bluuebunny/crossref_metadata_2025 huggingface dataset:
A Co-Evolution Theory of the Genetic Code
Authors: Wong J. Tze-Fei
Published: May 2006

The theory is proposed that the structure of the genetic code was determined by the sequence of evolutionary emergence of new amino acids within the primordial biochemical system.
Catecholamine-induced subsensitivity of adenylate cyclase associated with loss of beta-adrenergic receptor binding sites.
Authors: Mukherjee C , Caron M G , Lefkowitz R J
Published: May 2006

Injection of frogs with beta-adrenergic catecholamines for 1-24 hr produces marked subsensitivity of the erythrocyte membrane adenylate cyclase [ATP pyrophosphate-lyase (cyclizing); EC 4.6.1.1.] to in vitro stimulation by isoproterenol. The subsensitization is specific for catecholamine stimulation, since basal and fluoride-stimulated enzyme activity are unaffected. Maximum isoproterenol-stimulated adenylate cyclase activity declines by 75% in the isoproterenol-treated animals (P less than 0.001). The concentration of isoproterenol causing one-half maximal activation of adenylate cyclase, however, is unaltered. (-)[3H]Alprenolol, a potent competitive beta-adrenergic antagonist, was used to study directly the beta-adrenergic receptor binding sites in the erythrocyte membranes from control and subsensitized animals. A highly significant (P less than 0.005) 60% fall in the number of the beta-adrenergic receptor binding sites ("specific"(-)[3H]alprenolol binding sites) in the treated animals was found. The binding affinity of the sites was not markedly altered. These data suggest that beta-adrenergic catecholamines are able to regulate catecholamine sensitivity of tissues in vivo, by regulating the properties of the beta-adrenergic receptor binding sites.
Detection of new temperature-dependent conformational transition in lysozyme by carbon-13 nuclear magnetic resonance spectroscopy.
Authors: Cozzone P J , Opella S J , Jardetzky O , Berthou J , Jollès P
Published: May 2006

A specific temperature-dependent conformational transition of hen egg-white lysozyme, occurring between 20 degree C and 30 degree C in solution, has been detected by 13-C-nuclear magnetic resonance spectroscopy. Selective changes in the chemical shifts of aromatic residues, together with differences in the chemical shifts, and nuclear Overhauser enhancement in the carbonyl, carboxyl, and alpha-carbon regions of the spectrum point to the vicinity of subsites D and E as the primary locus of the structural change.
Beta-adrenergic stimulation of pineal N-acetyltransferase: adenosine 3':5'-cyclic monophosphate stimulates both RNA and protein synthesis.
Authors: Romero J A , Zatz M , Axelrod J
Published: May 2006

The lag period in the induction of rat pineal N-acetyltransferase (arylamine acetyltransferase or acetyl-CoA:arylamine N-acetyltransferase EC 2.3.1.5) by catecholamines via the beta-adrenergic receptor varies with the length of exposure of the rat to light or darkness. If rats have been exposed to light and reduced sympathetic nerve activity for more than 12 hr, this lag period is 1-2 hr long. Under these conditions, actinomycin D completely blocks the induction of N-acetyltransferase by isoproterenol and by dibutyryl adenosine 3':5'-cyclic monophosphate (cyclic AMP). In contrast, if enzyme activity is caused to fall by brief exposure to light at night when N-acetyltransferase activity is high, reinduction by catecholamines occurs almost immediately. In this case, actinomycin D does not block the reinduction of N-acetyltransferase by isoproterenol or by dibutyryl cyclic AMP. Cycloheximide blocks N-acetyltransferase induction under all conditions tested. Thus, new protein synthesis is always required for N-acetyltransferase induction; however, the requirement for RNA synthesis is variable, and contributes to the length of the lag period for induction. It is postulated that both beta-adrenergic stimulation and dibutyryl cyclic AMP act intracellularly at two separate sites in the induction of pineal N-acetyltransferase. One site is in the stimulation of transcription, and the other is in the stimulation of post-transcriptional events.
Tumor-associated transplantation antigens in immune rejection of mouse malignant cell hybrids.
Authors: Jami J , Ritz E
Published: May 2006

The inability of mouse cell hybrids derived from two malignant parental cells to produce tumors in syngeneic F1 hosts was analyzed. YC hybrids, derived from the fusion of C1.1D and L1210 clls, failed to induce any tumor in adult mice, while 91% of x-irradiated newborn mice developed tumors and died. Some telocentric chromosomes were lacking in hybrid tumors; however, none of the immunologically intact adult mice developed tumors when grafted with tumors grown in x-irradiated newborn mice. This indicates that histocompatibility factors interfered in the failure of YC tumors to grow to adult hosts. Syngeneic F1 mice immunized with Y2C hybrid cells had cytotoxic antibodies against Y2C hybrids and C1.1D and L1210 parental cells. Complete absorption of cytotoxic antibodies directed against hybrid cells by mixtures of both parental cell lines demonstrates the absence of any new antigen on hybrid cells.Hyb rid cells had a higher density of C1.1D-tumor antigenic sites, as compared to C1.1D parental cells.This possibly explains the higher antigenicity and/or the higher sensitivity to immune lysis of hybrid cells.
Changes in cyclic nucleotide metabolism in aorta and heart of neurogenically hypertensive rats: possible trigger mechanism of hypertension.
Authors: Amer M S , Doba N , Reis D J
Published: May 2006

Changes in cyclic nucleotide metabolism similar to those characteristic of the chronic forms of hypertension were observed in an acute neurogenic form of hypertension in rats produced by electrolytic lesions of the nucleus tractus solitarii. These changes that were evident 2 hr after the lesions were made included decreased cyclic AMP levels in the heart, increased cGMP:cAMP ratio, cAMP phosphodiesterase (3':5'-cAMP 5'-nucleotidohydrolase, EC 3.1.4.17) and guanylyl cyclase (GTP pyrophosphate-lyase (cyclizing), EC 4.6.1.2) activities in the aorta and decreased snesitivity of adenylyl cyclase (ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1) in both the aorta and heart to stimulation by the beta-adrenergic stimulant isoproterenol. These changes appear to depend on catecholamine release and are not due to mechanical distortion secondary to the increased arterial pressure. These studies provide biochemical support to the concept that the sympathetic nervous system may play a critical role in the initiation of the hypertensive syndrome and that chronic hypertension could result from the fixation of the biochemical effects of increased sympathetic activity.
Language boundaries and biological differentiation of Bougainville: multivariate analysis of variance.
Authors: Rhoads J G , Friedlaender J S
Published: May 2006

Blood genetic and antropometric data on Melanesians from Bougainville, Papus New Guinea, are analyzed by random-effects analysis of variance to partition the observed variation into components for the individuals, village, and language group level. Both clinial and unpatterned group differences exist. The differences between language groups appear to be substantial, even when the results are corrected for clinal effects. The amounts of variation of each level correspond roughly to a similar analysis of heterozygosity in blood polymorphisms. Observed current migration figures suggest that language and village constitute a 2-fold hierarchical subdivision of the breeding system, and this analysis shows probable random drift effects between groups at both the village and language level. Both language and genetic constitution of these villages are the result of differentiation in place.
Human globin gene analysis for a patient with beta-o/delta beta-thalassemia.
Authors: Ottolenghi S , Lanyon W G , Williamson R , Weatherall D J , Clegg J B , Pitcher C S
Published: May 2006

Complementary DNA (cDNA) was prepared with RNA-dependent DNA polymerase from human globin messenger RNA (mRNA). Annealing and translation experimenta with total mRNA from circulating cells from a patient with heterozygous beta/heterozygous beta-delta-o thalassemia (beta-o/delta beta-o-thalassemia) demonstrated no detectable mRNA for beta-globin. cDNA enriched in sequences homologous to beta-globin mRNA was prepared by hydroxylapatite fractionation of hybrids formed between beta-o/delta beta-o-thalassemic mRNA and cDNA made from mRNA from a patient with alpha-thalassemia (hemoglobin H disease). The rate of annealing of this beta-enriched cDNA to normal human nuclear DNA was that of a sequence present as only a single copy per haploid genome. The beta-enriched cDNA annealed to the beta-o-delta beta-o-thalassemia total DNA with approximately the same kinetics as to normal DNA, indicating that no total gene deletion of beta-globin genes from the diploid genome has occurred, although the accuracy of the technique could not exclude with certainty a partial deletion or a deletion of a beta-globin gene from only one of the haploid genomes. This demonstrates that at least one of the beta-o- or the delta beta-o-thalassemia haploid genomes in this case contains a substantially intact beta-globin gene.
Poly(adenylic acid) sequences in the RNA of Caulobacter crescenus.
Authors: Ohta N , Sanders M , Newton A
Published: May 2006

Poly(adenylic acid) sequences have been isolated from the Gram-negative bacterium Caulobacter crescentus. Most of these A-rich tracts are associated with large RNA molecules that constitute an important fraction of the unstable RNA in these bacteria, and, as estimated by poly(U) filter binding, they are not present in the 16S or 23S ribosomal RNA. Preliminary estimates of size from polyacrylamide gel electrophoresis suggest that the majority of the A-rich tracts ranges from 15 to approximately 50 residues in length.
A genetic determinant of the phenotypic variance of the molecular weight of low density lipoprotein.
Authors: Fisher W R , Hammond M G , Mengel M C , Warmke G L
Published: May 2006

The molecular weight of monodisperse human plasma low densitylipoprotein has been measured in 69 individuals and found to vary over the range of 2.4 to 3.9 X 10-6. By contrast, the molecular weight of low density lipoprotein measured on two separate occasions for specific individuals shows a mean difference of 0.07 X 10-6 and a standard deviation of 0.08 X 10-6; hence low density lipoprotein differing in molecular weight by greater than 0.2 X 10-6 may be considered different macomolecules. The distribution of the molecular weight of low density lipoprotein does not differ as a function of age or sex. Hyperlipemic subjects having monodisperse low density lipoprotein show similar molecular weight distribution to normal subjects, as do subjects with premature coronary artery disease. Family studies reveal a correlation coefficient of 0.82 between average molecular weights of parents and offspring, with significance at 0.01. In order to assess the influence of environment on molecular weight of low density lipoprotein, the correlation coefficient between the fathers' and mothers' low density lipoprotein was measured and no statistically significant correlation was found. These data are interpreted as strong evidence for a genetic determination of molecular weight of low density lipoprotein. A study of individuals in five families yields molecular weight data consistent with a single gene locus genetic mode of inheritance without dominance. The regression coefficient of the mean low denisty lipoprotein parental molecular weight on the offspring molecular weight is 0.30. If the variability of molecular weight is considered as an expression of phenotypic variance, then the regression analysis identified 30% of this phenotypic variance as arising from additive gene action presumably at a single locus. Segregation in the family data is consistent. Since the differences in molecular weight of low density lipoprotein arise from differences in the amount of lipid bound to the apoprotein, it is likely that an additional portion of the phenotypic variance of the molecular weight results from individual variations in the metabolism of low density lipoprotein, which yield differences in lipid content. The individual variation in molecular weight is only approximately 5%; hence those metabolic sequences that influence molecular weight of low density lipoproteins must be precisely controlled.
Circadian Rhythm of Neuron R15 of
Aplysia californica: In Vivo
Photoentrainment
Authors: Audesirk Gerald , Strumwasser Felix
Published: May 2006

(
1
) The neuron R15 in the parietovisceral ganglion of
Aplysia
has a circadian rhythm of spiking activity when recorded in the isolated ganglion. The rhythm is entrained
in vivo
by light-dark cycles. (
2
) The phase of the R15 rhythm is a function not only of the entraining light schedule, but also of the time of dissection. Changes in the dissection time during the light portion of the light-dark cycle yield little change in the subsequent R15 peak time. Dissections during the dark portion produce peak times that vary with dissection time with a slope that is approximately one. (
3
) The circadian rhythm of R15 can be phase-shifted
in vivo
by changes in the phase of the entraining light-dark cycle in one to two weeks. R15 neurons of blinded
Aplysia
, however, show little or no phase shift in this time. (
4
) It is concluded that the eyes are important as receptors for the photoentrainment of the R15 rhythm
in vivo
, but that neural connections from the eyes to R15 are not required.
Observational validation of the chronometric cosmology: I. Preliminaries and the redshift-magnitude relation
Authors: Segal I. E.
Published: May 2006

Results of comprehensive observational tests of the cosmological theory proposed by Segal in 1972 are summarized in this and succeeding notes. Within a framework of natural and general causality and symmetry considerations, the theory is mathematically essentially unique. It implies definite relations between directly observed quantities (redshift, apparent magnitude and diameter, number counts, and the like) which are entirely free of adjustable parameters, such as the
q
0
and Λ of general relativistic cosmologies. Rigorous statistical analysis of large (≳100) or systematically defined samples of galaxies or quasars accepts these relations at substantial probability levels. Note I establishes uniform statistical procedures and compares the redshift—magnitude relations of major samples with the theoretical relations of the present and expansion theories.
Well-posed inverse spectral problems
Authors: Hochstadt Harry
Published: May 2006

It is known that if complete spectral data are provided, the potential function in a Sturm-Liouville operator is uniquely determined almost everywhere. If two such operators have spectra that differ in a finite number of eigenvalues, then the corresponding potential functions will no longer be the same. However, as is demonstrated when the nonidentical eigenvalues are almost equal, then the corresponding potential functions will also be nearly equal almost everywhere. Furthermore, if an operator and its spectrum are given and the potential is presumably known and if a second operator is defined in such a way such that its eigenvalues agree with the eigenvalues of the first operator except for a finite set, then the potential corresponding to the second operator can be explicitly found by solving a set of nonlinear ordinary differential equations. Lastly, it is shown that the procedures discussed here and the Gelfand-Levitan procedures are significantly different, and in fact that the Gelfand-Levitan procedure is almost certainly not well posed.
Cholera toxin activation of adenylate cyclase in cancer cell membrane fragments.
Authors: Bitensky M W , Wheeler M A , Mehta H , Miki N
Published: May 2006

Activation of adenylate [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] by cholera toxin (84,000 daltons, 5.5 S) is demonstrated in plasma membrane fragments of mouse ascites cancer cells. The activation of adenylate cyclase is mediated by a macromolecular cyclase activating factor (MCAF), which has a sedimentation constant of 2.7 S and a molecular weight of about 26,000. MCAF is derived from, and may be identical to the "A fragment" of cholera toxin. Generation of MCAF depends on prior interaction of cholera toxin with either dithiothreitol, NADH, NAD, or a low-molecular-weight component (less than 700 daltons) present in cytoplasm. Subsequent exposure of this pretreated cholera toxin to cell membranes from a variety of mouse ascites cancer cells is followed rapidly by the appearance of MCAF, which no longer requires dithiothreitol, NADH, or NAD for the activation of adenylate cyclase. Activation of adenylate cyclase by MCAF in ascites cancer cell membrane fragments is not reversed by repeated washing of these membrane fragments. Adenylate cyclase in normal cell membrane fragments fails to respond either to cholera toxin or MCAF in the presence of dithiothreitol. In striking contrast, the adenylate cyclase in membrane fragments from five ascites cancer cells responds to either MCAF or native cholera toxin preincubated with dithiothreitol, NADH, or NAD.
Methylation of messenger RNA of Newcastle disease virus in vitro by a virion-associated enzyme.
Authors: Colonno R J , Stone H O
Published: May 2006

Purified Newcastle disease virus contains an enzyme that incorporates the methyl group from S-adenosyl-L-methionine into RNA synthesized in vitro by the virion-associated RNA polymerase (RNA nucleotidyltransferase). Incorporation of radioactivity from S-adenosyl-L-[methyl-3H]methionine was totally dependent upon RNA synthesis. The methylation reaction was completely inhibited by S-adenosyl-L-homocysteine, suggesting the transfer of only the methyl group of S-adenosyl-methionine to RNA products. Velocity sedimentation and hybridization of the in vitro product RNA indicated that both [3H]methyl and [32P]GMP labels resided in single-stranded 18S RNA molecules which were virus specific. Approximately 1 to 2 methyl groups were incorporated per RNA molecule. DEAE-cellulose chromatography of product RNA after alkaline hydrolysis suggested that the 5' terminus was the site of methylation.
Establishment of tight junctions between epithelial cells.
Authors: Hudspeth A J
Published: May 2006

Epithelia serve as barriers to the diffusion of solutes between body compartments, and must do so despite the frequent loss of cells. When single cells are experimentally removed from the Necturus gallbladder epithelium, contiguous cells migrate to fill the defect within 30 min. Electrophysiological measurements show that the local electrical resistance across the epithelium in the region of a wound returns to normal in the same period of time; electron microscopy demonstrates that tight junctions are formed concurrently. Physiologically functional and morphologically recognizable tight junctions can thus be established within 30 min, demonstrating a mechanism for the rapid restoration of epithelial integrity after cell loss.
Fetal mouse hearts: a model for studying ischemia.
Authors: Ingwall J S , DeLuca M , Sybers H D , Wildenthal K
Published: May 2006

A new experimental model for the study of two important aspects of ischemia, namely, oxygen and substrate deprivation, is proposed: the intact, beating fetal mouse heart in organ culture. This model offers long-term stability, ease and reproducibility of preparation, and the ability to manipulate experimental conditions. Hearts deprived of oxygen and glucose ceased beating immediately. After 3-4 hr of deprivation, biochemical and ultrastructural changes consistent with ischemic injury were evident. These include depletion of ATP and glycogen levels, loss of cytoplasmic enzymes, and extensive swelling and disruption of mitochondrial structure. Glucose and insulin partially protected against ATP depletion. Upon resupply of oxygen and glucose , beating resumed immediately, ATP levels rapidly increased to control levels and, consistent with this, mitochondrial structure returned toward normal. During the recovery phase autophagic vacuoles containing damaged mitochondria and myofibrils were seen, indicating that repair mechanisms were activated. Consistent with this, the proportion of lysosomal enzymes that were present in the nonsedimentable fraction of the tissue homogenate increased. We conclude that the cultured fetal mouse heart is a model useful for studying myocardial responses to anoxia and/or substrate deprivation and for assessing interventions designed to limit damage or to stimulate repair after ischemic injury.
Location of histones on simian virus 40 DNA.
Authors: Polisky B , McCarthy B
Published: May 2006

The physical location of histone molecules in a simian virus 40 DNA-histone complex isolated from purified virions was examined using site-specific restriction endonucleases. The complex contains four host histone species but lacks histone F1. Histones prevent complete cleavage of SV40 DNA by two restriction enzymes, HindIII and EcoRI. From the pattern of DNA fragments resulting from cleavage of the histone-DNA complex by the HindIII endonuclease, which makes six breaks on purified SV 40 DNA, we have concluded that histones are randomly arranged on SV40 DNA relative to restriction enzyme cleavage sites. The EcoRI endonuclease, which makes one break in SV40 NDA, was used to determine the degree of physical coverage of the SV 40 DNA molecule by histones. We observed that 80% of the EcoRI sites in the complex are accessible to the enzyme while 20% are "closed." This degree of coverage is consistent with the mass ratio of DNA:histone in the complex as revealed by the buoyant density of the formaldehyde-fixed complex. We conclude that the histones in the complex are located randomly on the SV 40 genome and cover approximatley 20% of the DNA. These results suggest that the histone species F2b, F2al, F2a2, and F3 are bound without regard to nucleotide sequence of SV 40 DNA.
On primary structure and biosynthesis of histidine-rich polypeptide from malarial parasite Plasmodium lophurae.
Authors: Kilejian A , Liao T H , Trager W
Published: May 2006

Experiments on the primary structure of a histidine-rich polypeptide isolated from the malarial parasite Plasmodium lophurae indicate that the smaller quantities of amino acids other than histidine form an integral part of the polypeptide and do not arise from a protein contaminating a histidine homopolymer. In culture, the parasites incorporate over 50% of exogenously supplied histidine into the histidine-rich polypeptide and this incorporation is inhibited by puromycin.
Recovery of infectious proviral DNA from mammalian cells infected with respiratory syncytial virus.
Authors: Simpson R W , Iinuma M
Published: May 2006

The DNA fraction from a line of bovine embryonic kidney cells originally exposed as primary cultures several months earlier to a temperature-sensitive (ts) mutant of respiratory syncytial (RS) virus could be used to transfect human HEp-2 cells with the production of infectious RS virus. The DNA donor cells, designated BEK/RS ts, retained their healthy fibroblastic appearance during continuous cultivation at a temperature (39 degrees) restrictive for growth of the original infecting mutant and showed no evidence for RS virus replication or viral antigen synthesis when directly examined for these activities by conventional methods. The infectious property of the DNA from BEK/RS ts cells was abolished by exposure of the nucleic acid preparation to DNase (but not RNase) or by pretreatment of recipient HEp-2 cells with actinomycin D or mitomycin C. The latter drug treatments substantially enhanced the replication of infecting wild-type RS virus in HEp-2 cells. Viral isolates derived from the progeny of a DNA transfection included clones possessing several genetic markers of the RS ts mutant originally used to infect BEK/RS ts cells and other virus clones that appeared to be either hybrid or wild-type for phenotypic properties such as their temperature sensitivity. An infectious proviral DNA was also detected in a line of virogenic HEp-2 cells (HEp-2/RS) persistently infected with respiratory syncytial virus after exposure to the wild-type strain 2 years earlier.
Ribonuclease-sensitive DNA-synthesizing complex in human sperm heads and seminal fluid.
Authors: Witkin S S , Korngold G C , Bendich A
Published: May 2006

An endogenous DNA-synthesizing complex sensitive to ribonuclease has been found in purified preparations of swollen human sperm heads. Incorporation of [3H]dTTP into acid-precipitable material occurred in the presence of actinomycin D and required addition of dGTP, dCTP, dATP, plus Mg++. Polymerization was sensitive to pretreatment of the complex with pancreatic RNase A or Triton X-100. Exogenous activity was elicited by the synthetic template (dT)12--18-(rA)n but not by (dT)12--18-(dA)n or (dT)10. The complex sedimented from a 10,000 X g supernatant by centrifugation at 165,000 X g for 60 min and banded in sucrose at a density of 1.21--1.25 g/cm3. Endogenous RNase-sensitive DNA polymerase activity from cell-free seminal fluid was also detected in a fraction in sucrose at a density of 1.22 g/cm3. This activity was labile to freezing and stimulated by 0.04% Triton X-100, and thus differed from that of sperm heads.
Specificity of substrate recognition by the EcoRI restriction endonuclease.
Authors: Polisky B , Greene P , Garfin D E , McCarthy B J , Goodman H M , Boyer H W
Published: May 2006

The substrate specificity of the EcoRI restriction endonuclease can be varied in vitro by changing the pH and the ionic environment of the reaction. Phosphodiester bond cleavage occurs at a DNA hexanucleotide sequence d(N-G-A-A-T-T-C-N)/d(N-C-T-T-A-A-G-N) when the ionic strength is high, 100 mM Tris-HCl, 50 mM NaCl, 5 mM MgCl2, and the pH is approximately 7.3. Lowering the ionic strength to 25 mM Tris-HCl, 2 mM MgCl2, and adjusting the pH to 8.5 reduces the recognition specificity of the EcoRI endonuclease to the tetranucleotide sequence, d(N-A-A-T-T-N)/d(N-T-T-A-A-N). The enzymatic activity responsible for this substrate recognition is referred to as EcoRI. Cleavage of pVH51 plasmid DNA under EcoRI conditions results in a number of partial digest fragments, some of which disappear slowly over a prolonged digestion period. This suggests that different recognition sites are cleaved at different rates. Comparison of DNA fragment patterns of modified and unmodified pVH51 DNA indicates that the canonical EcoRI sequence is the most rapidly cleaved site under EcoRI conditions. DNA modified in vivo by the EcoRI methylase is not cleaved by the EcoRI endonuclease under standard conditions, but is cleaved under EcoRI conditions at sites other than the standard EcoRI substrate.
Fatty acids as modulators of membrane functions: catecholamine-activated adenylate cyclase of the turkey erythrocyte.
Authors: Orly J , Schramm M
Published: May 2006

Activation of the adenylate cyclase [ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1[ from turkey erythrocytes by isoproterenol decreased precipitously below 26 degrees. Certain unsaturated fatty acids enhanced the activation by isoproterenol up to 25-fold at reduced temperatures. The fatty acid also enhanced the formation of a persistent active state of the enzyme which was produced by preincubation with guanosine 5'-(beta,gamma-imino)triphosphate [Gpp(NH)p]. Once the enzyme had been activated by Gpp(NH)p plus isoproterenol the reaction rate was no longer as temperature sensitive and the fatty acid had little effect. The synthetic Gpp(NH)p apparently substituted for the natural GTP, which is known to play a regulatory role in the adenylate cyclase system. The findings suggest that the function of GTP which is mediated by the hormone is the temperature-sensitive event which is enhanced by the fatty acid. The use of free fatty acid to probe membrane-associated reactions in intact cells and in isolated membrane preparations is proposed.
31P magnetic resonance of tRNA.
Authors: Guéron M , Shulman R G
Published: May 2006

We have observed well-resolved 31P resonances at 65 kG (109 MHz) in solutions of Escherichia coli tRNAGlu and yeast tRNAPhe. One resolved resonance, identified as the terminal phosphate, titrates with a pK = 6.35. Upon melting the yeast tRNAPhe in 0.1 M NaCl, without Mg++, the resolved peaks broaden and disappear in the vicinity of 35 degrees while the central cluster narrows drastically, shifts slightly, and loses its structure. Addition of Mg++ shifts one resolved peak and changes the shape of the cluster. The 31P lines are broader when observed at 65 kG than at 24 kG. The broadening is shown to come from chemical shift anisotropy which is estimated to be about 140 ppm.
Ultrastructural localization of insulin receptors on adipocytes.
Authors: Jarett L , Smith R M
Published: May 2006

The method for preparing a stable, biologically active, covalently linked ferritin--insulin complex has been modified to provide a 25-fold increase in yield compared to the original procedure while reducing the molar fatio of ferritin to insulin to 1:1 from 40:1. Ultrastructural studies of isolated adipocytes revealed specific binding of ferritin--insulin to the cell surface in irregular clusters associated with the glycocalyx coating. The number of ferritin--insulin molecules observed was consistent with the number of sulin molecules observed was consistent with the number of receptors calculated from 125I-labeled insulin binding studies. The ferritin--insulin was not observed in the cytoplasm of the cell but was found on the convave side of surface connected vesicles. These surface connected vesicles were part of an alveolar-like system of plasma membrane invaginations which project in various directions in the cytoplasm and by thin sectioning can appear as pinocytotic-like microvesicles. The morphological observations on ferritin--insulin binding were supported by the finding that 125I-labeled insulin binding was almost exclusively localized to highly purified plasma membranes isolated by fractionation of adipocytes after incubation with 125I-labeled insulin. These data supported the theory that insulin did not need to enter a cell to cause biological effects and was consistent with the negative cooperativity concept of insulin binding to cell receptors.
Erythroid colony induction without erythropoietin by Friend leukemia virus in vitro.
Authors: Clarke B J , Axelrad A A , Shreeve M M , McLeod D L
Published: May 2006

Erythroid colonies could be produced without the addition of erythropeietin in plasma cultures seeded with bone marrow cells from normal C3Hf/Bi mice by exposure of the cells in vitro to medium from a cell line (IS) that continuously produces Friend leukemia virus in culture. The activity in the culture medium was viral rather than erythropoietin-like, since it was sedimentable by high-speed centrifugation and heat labile. Erythroid colonies did not develop when the bone marrow cells exposed to virus-containing medium were from mice genetically resistant to Friend virus. IS culture medium contained both Friend spleen focus-forming and XC-plaque-forming activities. No erythroid colonies were induced when genetically sensitive cells were exposed to a preparation from which the spleen focus-forming activity had been removed, but which contained XC plaque-forming activity in high concentration. Thus the spleen focus-forming component of Friend virus appeared to be responsible for inducing erythroid colony formation without erythropoietin in vitro. Some erythroid colonies were also found in control cultures to which neither virus nor erythropoietin had been added. Reduction in the concentration of fetal calf serum in the culture medium substantially decreased the number of these colonies but had only a minor effect on the number of virus-induced colonies. The number of erythroid colonies produced after 2 days of culture without erythropoietin or fetal calf serum was approximately proportional to the titer of Friend spleen focus-forming virus to whcih the bone marrow cells had been exposed. This system should prove useful for investigation in vitro of Friend virus--host cell interactions which lead to erythropoietin-independent erythropoiesis.
Competitive protein binding assay for methotrexate.
Authors: Myers C E , Lippman M E , Elliot H M , Chabner B A
Published: May 2006

A competitive protein binding assay has been developed for methotrexate based on the tight binding of this drug to Lactobacillus casei dihydrofolate reductase (= tetrahydrofolate dehydrogenase; 5,6,7,8-tetrahydrofolate: NADP+ oxidoreductase; EC 1.5.1.3). Free drug may be separated from that bound to reductase by adsorption with dextran--albumin coated charcoal. Scatchard plot analysis of the enzyme--drug interaction confirmed the presence of a single homogeneous class of binding sites with an association constant Ka of 2.1 X 10(8) M-1. This high affinity binding permits detection of methotrexate in the range of 0.3--30 pmol with a coefficient of variation of 15% or less. The predominant circulating folate, 5-methyl tetrahydrofolate, and the clinically useful rescue agent leucovorin (5-formyl tetrahydropteroyl-glutamic acid) do not interfere with the assay, nor does the methotrexate metabolite 4-amino-4-deoxy-10-methylpteroic acid. Assay of clinical samples, including plasma and cerebrospinal fluid, showed close agreement between the previously described enzyme inhibition assay and the more rapid competitive binding method.
Group selection, altruism, reinforcement, and throwing in human evolution.
Authors: Darlington P J
Published: May 2006

Evolution of altruism by group selection involves sacrifice of some individuals, not to the "group as a whole," but to other individuals in the group. Deme-group selection may establish strictly altruistic genes in a population, but only under limited conditions, and perhaps never among vertebrates, among which apparently altruistic behaviors may always potentially benefit the altruists. Responsive-group selection is a more effective mode of evolution of altruism, conspicuous in man. Evolutionary reinforcement increases the force of selection of advantageous behaviors, including altruistic ones, by making them pleasant or rewarding. It is probably involved also in ecological habitat selection, and may be the source of many human emotions, including esthetic ones. Throwing (of stones and weapons) exemplifies both the possible importance of a difficult-to-measure evolutionary factor and the role of reinforcement; in human evolution throwing may have been decisive in food-getting and fighting, in shifting emphasis from brute force to skill, and in inducing evolution of a brain able to handle three-body geometric problems precisely and thus preadapted for more complex functions.
Applications of synchrotron radiation to protein crystallography: preliminary results.
Authors: Phillips J C , Wlodawer A , Yevitz M M , Hodgson K O
Published: May 2006

X-ray diffraction photographs of protein single crystals have been obtained using synchrotron radiation produced by an electron-positron storage ring. The diffracted intensities observed with this unconventional source are a factor of at least 60 greater than those obtained with a sealed x-ray tube using the same crystal and instrumental parameters. Diffraction data have been collected by the precession method to higher resolution and using smaller protein crystals than would have been possible with a conventional source. The crystal decay rate in the synchrotron beam for several proteins appears to be substantially less than that observed with Ni-filtered Cu radiation. The tunable nature of the source (which allows selective optimization of anomalous contributions to the scattering factors) and the low angular divergence of the beam make the source very useful for single crystal protein diffraction studies.
Localization of phytochrome in etioplasts and its regulation
in vitro
of gibberellin levels
Authors: Evans A. , Smith H.
Published: May 2006

Etioplasts isolated from barley leaves and purified on a Sephadex G-50 (coarse) column were characterized by electron microscopy and nucleic acid analysis. The majority of etioplasts retained an intact outer envelope, and contamination by other fragments was extremely low. The level of gibberellin-like substances extractable from intact etioplast suspensions was enhanced within 5 min of the termination of a saturating red irradiation, and the response was far-red reversible. Ultra-sonication caused a 3-fold increase in extractable activity both in dark control suspension and suspensions treated with red light. It is concluded that phytochrome, as a function of its interconversions, probably causes the transport of gibberellin from inside the etioplast into the surrounding medium. This leads to increased production of active gibberellins, possibly by release of feedback control of late steps of the biosynthetic pathway. Dual wavelength difference spectrophotometry has demonstrated the presence of a proportion of total cellular phytochrome within the etioplast.
Synthesis of acetylcholine receptors by cultured chick myotubes and denervated mouse extensor digitorum longus muscles.
Authors: Devreotes P N , Fambrough D M
Published: May 2006

Mono-[125I]iodo- alpha - bungarotoxin - receptor complexes extracted from chick myotube cultures as well as from adult denervated extensor digitorum longus muscles of the mouse have been banded at their buoyant density in gradients of metrizamide-deuterium oxide. When cultures or denervated adult muscles are preincubated in media containing 2H- or 13C-substituted amino acids under conditions of active receptor accumulation, the mono-[125I]iodo-alpha-bungarotoxin-receptor complexes have an increased buoyant density and band in a position of higher density in the gradient relative to a marker of mono-[131I]iodo-alpha-bungarotoxin-receptor complexes extracted from cells preincubated in normal media. It is concluded that the accumulation of receptors on the surfaces of cultured chick myotubes and on the non-synaptic surfaces of extensor digitorum longus muscles following denervation are the result of de novo synthesis.
Analysis of membrane halves: cholesterol.
Authors: Fisher K A
Published: May 2006

Membrane splitting by freeze-fracture has been used as a preparative tool for chemical analysis of outer and inner "half"-membranes. In a previous report I showed that monolayers of human erythrocytes, bound to cationized glass, fracture nonrandomly, producing membrane fractions substantially enriched in outer or inner "halves." For the present study cells were used in quantities compatible with microanalysis. For quantitation the total amount of membrane present and the fractional enrichment of outer and inner "half"-membranes were determined. Cholesterol was examined by quantitative thin-layer chromatography modified to assay nanogram amounts. Comparison of lipids extracted from intact membranes with lipids from fractured membranes indicated that cholesterol was asymmetrically distributed across the plane of the membrane, more being present on the exterior side than on the interior side.
Transduction of chemical into electrical energy.
Authors: Nachmansohn D
Published: May 2006

The paper recalls some fundamental notions, developed by Otto Meyerhof, which were used in the analysis of the transduction of chemical into mechanical energy during muscular contraction. These notions formed the basis of the approach to the analysis of the transduction of chemical into electrical energy, i.e., the very principle underlying nerve and muscle excitability and bioelectricity. Instrumental for this purpose was the use, since 1937, of electric organs of fish, a tissue highly specialized for bioelectrogenesis.
Incorporation of fatty acids containing photosensitive groups into phospholipids of Escherichia coli.
Authors: Greenberg G R , Chakrabarti P , Khorana H G
Published: May 2006

A series of new fatty acids containing photosensitive groups at different positions on the paraffin chains supported the growth of an auxotroph of E. coli requiring unsaturated fatty acids. The derivatives were 6-, 9-, 11-, and 12-azidostearic acids, 12-azido-oleic acid, 16-azidopalmitelaidic acid, and 12-(4-azido-2-nitrophenoxy)-stearic and -oleic acids. Analyses of the phospholipids from cultures grown in the presence of the first six compounds showed that these derivatives accounted for 16-43% of the total fatty acids. Further analysis of phospholipids from cultures grown with 12-azido-oleic acid, 11-azidostearic acid, or 16-azidopalmitelaidic acid indicated that the azido fatty acids were at the 2-position of the glycerol moieties. The incorporation of these fatty acid derivatives offers a new approach to the study of membrane structure and, in particular, phospholipid-protein interactions by photolysis-induced crosslinking of the fatty acids to the structures in their immediate vicinity.
Anticomplementary nature of smaller DNA produced during synthesis of extensive DNA copies of poliovirus RNA.
Authors: Kacian D L , Myers J C
Published: May 2006

The reverse transcriptase (RNA-directed DNA nucleotidyltransferase) from avian myeloblastosis virus is able to make an extensive, possibly complete, complementary DNA copy of intact poliovirus RNA. In the presence of high concentrations of deoxyribonucleoside triphosphates, ribonucleoside triphosphates, or sodium pyrophosphate, this DNA is the only species produced. Without these additives, however, a second size class of DNA is also synthesized. This material has a sedimentation coefficient between roughly 4 and 10 S and is produced later in the reaction, largely after synthesis of the larger complementary DNA has ceased. The smaller DNA consists primarily of material anticomplementary to the RNA template and contains a faithful and uniform representation of the viral sequences. It most likely arises by transcription of the larger DNA species.
Amplification of moderately repetitive DNA sequences during chick cartilage differentiation.
Authors: Strom C M , Dorfman A
Published: May 2006

A 5-bromo-2'-[3H]deoxyuridine (BrdUrd) probe was isolated to analyze DNAs obtained from various chick tissues and cell types. [3H]BrdUrd-substituted DNA, prepared from limb bud cultures, was sheared and freed from palindromic DNA. Nonradioactive DNA was prepared from embryonic liver, undifferentiated limb bud mesenchyme, sternal cartilage, differentiated limb bud cultures, and BrdUrd-blocked cultures, and was sheared. These DNAs were used in 100-fold excess to drive the reassociation of the [3H]-BrdUrd-DNA probe. Purified mature cartilage DNAs of embryonic sternae or differentiated limb bud cultures drove the reassociation of the probe approximately two times faster than did DNA from liver, undifferentiated limb bud, or BrdUrd-blocked cells. These data indicate that cartilage DNA contains a greater number of sequences complementary to the BrdUrd probe than do DNAs of noncartilage or undifferentiated precartilage cells. Calculations determined an average substitution of 10% of thymidine residues by BrdUrd in purified probe, whereas CsCl density gradients of unsheared probe revealed radioactive peaks of greater than 20% substitution. The BrdUrd appears to be clustered in the genome.
Isolation of retina-specific cell-aggregating factor from membranes of embryonic neural retina tissue.
Authors: Hausman R E , Moscona A A
Published: May 2006

There is increasing evidence that developmental associations among embryonic cells are mediated by specific components of the cell surface. Earlier work has indicated that such components are extruded into the medium of primary monolayer cultures of embryonic cells, and that they represent the active constituents of the tissue-spedific cell-aggregating factors isolated fro- the supernatant medium of such cultures. We presently report that tissue-specific cell-aggregating factors can be obtained directly from embryonic tissues, and describe the isolation and partial purification of retina-spedific factor from a cell-membrane preparation derived from retina tissue of the chick embryo. Extraction of the purified membrane preparation with 1-butanol yielded an activity in the aqueous phase which resides in a protein probably a glycoprotein, with an estimated molecular weight of 50,000 in solution. This material could be obtained from retinas of embryos not older than 13 days, and only pre-13-day cells responded to its cell-aggregating activity. By these characteristics, this membrane-derived retina factor closely resembles the retina cell-aggregating glycoprotein previously purified from the supernatant medium of retina cell cultures. It is of special interest that the cell-aggregating protein is obtainable from cellular membranes during those stages of development when retina cells are most actively engaged in histological organization. Work in progress indicates that, by the procedures described herein, tissue-specific cell-aggregating factors can also be obtained from membrane preparations of other embryonic tissues.
Evidence for the lack of feedback regulation of gene activity and for the absence of subunit exchange between lactate dehydrogenase tetramers in vivo.
Authors: Nadal-Ginard B
Published: May 2006

Frog and rat lactate dehydrogenase (LDH, L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) isozymes, highly purfied by affinity chromatography, were injected into newly fertilized from (Rana pipiens) eggs to study the effect of the injected isozymes on the expression of the LDH genes of the developing frog embryo. From these experiments three conclusions can be drawn: (i) homologous and heterologous LDH, even when present in more than twice the normal amount of LDH, does not play any role in the switching on or off of the frog LDH genes or in regulating the level of gene activity. No evidence was found in support of any feedback regulation of LDH synthesis by the LDH molecule itself. (ii) Injected rat LDH is very stable in the frog embryo, and the contrary to what can be demonstrated in vitro, all the isozymes show the same stability. No evidence was found for selective degradation of any of the injected five isozymes. (iii) The fact that the injected isozymes disappear 12 days after the injection without forming hybrid molecules between themselves or with the stored or newly synthesized frog LDH is evidence in favor of the hypothesis that the tetramer is the degradative unit of LDH, not the monomer as others have postulated. The LDH tetramers do not dissociate the recombine under the physiological conditions present in frog cells.
Cellular cholesterol ester accumulation induced by free cholesterol-rich lipid dispersions.
Authors: Arbogast L Y , Rothblat G H , Leslie M H , Cooper R A
Published: May 2006

The influence of lipoprotein composition on free cholesterol (RC) esterification and accumulation of esterified cholesterol (EC) was studied in cells of Fu5AH rat hepatoma exposed to culture media supplemented with (FC/P) ranging from 0.6 to 2.8. In the presence of normal human serum, FC-lecithin dispersions witb a FC/P greater than one stimulated both the esterification of FC and the accumulation of cellular EC. Conversely, dispersions with FC/P values of approximately one had no effect on either FC esterification or the accumulation of EC when compared to cells grown in the absence of lipid dispersions. No stimulation of cellular response was obtained when FC-rich dispersions were added to cells in the absence of serum; however, stimulation was observed when cells were exposed to isolated human serum lipoproteins (very low density, low density, and high density). With each lipoprotein, FC-lecithin dispersions with FC/P values less than one inhibited cholesteryl ester accumulation, FC-rich dispersions stimulated, and dispersions with FC/P values of approximately one had an effect equivalent to that of isolated lipproteins alone. These studies extablish that cellular FC esterification and EC content are not influenced solely by the FC concentration of medium, but rather respond to the ratio of FC to phospholipidl These studies also suggest that serum lipoproteins participate in the transfer of FC carried by FC-rich lecithin dispersions to cells.