thus it may be important in the degenerative and regenerative responses of neurons to axotomy.
Strongly nonlinear parabolic variational inequalities
Authors: Browder Felix E. , Brézis Haim
Published: May 2006

An existence and uniqueness result is established for a general class of variational inequalities for parabolic partial differential equations of the form ∂
u
/∂
t
+
A
(
u
) +
g
(
u
) =
f
with
g
nondecreasing but satisfying no growth condition. The proof is based upon a type of compactness result for solutions of variational inequalities that should find a variety of other applications.
Geometrical gauge theory of ghost and Goldstone fields and of ghost symmetries
Authors: Ne'eman Yuval , Thierry-Mieg Jean
Published: May 2006

We provide a geometrical identification of the ghost fields, essential to the renormalization procedure in the non-Abelian (Yang-Mills) case. These are some of the local components of a connection on a principal bundle. They multiply the differentials of coordinates spanning directions orthogonal to those of a given section, whereas the Yang-Mills potential multiplies the coordinates in the section itself. In the case of a supergroup, the ghosts become commutative for the odd directions, and represent Nambu-Goldstone fields. We apply the results to chiral “flavor”
SU
(3)
L
×
SU
(3)
R
and to
SU
(2/1). The latter reproduces a highly constrained Weinberg-Salam model.
Plasminogen activator of islets of Langerhans: modulation by glucose and correlation with insulin production.
Authors: Virji M A , Vassalli J D , Estensen R D , Reich E
Published: May 2006

Islets of Langerhans isolated from rat pancreas contain and secrete plasminogen activator (PA). Production of PA is increased up to 15-fold by culture in the presence of high concentrations of glucose, and the dose-response curves for the effect of glucose on secretion of PA and on insulin are superimposable. Alloxan, a diabetogenic agent that is selectively cytotoxic for beta cells, abolishes the PA response to glucose. Various agents and hormones that affect beta-cell function affect the secretion of PA in a manner parallel to their modulation of insulin secretion. These observations suggest that PA is produced by beta cells and that enzyme synthesis and secretion are regulated in concert with those of the hormone. The potential role of PA and of plasmin in the physiology of the islets is considered. In particular, because plasmin cleaves proinsulin to a product that is electrophoretically indistinguishable from insulin, it is suggested that the PA/plasmin system may play a part in the conversion of proinsulin to the active hormone.
Digitalis metabolism and human liver alcohol dehydrogenase.
Authors: Frey W A , Vallee B L
Published: May 2006

Human liver alcohol dehydrogenase (alcohol: NAD" oxidoreductase, EC 1.1.1.1) catalyzes the oxidation of the 3 beta-OH group of digitoxigenin, digoxigenin, and gitoxigenin to their 3-keto derivatives, which have been characterized by high performance liquid chromatography and mass spectrometry. These studies have identified human liver alcohol dehydrogenase as the unknown NAD(H)-dependent liver enzyme specific for the free hydroxyl group at C3 of the cardiac genins; this hydroxyl is the critical site of the genins' enzymatic oxidation and concomitant pharmacological inactivation in humans. Several kinetic approaches have demonstrated that ethanol and the pharmacologically active components of the digitalis glycosides are oxidized with closely similar kcat/Km values at the same site on human liver alcohol dehydrogenase, for which they compete. Human liver alcohol dehydrogenase thereby becomes an important biochemical link in the metabolism, pharmacology, and toxicology of ethanol and these glycosides, structurally unrelated agents that are both used widely. Both the competition of ethanol with these cardiac sterols and the narrow margin of safety in the therapeutic use of digitalis derivatives would seem to place at increased risk those individuals who receive digitalis and simultaneously consume large amounts of ethanol or whose alcohol dehydrogenase function is impaired.
Polyamine and differentiation: induction of ornithine decarboxylase by parathyroid hormone is a good marker of differentiated chondrocytes.
Authors: Takigawa M , Ishida H , Takano T , Suzuki F
Published: May 2006

The activity of ornithine decarboxylase (OD-Case:L-ornithine carboxy-lyase, EC 4.1.1.17) in rabbit costal chondrocytes in culture increased markedly after addition of parathyroid hormone (PTH), reaching a maximum 4 to 5 hr after PTH addition. The increase in ODCase activity was followed by increase in the intracellular concentrations of polyamines, especially putrescine, which increased in 6 hr to about 3-fold that of untreated cultures. The induction of ODCase by PTH was not observed in L, 3T3, HeLa, buffalo rat liver, or BHK cells. Retinyl acetate and retinoic acid both inhibited expression of the differentiated phenotype of chondrocytes by rabbit costal chondrocytes in culture within 3 days after their addition, as judged by morphological change and decrease in sulfate incorporation into glycosaminoglycans but did not inhibit cell proliferation. PTH could not induce an increase in ODCase in de-differentiated cells that had been pretreated with retinyl acetate or retinoic acid for 3 days. but 4 days after removal of the retinoids, these de-differentiated cells regained the ability to synthesize ODCase in response to PTH. These facts suggest that the induction of ODCase and the formation of putrescine by PTH are good markers of the differentiated phenotype of cultured chondrocytes.
Dissociation of vasoconstrictor and platelet aggregatory activities of thromboxane by carbocyclic thromboxane A2, a stable analog of thromboxane A2.
Authors: Lefer A M , Smith E F , Araki H , Smith J B , Aharony D , Claremon D A , Magolda R L , Nicolaou K C
Published: May 2006

Carbocyclic thromboxane A2 [2 beta (Z),3 alpha- (1E,3R*)-3-(3-hydroxy(1-octenyl)-bicyclo[3.1.1]hept-2-yl-5-heptenoic acid], a stable analog of thromboxane A2, has been tested for its physiologic properties. Carbocyclic thromboxane A2 is a potent coronary vasoconstrictor, stimulating cornonary vascular smooth muscle at concentrations as low as 29 pM. At 1-5 micro M it is also an inhibitor of arachidonic-acid- and endoperoxide-induced aggregation of platelets. At 200 nM it stimulated the release of lysosomal hydrolases from large granule fractions of liver homogenate. It inhibited thromboxane synthesis in platelets, although it did not inhibit synthesis of prostacyclin in ram seminal vesicles. Thus, carbocyclic thromboxane A2, a molecule closely related to thromboxane A2, separates coronary vasoconstrictor from platelet-aggregating activity. The constrictor activity predominates in vivo; carbocyclic thromboxane A2 induces coronary vasoconstriction leading to myocardial ischemia and sudden death in rabbits in the absence of pulmonary or coronary thrombosis.
Regulation of early adenovirus transcription: A protein product of early region 2 specifically represses region 4 transcription
Authors: Nevins Joseph R. , Winkler Janet Jensen
Published: May 2006

An aspect of the regulation of early adenovirus gene expression has been studied by measuring the rates of transcription of several viral transcription units during infection by wild-type (WT) adenovirus type 5 and a temperature-sensitive mutant, H5ts125. It has previously been shown that two transcription units (regions 2 and 4) are subject to negative control. During the course of early infection, transcription of regions 2 and 4 increased to a maximal rate and then declined. The decline from each of these transcription units appeared to be a specific shutoff of transcription, because the transcription of another early transcription unit (region 1A) remained constant during early infection and, in the absence of protein synthesis, the apparent shutoff of region 2 and region 4 transcription did not occur. At the nonpermissive temperature for the mutant, the kinetics of transcription of early region 2 were identical to the kinetics of transcription of WT. Thus, the repression of transcription of region 2 occurred in H5ts125-infected cells as well as in WT-infected cells. Region 4 transcription, however, was not repressed during H5ts125 infection at the nonpermissive temperature. After a maximal rate of transcription was reached, this rate was maintained throughout the course of the experiment. Furthermore, the repression of region 4 transcription appears to be the result of a block of initiation of transcription rather than the result of an inducement of premature termination of transcription. Therefore, it can be concluded that a product of the adenovirus region 2 gene, which is the site of the mutation for H5ts125, is responsible for the specific shutoff of region 4 transcription.
Propagation of conformational changes in Ni(II)-substituted aspartate transcarbamoylase: Effect of active-site ligands on the regulatory chains
Authors: Johnson Ronald S. , Schachman H. K.
Published: May 2006

Although the importance of ligand-promoted conformational changes in allosteric enzymes has been recognized, it often has been difficult to determine whether the effects of binding are propagated to remote positions in different chains. Efforts were made, therefore, to demonstrate that changes due to ligand binding to the catalytic chains of aspartate transcarbamoylase (carbamoylphosphate:L-aspartate carbamoyltransferase, EC 2.1.3.2) of
Escherichia coli
are “communicated” to the regulatory chains. For these studies the endogenous zinc in the latter chains was replaced by nickel, which served as a discriminating spectral probe. The Ni(II)-enzyme was constructed by dissociating the native enzyme, separating the catalytic and regulatory subunits, removing Zn(II) from the latter, replacing it with Ni(II), and reconstituting the enzyme from native catalytic and Ni(II)-containing regulatory subunits. Ni(II) derivatives containing either six Ni(II) or five Ni(II) and one Zn(II) possess the allosteric properties of the native enzyme and exhibit absorption bands at 360 and 440 nm due to charge transfer transitions. Smaller bands were also observed at 665 and 720 nm from
d-d
transitions, which are consistent with tetrahedral geometry in the coordination sphere of nickel. Binding of the bisubstrate ligand
N
-(phosphonacetyl)-L-aspartate to the catalytic subunit of Ni(II)-aspartate transcarbamoylase perturbed the Ni(II) chromophore, giving rise to two difference spectral bands (at 390 and 465 nm). Spectral titrations showed that the conformational changes at the metal-ion-binding sites were complete even though about one-third of the active sites were unoccupied. This propagation of conformational changes is in accord with other evidence indicating that the allosteric transition in aspartate transcarbamoylase is concerted.
Net proton-hydroxyl permeability of large unilamellar liposomes measured by an acid-base titration technique.
Authors: Nichols J W , Deamer D W
Published: May 2006

The net proton-hydroxyl permeability of large unilamellar liposomes has been measured by an acid-base pulse titration technique and has been determined to be several orders of magnitude greater than that measured for other monovalent ions. This permeability is relatively insensitive to variations in lipid composition. Proton permeability and hydroxyl permeability vary with pH 6 to 8, and this variation can occur in the absence of alterations in surface charge density resulting from titrations of acidic and basic groups on the lipids. In order to account for the exceptionally high proton-hydroxyl permeability with respect to other monovalent ions, we propose that protons or hydroxyls or both interact with clusters of hydrogen-bonded water molecules in the lipid bilayer, such that they are transferred across the bilayer by rearrangement of hydrogen bonds in a manner similar to their transport in water and ice.
Cytogenetic effects of inhaled benzene in murine bone marrow: induction of sister chromatid exchanges, chromosomal aberrations, and cellular proliferation inhibition in DBA/2 mice.
Authors: Tice R R , Costa D L , Drew R T
Published: May 2006

Exposure of adult male and female DBA/2 mice to 3100 ppm benzene for 4 hr significantly increased the frequency of sister chromatid exchanges in bone marrow cells of both sexes, inhibited marrow cellular proliferation (but only in male mice), and did not significantly increase the frequency of chromosomal aberrations in either sex. Phenobarbital pretreatment synergistically interacted with benzene exposure to further increase sister chromatid exchanges in female mice, induce greater inhibition of cellular proliferation in male mice, and induce a significant level of chromatid-type chromosomal aberrations in both sexes. During the second day after exposure to benzene there was increased inhibition of cellular proliferation in male mice and both new DNA damage and persistance of old DNA damage in female mice. The differences in both the type and magnitude of the response of bone marrow cellular populations, as determined by different cytogenetic end points in male and female DBA/2 mice exposed to benzene or to phenobarbital and benzene, suggest not only that a metabolite of benzene is responsible for the observed effects, but that different metabolites may be involved in different end points.
Unreliability of tritiated cholesterol: studies with [1,2-3H]-cholesterol and [24,25-3H]cholesterol in humans.
Authors: Davidson N O , Ahrens E H , Bradlow H L , McNamara D J , Parker T S , Samuel P
Published: May 2006

Over the past 18 months different lots of [1,2-3H]cholesterol and [24,25-3H]cholesterol were found to be radiochemically acceptable by conventional chemical and other in vitro tests, yet, when co-administered with [4-14C]cholesterol to human subjects, an abrupt fall in the 3H/14C specific activity ratio in plasma cholesterol was discovered in every case. We have concluded that all batches of [3H]cholesterol should be regarded as radiochemically unreliable unless they are shown to behave identically in all respects to [4-14C]cholesterol in an appropriate in vivo assay system.
Activation, inactivation, and desensitization of acetylcholine receptor channel complex detected by binding of perhydrohistrionicotoxin
Authors: Eldefrawi Mohyee E. , Aronstam Robert S. , Bakry Nabil M. , Eldefrawi Amira T. , Albuquerque Edson X.
Published: May 2006

The effects of receptor activation were studied on the interaction of perhydrohistrionicotoxin (H
12
-HTX) with the ionic channel of the nicotinic acetylcholine (AcCho) receptor in membranes from the electric organ of
Torpedo ocellata
and with the endplate region of the soleus muscle of the rat. In
Torpedo
membranes, the initial rate (i.e., within 30 sec) of [
3
H]H
12
-HTX bindings to the ionic channel of the AcCho receptor was accelerated 10
2
- to 10
3
-fold in the presence of carbamoylcholine (Carb). H
12
-HTX also inhibited Carb-activated
22
Na
+
influx, over 95% inhibition at 10 μM H
12
-HTX. At this concentration H
12
-HTX did not inhibit [
3
H]AcCho binding to the AcCho-receptor sites. There was good correspondence between the degree of acceleration of [
3
H]H
12
-HTX binding and the stimulation of
22
Na
+
influx over a wide range of Carb concentrations (up to 100 μM). Preincubation of
Torpedo
membranes with Carb decreased the initial rate of [
3
H]H
12
-HTX binding, as well as the rate of
22
Na
+
influx, which may reflect desensitization of the AcCho-receptor.
d
-Tubocurarine inhibited the agonist-mediated acceleration of [
3
H]H
12
-HTX binding and
22
Na
+
influx. In the soleus muscle endplate, H
12
-HTX inhibited the transient depolarization induced by microiontophoretic application of AcCho; the more receptors activated and channels opened, the stronger was the inhibition by H
12
-HTX. These findings suggest that H
12
-HTX binds to closed and open ionic channels, with a preference for the latter conformation. It is also suggested that the conformational changes associated with activation or desensitization of the receptor can be monitored by studying binding of [
3
H]H
12
-HTX to the ionic channel sites as well as by the AcCho-receptor-regulated
22
Na
+
influx.
Very low energy cross sections for collision-induced rotational relaxation of I
2
seeded in a supersonic free jet
Authors: Tusa James , Sulkes Mark , Rice Stuart A.
Published: May 2006

We report a study of rotational relaxation of I
2
seeded in He, Ne, and Ar free jets. It is inferred that in the very low collision energy range covered by these experiments the cross section for rotational relaxation is independent of energy.
Multiple methylation in processing of sensory signals during bacterial chemotaxis.
Authors: DeFranco A L , Koshland D E
Published: May 2006

The multiple banding pattern on NaDodSO4/polyacrylamide electrophoresis of a subset of the signal transduction proteins involved in bacterial chemotaxis has been shown to be caused by multiple methylation of a single gene product. At least four methyl groups are introduced per molecule of the sensing membrane protein to give a total of five bands. The separation of the bands appears to correspond to a Mr difference of 1500, probably caused by the binding of different amounts of DodSO4 molecules as carboxyl groups are modified. Multiple methylation, similar to multiple phosphorylation in other systems, appears to offer advantages in control and sensory processing.
Heterogeneity of apolipoprotein B: isolation of a new species from human chylomicrons.
Authors: Kane J P , Hardman D A , Paulus H E
Published: May 2006

Low density lipoproteins and the triglyceride-rich lipoproteins of human serum each contain proteins of high molecular weight termed apolipoprotein B, which have previously been thought to be identical. We have isolated four species of apolipoprotein B with unique molecular weights and amino acid compositions. We have assigned numerical designations to these species in a centile system based upon their relative apparent Mr in NaDodSO4. One which we term B-100, with an apparent Mr of 549,000 +/- 7650 (SD) determined by NaDodSO4 gel electrophoresis, predominates in low density and very low density lipoproteins and is also present in chylomicrons from thoracic duct lymph or from plasma. Substantial amounts of two large proteins designated B-74 (apparent Mr 407,000 +/- 5790) and B-26 (apparent Mr 144,500 +/- 8970), which appear to be complementary fragments or constituents of the B-100 protein, are found in the low density lipoproteins of many individuals. A distinct protein, B-48, with an apparent Mr of 264,000 +/- 8150 is a major and constant constituent of chylomicrons from thoracic duct lymph or from plasma.
Phase equilibria for highly unsymmetrical plasmas and electrolytes
Authors: Pitzer Kenneth S.
Published: May 2006

The conclusion of classical Debye-Hückel theory that a phase separation may occur in highly unsymmetrical plasmas or electrolytes is shown to be false and to arise from a serious error in the treatment of the interaction of pairs of the most highly charged ions. After an approximate correction for this error, no phase separation is predicted. Specific application to iron in the solar plasmas is discussed.
Excretion of cysteine and gamma-glutamylcysteine moieties in human and experimental animal gamma-glutamyl transpeptidase deficiency.
Authors: Griffith O W , Meister A
Published: May 2006

Animals treated with potent gamma-glutamyl transpeptidase inhibitors and a patient with severe gamma-glutamyl transpeptidase deficiency excrete much larger than normal amounts of glutathione, gamma-glutamylcysteine, and cysteine in their urine; these compounds were found in disulfide forms. The findings indicate that the metabolic function of gamma-glutamyl transpeptidase is associated with the metabolism or transport (or both) of cysteine, gamma-glutamylcysteine, and glutathione, and that gamma-glutamylcysteine is a physiological substrate of the enzyme. The occurrence of gamma-glutamylcysteine in urine and other considerations suggest that this dipeptide is formed as an extracellular metabolite of glutathione in addition to its recognized role as an intrcellular precursor of glutathione. The dipeptide may be formed by a pathway involving transpeptidation or by cleavage of the Cys-Gly bond of glutathione. In the course of this work it was found that the mixed disulfide between glutathione and gamma-glutamylcysteine is a good substrate of glutathione reductase.
Lipid phase transition in planar bilayer membrane and its effect on carrier- and pore-mediated ion transport.
Authors: Boheim G , Hanke W , Eibl H
Published: May 2006

Using mixed-chain lipids, we have recorded cooling and heating curves of planar bilayer membranes while they passed the lipid phase transition range. With unmodified planar bilayers, spontaneous current fluctuations are observed near the lipid phase transition temperature (tc approximately 29 degrees C). This effect coincides with the expected and measured decrease in membrane capacitance. Carrier (valinomycin)-modified planar bilayers exhibit near tc an abrupt change from a high-conducting state above tc to the state of bare membrane conductance below tc. In contrast to this behavior, planar bilayers modified by pore-forming antibiotics (gramicidin A, alamethicin) do not show any peculiar effect at tc. However, at 22--23 degrees C a pronounced maximum in pore-induced conductance is seen. Whereas the gramicidin A pore abruptly stops stepwise fluctuations below approximately 16 degrees C, with alamethicin a few long-lasting pore and pore state fluctuations persist down to 10 degrees C. It is suggested that the carrier may freeze out into the membrane/water interface. The effects observed with pore-forming substances, on the other hand, are interpreted in terms of lateral phase separation into pure lipid and lipid/antibiotic domains.
Aggregation of luteinizing hormone receptors in granulosa cells: a possible mechanism of desensitization to the hormone.
Authors: Amsterdam A , Berkowitz A , Nimrod A , Kohen F
Published: May 2006

The temporal relationship between redistribution of receptors to lutropin (luteinizing hormone)/human chorionic gonadotropin in cultured rat ovarian granulosa cells and the cellular response to hormonal challenge were studied. Visualization of receptor-bound human chorionic gonadotropin by indirect immunofluorescence, with hormone-specific antibodies after fixation with 2% formaldehyde, revealed the existence of small clusters around the entire cell circumference 5--20 min after exposure to the hormone at 37 degrees C. Such small receptor aggregates were also evident if hormone incubation was at 4 degrees C or if cells were fixed with 2% formaldehyde before incubation. Larger clusters were evident after prolonged incubation with the hormone (2--4 hr) at 37 degrees C. The later change coincided with diminished cyclic AMP accumulation in respose to challenge with fresh hormone. When the fixation step was omitted and antibodies to human chorionic gonadotropin were applied after hormonal binding, acceleration of both receptor clustering and the desensitization process was observed. This maneuver also induced capping of the hormone receptors. In contrast, monovalent Fab' fragments of the antibodies were without effect. Internalization of the bound hormone in lysosomes, and subsequent degradation, was evident 8 hr after hormonal application and was not accelerated by the antibodies. It is suggested that clustering of the luteinizing hormone receptors may play a role in cellular responsiveness to the hormone. Massive aggregation of the receptors may desensitize the cell by interferring with coupling to adenylate cyclase.
Membrane differentiation in human erythroid cells: Unique profiles of cell surface glycoproteins expressed in erythroblasts
in vitro
from three ontogenic stages
Authors: Fukuda Minoru , Fukuda Michiko N. , Papayannopoulou Thalia , Hakomori Sen-itiroh
Published: May 2006

Human erythroblasts in culture, irrespective of the ontogenic stage of their progenitors, are characterized by: (
i
) the barely detectable amount of band 3 glycoprotein, (
ii
) the presence of two glycoproteins with molecular weights 105,000 and 95,000, (
iii
) the high concentration of glycophorin, and (
iv
) a minimum quantity of the carbohydrate chain susceptible to endo-β-galactosidase (“polylactosaminoglycan”). In contrast, mature erythrocytes, whether of fetal, neonatal, or adult origin, are characterized by a high concentration of band 3 glycoprotein, polylactosaminoglycan, and glycophorins, but do not contain 105- and 95-kilodalton-glycoproteins. Thus, the process of erythroid maturation from erythroblasts to erythrocytes is accompanied by the appearance of band 3, the disappearance of 105- and 95-kilodalton glycoproteins, and a great increase in the quantity of polylactosaminoglycan. The structure of polylactosaminoglycan may not be different between mature erythrocytes and erythroblasts from the same ontogenic stage, but it is distinctively different from one stage to the other. The profiles of oligosaccharides released by endo-β-galactosidase and immunofluorescence studies with anti-Ii antibodies indicated that a linear polylactosaminoglycan structure was present in erythroblasts as well as in erythrocytes of the fetal and newborn stage, whereas a branched polylactosaminoglycan structure was present in erythroblasts as well as erythrocytes of adult blood. Thus, two membrane characteristics are closely associated with the process of erythroid cell development; one—the membrane proteins band 3, band 4.5, and 95- and 105-kilodalton glycoproteins—determines the degree of maturation, and the other—polylactosaminoglycan—may determine the ontogenic stage of the erythroblast progenitors.
Adhesion plaques of Rous sarcoma virus-transformed cells contain the src gene product.
Authors: Rohrschneider L R
Published: May 2006

Another intracellular location of the Rous sarcoma virus (RSU) src gene product (pp60src) has been detected within RSV-transformed cells by indirect immunofluorescence. By using rabbit anti-tumor serum specific for pp60src, a speckled pattern of fluorescence was found on the ventral surface of RSV (Schmidt-Ruppin strain)-transformed normal rat kidney cells. Several tests indicated that this pattern was specific for pp60src. In addition, interference-reflection microscopy was used to visualize cellular adhesion plaques, which are the points at which cells attach to the substratum. Simultaneous immunofluorescence and interference-reflection microscopy indicated that the speckles of pp60src fluorescence corresponded exactly to the adhesion plaque structures. The presence of pp60src within the adhsion plaques was further demonstrated by indirect immunofluorescences on isolated adhesion plaques that remained bound to glass after removal of the cells. pp60src also was observed in adhesion plaques of RSV-tranformed chicken embryo fibroblasts (CEF) and mouse fibroblasts, as well as CEF infected with the temperature-sensitive RSV mutant tsNY68 and grown at permissive temperature. At nonpermissive temperature, pp60src was not detectable in adhesion plaques of the tsNY68-infected CEF. Adhesion plaques serve as focal points of microfilament bundle attachment, and thse results suggest that pp60src interacts directly with cellular cytoskeletal components.
Cultured mouse embryos metabolize benzo[a]pyrene during early gestation: genetic differences detectable by sister chromatid exchange.
Authors: Galloway S M , Perry P E , Meneses J , Nebert D W , Pedersen R A
Published: May 2006

Mouse embryos explanted at 7 1/2 or 8 1/2 days of gestation were cultured in medium containing benzo[a]pyrene and supplemented with 5-bromodeoxyuridine to allow detection of sister chromatid exchanges. The murine Ah locus regulates the inducible metabolism of polycyclic hydrocarbons such as benzo[a]pyrene. A high frequency of sister chromatid exchange was induced by benzo[a]pyrene in embryos from three Ah-"responsive" inbred strains (BALB/cDub, C3H/AnfCum, and C57BL/6N); there was little or no increase in two Ah-"nonresponsive" inbred strains (AKR/J and DBA/2J). Benzo[a]pyrene also induced sister chromatid exchanges in the Ah-responsive recombinant inbred line B6NXAKN-12 but not in the Ah-nonresponsive recombinant inbred line B6NXAKN-3. Sister chromatid exchange in cultured Ah-responsive mouse embryos was thus shown to be a sensitive assay. These data provide direct evidence that genetically responsive mouse embryos (early postimplantation stage) possess the subcellular processes necessary for induction of enzymes that metabolize benzo[a]pyrene to its chemically active forms(s). Both the Ah regulatory gene product (a cytoslic receptor) and the structural gene product (inducible cytochrome P1-450) therefore appear to be functional at an early embryonic age. Furthermore, this metabolic capacity may play an important role in the damage to embryonic cells by polycyclic hydracarbons.
Biparental products of bacterial protoplast fusion showing unequal parental chromosome expression.
Authors: Hotchkiss R D , Gabor M H
Published: May 2006

Efficiently regenerated single colonies from mixed multiply auxotrophic Bacillus subtilis protoplasts, fused with polyethylene glycol, reveal colonies carrying each of the parent types (biparentals) and recombinant colonies. The latter appear in high yields (up to 1% of certain recombinant classes); the yield of biparentals may be as large as 10%, in the range of the indicated physical fusion events. Many of the biparentals are diploids although, contrary to expectation, they are not complementing prototrophs, but show precisely the phenotype of one (either one) of the parent strains. Extensive pedigree analysis and subcloning of diploid lines show that they can propagate with varying stability on the appropriate parental selective medium to reproduce diploid progeny, parental segregants, and late-appearing recombinants, including some prototrophs. Thus, the principal product of intertype protoplast fusion is a diploid carrying two chromosomes, only one of which is expressed in each particular clone. The extinction of one parental genome is especially well demonstrated when it includes suppression of a normally dominant antibiotic sensitivity marker. In transformation experiments, DNA made from a selected diploid clone was able to transfer several of the unexpressed genes. The structural or topological character of DNA associated with the chromosome extinction remains unexplained.
Multiple opiate receptors: [3H]ethylketocyclazocine receptor binding and ketocyclazocine analgesia.
Authors: Pasternak G W
Published: May 2006

The receptor binding of the kappa agonist [3H]ethylketocyclazocine to brain homogenates in vitro and ketocyclazocine (kappa) analgesia in vivo has been investigated and compared to morphine, a mu agonist. Saturation analysis of [3H]ethylketocyclazocine binding in both mice and rats yielded biphasic Scatchard plots similar to those of opiate mu agonists, antagonists, enkephalins, and endorphins. Treatment of brain membranes with monovalent and divalent cation, chelating agents, protein-modifying reagents, and enzymes affected [3H]ethylketocyclazocine binding in a manner similar to that of [3H]morphine. Naloxazone, a long-acting antagonist that selectively abolished high-affinity [3H-DAla2,Met5]enkephalinamide binding in vivo, also selectively blocked high-affinity [3H]ethylketocyclazocine binding. Evaluation of analgesia with writhing and tail-flick assays in animals whose high-affinity binding sites were blocked by naloxazone demonstrated a 6- to 7-fold increase in median effective dose (ED50) values of ketocyclazocine. This decrease in analgesic potency was comparable to morphine's decreased potency in similarly treated mice. These biochemical and pharmacological results suggest that the analgesic properties of both kappa and mu agonists may be mediated through the same subpopulation of receptors, the high-affinity binding sites.
Comment on the rotational state dependence of indirect photodissociation of a polyatomic molecule
Authors: Novak Frank A. , Rice Stuart A.
Published: May 2006

The rotational state dependence of indirect photodissociation has been examined in the light of some recent developments in the theory of radiationless transitions in rotating molecules. A simple argument based on projection operators leads to the prediction of sequential kinetics from an initial singlet state through an intermediate triplet bound state and on to a final dissociative state. The results indicate that as long as the geometry of the molecule in the triplet state is not very different from that of the singlet state, only a very small rotational dependence will show up in the singlet state radiationless lifetime, in spite of the fact that the overall dissociation constitutes a large geometry change.
Structure of a cloned circular Moloney murine leukemia virus DNA molecule containing an inverted segment: implications for retrovirus integration.
Authors: Shoemaker C , Goff S , Gilboa E , Paskind M , Mitra S W , Baltimore D
Published: May 2006

Closed circular Moloney murine leukemia virus (M-MuLV) DNA was prepared from recently infected cells and cloned in a lambda vector. Four classes of cloned M-MuLV inserts were found: Class I, full length 8.8-kilobase (kb) inserts with two tandem long terminal repeats (LTRs) of 600 base pairs; class 2, 8.2-kb inserts with a single copy of a LTR; class 3, M-MuLV DNA inserts with various portions deleted; and class 4, an 8.8-kb insert with an internal sequence inversion. Determination of nucleotide sequence at the junction between the two LTRs from a class 1 insert suggested that circularization occurred by blunt-end ligation of an 8.8-kb linear DNA. The class 4 molecule had an inversion that was flanked by inverted LTRs, each of which had lost two terminal base pairs at the inversion end points. Also, four base pairs that were present only once in standard M-MuLV DNA were duplicated at either end of the inversion. This molecule was interpreted as resulting from an integrative inversion in which M-MuLV DNA has integrated into itself. Its analysis thus provided explicit information concerning the mechanism by which retrovirus DNA integrates into host cell DNA. Models of retrovirus integration based on bacterial DNA transposition mechanisms are proposed.
Both the 7-methyl and the 2'-O-methyl groups in the cap of mRNA strongly influence its ability to act as primer for influenza virus RNA transcription.
Authors: Bouloy M , Plotch S J , Krug R M
Published: May 2006

The ability of eukaryotic mRNAs to serve as primers for influenza virus RNA transcription depends on the presence of a 5'-terminal methylated can structure, the absence of which eliminates essentially all priming activity [Plotch, S. J., Bouloy, M. & Krug, R. M. (1979) Proc. Natl. Acad. Sci. USA 76, 1618-1622]. The present study was undertaken to determine the extent to which each of the methyl groups in the cap influences the priming activity of a mRNA. To assess the importance of the 2'-O-methyl group on the penultimate base of the cap, we used several plant viral RNAs containing the monomethylated cap 0 structure, m7GpppG. Brome mosaic virus (BMV) RNA 4 stimulated influenza virus RNA transcription only about 10-15% as effectively as did globin mRNA, which has a cap with a 2'-O-methyl group. When the cap of BMV RNA 4 was enzymatically 2'-O-methylated, its priming activity was increased 14-fold. Qualitatively similar results were obtained with other plant virus RNAs. To assess the importance of the terminal 7-methyl group, BMV RNA 4 containing the cap structure GpppGm was prepared by a series of chemical and enzymatic steps. These molecules were found to be only about 15% as active in priming as BMV RNA 4 molecules containing the fully methylated cap, m7GpppGm, indicating that the terminal 7-methyl group also strongly enhances priming activity. These results indicate that the cap 1 structure (m7GpppXm) found in all mammalian cellular mRNAs is more stringently required for priming influenza virus RNA transcription than for translation in cell-free systems.