bluuebunny/crossref_metadata_2025 huggingface dataset:
Parasite sequestration in Plasmodium falciparum malaria: spleen and antibody modulation of cytoadherence of infected erythrocytes.
Authors: David P H , Hommel M , Miller L H , Udeinya I J , Oligino L D
Published: May 2006

Sequestration, the adherence of infected erythrocytes containing late developmental stages of the parasite (trophozoites and schizonts) to the endothelium of capillaries and venules, is characteristic of Plasmodium falciparum infections. We have studied two host factors, the spleen and antibody, that influence sequestration of P. falciparum in the squirrel monkey. Sequestration of trophozoite/schizont-infected erythrocytes that occurs in intact animals is reduced in splenectomized animals; in vitro, when infected blood is incubated with monolayers of human melanoma cells, trophozoite/schizont-infected erythrocytes from intact animals but not from splenectomized animals bind to the melanoma cells. The switch in cytoadherence characteristics of the infected erythrocytes from nonbinding to binding occurs with a cloned parasite. Immune serum can inhibit and reverse in vitro binding to melanoma cells of infected erythrocytes from intact animals. Similarly, antibody can reverse in vivo sequestration as shown by the appearance of trophozoite/schizont-infected erythrocytes in the peripheral blood of an intact animal after inoculation with immune serum. These results indicate that the spleen modulates the expression of parasite alterations of the infected erythrocyte membrane responsible for sequestration and suggest that the prevention and reversal of sequestration could be one of the effector mechanisms involved in antibody-mediated protection against P. falciparum malaria.
Localization of a poliovirus type 1 neutralization epitope in viral capsid polypeptide VP1.
Authors: van der Werf S , Wychowski C , Bruneau P , Blondel B , Crainic R , Horodniceanu F , Girard M
Published: May 2006

Poliovirus type 1 cDNA sequences coding for viral capsid polypeptide VP1 were inserted into the beta-lactamase sequence of Escherichia coli plasmid pBR322. Resulting recombinant plasmid pSW119 expressed in Escherichia coli a VP1-beta-lactamase fusion protein that reacted with antibodies raised against poliovirus capsid polypeptide VP1 and with a monoclonal poliovirus type 1 neutralizing antibody, C3. Deletions of various lengths were generated within the VP1 sequence. The hybrid proteins expressed by the deleted plasmids did not react any more with C3 when the region of VP1 amino acids 95-110 (poliovirus nucleotides 2,754-2,806) was deleted. Therefore, the C3 epitope responsible for virus neutralization is most probably located in this region of the capsid polypeptide.
Expression of a preproinsulin-beta-galactosidase gene fusion in mammalian cells.
Authors: Nielsen D A , Chou J , MacKrell A J , Casadaban M J , Steiner D F
Published: May 2006

As an approach to the study of mammalian gene expression, the promoters and translation initiation regions of the rat preproinsulin II and the simian virus 40 early genes were fused to the structural gene of Escherichia coli beta-galactosidase, a sensitive probe for gene expression. These fusions were introduced into COS-7 cells, a simian virus 40 large tumor-antigen-producing monkey kidney cell line, where they directed the synthesis of enzymatically active hybrid beta-galactosidase proteins. Conditions for transfection were varied to optimize the expression of beta-galactosidase activity in the transfected cells. The pH optimum of this activity was found to be 7.0, the same as that of native E. coli beta-galactosidase and distinct from the major lysosomal "acid" beta-galactosidase. The fused preproinsulin-beta-galactosidase was further characterized by gel electrophoresis of nondenatured cell extracts stained by a fluorogenic substrate and by immunoprecipitation and gel electrophoresis of 3H-labeled cell proteins. These results all indicate that fully active tetrameric beta-galactosidase hybrids can be produced in mammalian cells. The expression of preproinsulin-beta-galactosidase activity was measured in the presence of high glucose, insulin, dexamethasone, or epidermal growth factor but no regulatory changes were observed.
DNA sequence of the Escherichia coli tonB gene.
Authors: Postle K , Good R F
Published: May 2006

The nucleotide sequence of a cloned section of the Escherichia coli chromosome containing the tonB gene has been determined. Transcription initiation and termination sites for tonB RNA have been determined by S1 nuclease mapping. The tonB promoter and terminator resemble other E. coli promoters and terminators; the sequence of the tonB terminator region suggests that it may function bidirectionally. The DNA sequence specifies an open translation reading frame between the 5' and 3' RNA termini whose location is consistent with the position of previously isolated tonB::IS1 mutations. The DNA sequence predicts a proline-rich protein with a calculated size of 26.1-26.6 kilodaltons (239-244 amino acids), depending on which of three potential initiation codons is utilized. The predicted NH2 terminus of tonB protein resembles the proteolytically cleaved signal sequences of E. coli periplasmic and outer membrane proteins; the overall hydrophilic character of the protein sequence suggests that the bulk of the tonB protein is not embedded within the inner or outer membrane. A significant discrepancy exists between the calculated size of tonB protein and the apparent size of 36 kilodaltons determined by sodium dodecyl sulfate/polyacrylamide gel electrophoresis.
pif mutation blocks recombination between mitochondrial rho+ and rho- genomes having tandemly arrayed repeat units in Saccharomyces cerevisiae.
Authors: Foury F , Kolodynski J
Published: May 2006

Three allelic nuclear mutants affected in the recombination of mtDNA have been characterized in Saccharomyces cerevisiae and assigned to the PIF locus. In the mutants, the general recombination measured by the recombination frequency between linked or unlinked alleles is normal. However, the pif mutations prevent the integration into the rho+ genome of the markers (oli1, oli2, diu1, ery, oxi1, oxi2) of those rho- genomes that have tandemly arrayed repeat units. Therefore, these rho- genomes characterize a PIF-dependent recombination system. The pif mutations have also revealed the existence of a PIF-independent recombination system used by those rho- genomes that have an inverted organization of their repeat units. The markers of such palindromic rho- genomes exhibit high integration frequency into the rho+ genome even in the presence of the pif mutation. In addition, the pif mutations greatly increase suppressiveness in crosses between pif rho+ strains and PIF-dependent as well as PIF-independent rho- clones. We conclude that the recombination between rho+ and rho- genomes involves at least two distinct systems that depend on the organization of the rho- genome.
Nucleotide-stimulated proteolysis of histone H1.
Authors: Surowy C S , Berger N A
Published: May 2006

We have identified a proteolytic system that selectively degrades histone H1 in normal human lymphocytes. Treatment of permeabilized human lymphocytes with a series of nucleotides produced a marked decrease in their histone H1 content compared to untreated cells. The nucleotide-stimulated process was selective for histone H1 because gel electrophoresis showed that almost all other lymphocyte protein bands remained constant while histone H1 disappeared. The elimination of histone H1 appears to be the result of proteolysis by a trypsin-like enzyme because it was inhibited by phenylmethylsulfonyl fluoride, antipain, soybean trypsin inhibitor, and diisopropyl fluorophosphate. Proteolysis was stimulated by P1,P4-di(adenosine-5') tetraphosphate, P1,P3-di(adenosine-5') triphosphate, P1,P5-di(adenosine-5') pentaphosphate, adenosine 5'-tetraphosphate, ATP, adenosine 5'-[alpha, beta-methylene]triphosphate, adenosine 5'-[beta, gamma-methylene]triphosphate, ADP, CTP, GTP, UTP, dATP, or pyrophosphate, whereas AMP, adenosine, adenosine diphosphoribose, NAD+, cAMP, or sodium phosphate did not show this stimulation of proteolysis. ATP, [alpha, beta-methylene]ATP, [beta, gamma-methylene]ATP, and pyrophosphate all stimulated proteolysis, suggesting that a pyrophosphate linkage was necessary for this process. Thus, resting human lymphocytes contain a trypsin-like protease that is stimulated by nucleotides or pyrophosphate to selectively degrade histone H1.
Molecular cloning and cell cycle-specific regulation of a functional human thymidine kinase gene.
Authors: Bradshaw H D
Published: May 2006

A functional thymidine kinase (TK; ATP:thymidine 5'-phosphotransferase, EC 2.7.1.21) gene has been molecularly cloned from human DNA. The gene was rescued from a genomic library of TK-deficient mouse L cells transformed to the TK+ phenotype with total HeLa cell DNA. Of 14 overlapping clones, only one contained the intact human TK gene. The cloned recombinant bacteriophage carries a 16-kilobase insert derived entirely from human DNA and is capable of transforming LTK- cells to TK+ with an efficiency of 10 TK+ colonies per ng of DNA per 10(6) cells. Restriction endonuclease mapping shows that the functional human TK gene is at least twice as long as that reported for chicken. A 1.6-kilobase Xho I/EcoRI fragment was subcloned and found to hybridize to a human mRNA of 1.5 kilobases. When introduced into LTK- cells, the cloned human TK gene is regulated in the cell cycle-specific manner characteristic of TK+ mammalian cells. That is, TK activity in synchronized cells increases markedly with the onset of DNA synthesis. The signals governing the S-phase induction of TK activity reside within 16 kilobases of human DNA and are correctly interpreted by mouse cells.
Control of H-2 antigen and beta 2-microglobulin gene expression in mouse trophoblast cell clones.
Authors: Tanaka K , Ozato K , Jay G , Parnes J R , Ramanathan L , Seidman J G , Chang K S , Appella E
Published: May 2006

We have investigated the expression of H-2 antigen and beta 2-microglobulin in recently established mouse trophoblast cell clones. These clones, derived from C57BL/6 (H-2b) or BALB/c (H-2d) strains, synthesized extra-embryonic endoderm- and trophectoderm-specific cytoskeletal proteins, termed "Endo A" and "Endo B," and no detectable SSEA-1 embryonic antigen. Flow microfluorometry indicated that H-2 antigen expression on the cell surface of trophoblast cells was very low, corresponding to 2-5% of the amounts found on differentiated teratocarcinoma cells (12-1a) and BALB/c 3T3 cells, respectively. Expression of beta 2-microglobulin was reduced to approximately equal to 14% of the amounts found on 12-la cells. Immunoprecipitation and polyacrylamide gel electrophoretic analysis indicated that the synthesis of H-2 antigen and beta 2-microglobulin in the trophoblast cells was lower than that found in normal spleen cells. In addition low, but unequal, levels of mRNA specific for H-2 antigen and beta 2-microglobulin were found in trophoblast cells by blot hybridization with cDNA probes. The low mRNA levels may be due to transcriptional control of the genes encoding H-2 antigen and beta 2-microglobulin.
Chromatid damage after G2 phase x-irradiation of cells from cancer-prone individuals implicates deficiency in DNA repair.
Authors: Parshad R , Sanford K K , Jones G M
Published: May 2006

Ten lines of skin fibroblasts from individuals with genetic disorders predisposing to a high risk of cancer were compared with nine lines from normal adult donors with respect to chromatid damage after x-irradiation [25, 50, and 100 rad (0.25, 0.50, and 1 gray)] during G2 phase. The 10 cell lines represented five genetic disorders: Bloom syndrome, familial polyposis, Fanconi anemia, Gardner syndrome, and xeroderma pigmentosum, complementation groups A(XP-A), C(XP-C), E(XP-E), and variant (XP-Va). The incidence of chromatid breaks in all cancer-prone lines except XP-E and XP-A was significantly higher than in the normal lines. The incidence of chromatid gaps in all cancer-prone lines except XP-A and XP-Va was significantly higher than in the normal lines. Because each chromatid apparently contains a single continuous DNA double strand, chromatid breaks and gaps represent unrepaired DNA strand breaks arising directly or indirectly during excision repair of x-ray-induced DNA damage. These cytogenetic data together with results from use of the DNA repair inhibitor arabinofuranosyl cytosine (cytosine arabinoside) suggest that cells from all of these cancer-prone individuals are deficient in some step of DNA repair, predominantly excision repair operative during the G2-prophase period of the cell cycle. It appears that these DNA repair deficiencies are associated with a genetic predisposition to a high risk of cancer.
A flow- and time-dependent index of ischemic injury after experimental coronary occlusion and reperfusion.
Authors: DeBoer L W , Rude R E , Kloner R A , Ingwall J S , Maroko P R , Davis M A , Braunwald E
Published: May 2006

The purpose of the present study was to determine whether an ischemic index--expressed as the product of flow deprivation (FD) and the duration of occlusion (T), FD X T--correlated with biochemical and early morphologic alterations of the subendocardial myocardium and could predict ultimate development of irreversible injury after coronary reperfusion. Myocardial biopsy specimens for measurement of ATP and other purines and for ultrastructure studies were obtained in vivo during coronary occlusion in a canine model and were considered relative to development of necrosis after coronary reperfusion. FD X T correlated negatively with ATP content [ATP, nmol/mg of cardiac protein = 23.6 - 0.24(FD X T) + 0.0007(FD X T)2; r = -0.81] and with a semiquantitative early histologic index of damage (r = 0.70). Values of (FD X T) less than 18 were associated with reversible injury--i.e., complete salvage after coronary reperfusion. (FD X T) greater than 18 was associated with varying degrees of necrosis; necrosis was severe (78 +/- 12% of subendocardial biopsy specimens) when ATP less than 10 nmol/mg of protein and total purine pool was decreased by 50%. FD X T correlated with the eventual percentage of subendocardial necrosis (r = 0.85). Accordingly, as an index of ischemic injury, FD X T may be useful in assessing whether ischemic myocardial tissue will benefit from early restoration of blood flow to the ischemic area.
Reversible uncoupling of oxidative phosphorylation at low oxygen tension.
Authors: Kramer R S , Pearlstein R D
Published: May 2006

The stoichiometry of oxidative phosphorylation at low oxygen tension (less than 3 torr; O2 less than 5 microM) has been measured in rat liver mitochondria. In a steady-state model in which respiration rate was experimentally controlled by either oxygen or substrate (succinate) limitation, flux-dependent variation in the phosphorylation efficiency (P/O ratio) of stimulated mitochondrial respiration was evaluated. P/O ratio remained constant over a wide range of respiration rates in mitochondria limited only by substrate availability. In contrast, oxygen-limited mitochondria demonstrated a continuous decline in P/O ratio as respiration was increasingly restricted. Significant differences in the two test conditions were demonstrated throughout the range of analysis. The effect of oxygen limitation on phosphorylation efficiency was shown to be completely reversed by restoring zero-order kinetics associated with high oxygen tension. These findings are discussed in regard to a proposed uncoupling of mitochondrial coupling site II at low oxygen tension arising as a consequence of energy-dissipating electron flux through the ubiquinone-cytochrome b-c1 region of the respiratory chain (complex III).
Cloning and expression of murine immune interferon cDNA.
Authors: Gray P W , Goeddel D V
Published: May 2006

The murine immune interferon (IFN-gamma) gene was cloned and expressed under control of the simian virus 40 early promoter in the monkey COS-1 cell line. A protein is secreted from these cells having the biological, antigenic, and biochemical characteristics of natural murine IFN-gamma. Cloned murine IFN-gamma cDNAs were obtained by using RNA from both mitogen-induced murine spleens and the transfected COS cells, and both code for identical proteins. The mature murine IFN-gamma encoded is 136 amino acids long, 10 amino acids shorter than human IFN-gamma. The nucleotide homology between the murine and human IFN-gamma genes is 60-65%, whereas the encoded proteins are only 40% homologous. Murine IFN-gamma cDNA was expressed in Escherichia coli under trp promoter control.
Bis-penicillamine enkephalins possess highly improved specificity toward delta opioid receptors.
Authors: Mosberg H I , Hurst R , Hruby V J , Gee K , Yamamura H I , Galligan J J , Burks T F
Published: May 2006

The conformationally restricted, cyclic, disulfide-containing, enkephalin analogs [2-D-penicillamine, 5-L-penicillamine]enkephalin [(D-Pen2,L-Pen5]enkephalin) and [2-D-penicillamine, 5-D-penicillamine]enkephalin [(D-Pen2,D-Pen5]enkephalin) were synthesized by solid-phase methods. Selectivities of these analogs for a single class of opioid receptor were investigated by examining relative potencies in the mouse vas deferens assay, in which the functional receptor is the delta receptor, versus the guinea pig ileum assay, in which the mu receptor is the functional receptor, and by determining their relative abilities to displace the prototypical delta receptor ligand [D-Ala2, D-Leu5]enkephalin and the prototypical mu receptor ligand naloxone from rat brain membrane preparations. Based on these comparisons [D-Pen2,L-Pen5]- and [D-Pen2,D-Pen5]enkephalin exhibited delta receptor selectivities of 1,088 and 3,164, respectively, in the bioassays, and 371 and 175, respectively, in the binding assays. Compared with the previously reported delta receptor selective analogs, [D-Ala2,D-Leu5]enkephalin, [D-Ser2,Leu5,Thr6]enkephalin, and [D-Thr2,Leu5,Thr6]enkephalin, the bis-Pen-containing analogs provide an order of magnitude increase in delta receptor selectivity.
Tumor induction by direct injection of cloned v-src DNA into chickens.
Authors: Fung Y K , Crittenden L B , Fadly A M , Kung H J
Published: May 2006

The in vitro transforming potential of Rous sarcoma viruses has been shown to reside in a gene segment, v-src. To provide direct proof that the isolated v-src sequence contains all the information necessary for sarcoma induction in vivo, we have injected subgenomic and genomic cloned DNA carrying v-src directly into chickens. We report that v-src, without the complements of other viral genes, can effectively induce sarcomas in the host within weeks after injection. The sarcoma tumors generated by subgenomic v-src DNA are of mono- or oligoclonal origin. They have acquired v-src sequences in the genome and expressed high levels of src messages. Intact long terminal repeats do not seem to be essential in the tumorigenic process.
Protein degradation in Escherichia coli: the lon gene controls the stability of sulA protein.
Authors: Mizusawa S , Gottesman S
Published: May 2006

Escherichia coli lon mutants are defective in the ATP-dependent proteolysis of abnormal proteins. The mutants are also sensitive to ultraviolet light (UV) in that septation is inhibited after exposure to UV. sulA mutations, isolated as suppressors of UV sensitivity unlinked to lon, do not affect proteolysis but allow septation to occur after DNA damage. We have confirmed the hypothesis that the product of the sulA gene is degraded by lon proteolysis. If sulA (the product of sulA) is a UV-inducible division inhibitor, as suggested by a variety of experiments, lon (the product of lon) may regulate cell division by regulating the half-life of sulA. We cloned the sulA gene in a bacteriophage lambda vector from a plasmid carrying the ompA region of E. coli. An 18-kilodalton polypeptide was identified as the product of the sulA gene. Pulse-chase labeling demonstrated that the half-life of the sulA protein is 1.2 min in lon+ cells and 19 min in lon- cells. This work demonstrates that lon proteolysis affects the stability of a native E. coli protein.
Regulation of a metallothionein-growth hormone hybrid gene in bovine papilloma virus.
Authors: Pavlakis G N , Hamer D H
Published: May 2006

We have constructed bovine papilloma virus recombinants carrying a hybrid gene in which human growth hormone structural sequences are fused to the promoter and presumptive control region of the mouse metallothionein-I gene. Mouse cells transformed with the recombinants synthesize metallothionein-growth hormone hybrid mRNA with the same 5' end as metallothionein mRNA. Hybrid mRNA is inducible by cadmium but not by dexamethasone, whereas the chromosomal metallothionein genes in the same cells are inducible by both agents. This indicates that heavy metals and glucocorticoids regulate the mouse metallothionein-I gene by independent mechanisms. Furthermore, because the transformed cells produce large quantities of human growth hormone polypeptide, it appears that bovine papilloma virus-metallothionein vectors may be useful for obtaining the efficient and regulatable expression of other eukaryotic gene products. We have constructed recombinants that allow the insertion of new structural sequences under the control of the metallothionein promoter.
Laminin receptor on human breast carcinoma cells.
Authors: Terranova V P , Rao C N , Kalebic T , Margulies I M , Liotta L A
Published: May 2006

Human MCF-7 breast carcinoma cells possess a receptor-like moiety on their surface that has a high binding affinity (Kd = 2 nM) for laminin, a glycoprotein localized in basement membranes. Laminin preferentially stimulates (8-fold) MCF-7 cells to attach to type IV (basement membrane) collagen, whereas fibronectin stimulates attachment only 2-fold for these cells on type I collagen. The attachment properties of two other human breast carcinoma cell lines to type IV collagen were also studied. The attachment of ZR-75-1 cells was stimulated 4-fold by laminin and 5-fold by fibronectin, whereas T47-D cell attachment was stimulated 2-fold by laminin and 7-fold by fibronectin. By employing protease-derived fragments of laminin, the major domains of the laminin molecule that participate in MCF-7 cell attachment to type IV collagen were identified. The whole laminin molecule has the configuration of a four-armed cross with three short arms and one long arm. A major cell-binding domain was found to reside near the intersection point of the short arms, and the type IV collagen-binding domain was associated with the globular end regions of the short arms. The receptor for laminin on the surface of these tumor cells may be involved in the initial interaction of tumor cells via laminin with the vascular basement membrane to facilitate invasion and subsequent promotion of metastasis.
Isolation and DNA sequence of a full-length cDNA clone for human X chromosome-encoded phosphoglycerate kinase.
Authors: Michelson A M , Markham A F , Orkin S H
Published: May 2006

Phosphoglycerate kinase (PGK), a major enzyme in glycolysis, is encoded by the X chromosome in mammals. We have initiated molecular analysis of the PGK structural gene by isolating a full-length cDNA clone from a human fetal liver cDNA library. Synthetic oligonucleotide mixtures encoding two different portions of PGK were used as direct in situ hybridization probes for the cDNA library. Several classes of clones were obtained based on their hybridization at different stringencies to one or both of the PGK oligonucleotide mixtures. One clone, designated pHPGK-7e, which hybridized at high stringency to each of the synthetic probes, encoded the complete PGK protein sequence as well as 82 base pairs of 5' and 437 base pairs of 3' untranslated regions. Southern blot analysis of human genomic DNAs revealed a complex pattern of hybridizing fragments, two of which were non-X in origin. These results suggest that the human genome contains a small family of dispersed PGK or PGK-like genes.
The murine Ia alpha chains, E alpha and A alpha, show a surprising degree of sequence homology.
Authors: Benoist C O , Mathis D J , Kanter M R , Williams V E , McDevitt H O
Published: May 2006

The I region of the murine major histocompatibility complex codes for a group of glycoproteins, the Ia antigens, thought to be involved in the control of immune responsiveness. Each Ia antigen complex contains a "heavy chain," a "light chain," and the "invariant chain." We describe here the isolation and characterization of genomic and cDNA clones for one of the heavy chains, Ak alpha. The complete nucleotide sequence of the cDNA clone is presented, and the predicted amino acid sequence is compared with that of another alpha chain, Ek alpha. About 50% of the amino acids are identical, a finding somewhat unexpected on the basis of preliminary protein sequence data.
Heat shock protein in mammalian brain and other organs after a physiologically relevant increase in body temperature induced by D-lysergic acid diethylamide.
Authors: Cosgrove J W , Brown I R
Published: May 2006

A physiologically relevant increase in body temperature from 39.7 to 42.5 degrees C, which was generated after the intravenous injection of D-lysergic acid diethylamide (LSD), caused the induction of synthesis of a 74,000-dalton heat shock protein in the brain, heart, and kidney of the young adult rabbit. A marked increase in the relative labeling of a 74,000-dalton protein was noted after analysis of both in vivo labeled proteins and cell-free translation products of isolated polysomes. A temporal decrease in the synthesis of this protein was noted as LSD-induced hyperthermia subsided. The 74,000-dalton protein, which is induced in various organs of the intact animal at a body temperature similar to that attained during fever reactions, may play a role in homeostatic control mechanisms.
Influence of nuclear spin on chemical reactions: Magnetic isotope and magnetic field effects (A Review)
Authors: Turro Nicholas J.
Published: May 2006

The course of chemical reactions involving radical pairs may depend on occurrence and orientation of nuclear spins in the pairs. The influence of nuclear spins is maximized when the radical pairs are confined to a space that serves as a cage that allows a certain degree of independent diffusional and rotational motion of the partners of the pair but that also encourages reencounters of the partners within a period which allows the nuclear spins to operate on the odd electron spins of the pair. Under the proper conditions, the nuclear spins can induce intersystem crossing between triplet and singlet states of radical pairs. It is shown that this dependence of intersystem crossing on nuclear spin leads to a magnetic isotope effect on the chemistry of radical pairs which provides a means of separating isotopes on the basis of nuclear spins rather than nuclear masses and also leads to a magnetic field effect on the chemistry of radical pairs which provides a means of influencing the course of polymerization by the application of weak magnetic fields.
Processing of transcription products of the gene encoding the RNA component of RNase P.
Authors: Sakamoto H , Kimura N , Shimura Y
Published: May 2006

The gene coding for the RNA component of RNase P from Escherichia coli was transcribed in vitro by using a restriction fragment carrying the gene as template. The terminal sequences of the transcription products were determined and mapped on the DNA sequence of the gene. The signals for transcription initiation and termination of the gene were thus identified. Transcription termination occurs within a region of two bases at positions 413 and 414 from the transcription start site. The transcripts carry extra stretches of 36 and 37 nucleotides at the 3' end of the RNA molecule isolated from the enzyme. The extra sequences were removed in vitro when the transcripts were incubated with a crude cell extract.
Evolution of a finite population under gene conversion.
Authors: Nagylaki T
Published: May 2006

Evolution at a multiallelic locus under the joint action of gene conversion, mutation, selection, and random genetic drift is studied. Generations are discrete and nonoverlapping; the diploid, monoecious population mates at random. Under the assumption that all four evolutionary forces are weak, a diffusion approximation is established for the dynamics of the gene frequencies. For two alleles, the inclusion of gene conversion merely alters one of the two selection parameters of the thoroughly investigated diffusion process without conversion. Therefore, all results for this classical process, some of which are reviewed and extended here, are immediately applicable to the biologically more general problem. Small conversional disparities can dramatically affect the fixation probability (and hence the rate of gene substitution) and can greatly reduce the mean conditional fixation time of a new mutant. The mean absorption and fixation times are often sufficiently short to imply that biased gene conversion can be an important mechanism for the loss of genetic variability in and the genetic divergence of isolated populations.
Functional immunoglobulin M production after transfection of cloned immunoglobulin heavy and light chain genes into lymphoid cells.
Authors: Ochi A , Hawley R G , Hawley T , Shulman M J , Traunecker A , Köhler G , Hozumi N
Published: May 2006

The rearranged immunoglobulin heavy (mu) and light (kappa) chain genes cloned from the Sp6 hybridoma cell line producing immunoglobulin M specific for the hapten 2,4, 6-trinitrophenyl were inserted into the transfer vector pSV2-neo and introduced into various plasmacytoma and hybridoma cell lines. The transfer of the mu and kappa genes resulted in the production of pentameric, hapten-specific, functional IgM.
Sarcomere length and tension changes in tetanized frog muscle fibers after quick stretches and releases.
Authors: Sugi H , Kobayashi T
Published: May 2006

The sarcomere length changes in tetanized frog muscle fibers in response to quick fiber length changes were examined along the fiber length with a high-sensitivity laser diffraction technique. The experiments were only performed with muscle fibers in which the uniform orientation and sarcomere length of the component myofibrils were well preserved during a tetanus. When the sarcomere length changes were recorded near the fixed fiber end, the delay of the onset of sarcomere length change in response to the applied fiber length change tended to be longer than that of the onset of tension changes recorded at the fixed fiber end. The magnitude of sarcomere length changes was larger near the moving fiber end than near the fixed fiber end. In the case of quick releases, the resulting sarcomere shortening tended to outlast the fiber shortening, so that the quick tension recovery started during the sarcomere shortening. These results indicate (i) that the tension changes in response to quick fiber length changes may not give direct information about the cross-bridge properties and (ii) that the viscoelastic multisegmental nature of muscle fibers should be taken into consideration in interpreting the tension responses to quick length changes.
Beta-globin locus is linked to the parathyroid hormone (PTH) locus and lies between the insulin and PTH loci in man.
Authors: Antonarakis S E , Phillips J A , Mallonee R L , Kazazian H H , Fearon E R , Waber P G , Kronenberg H M , Ullrich A , Meyers D A
Published: May 2006

Using a parathyroid hormone (PTH) cDNA probe we found a common Pst I polymorphic restriction site 3' to the PTH gene in all ethnic groups examined. Because the PTH, insulin, and beta-globin loci have been localized to the short arm of chromosome 11 (11p) we used DNA polymorphisms adjacent to each of these three loci to determine whether they are genetically linked and to determine their order. We found that the PTH and beta-globin loci are closely linked (estimated recombination fraction, 0.07; 95% confidence limits, 0.05-0.10; lod score, 4.63; odds favoring linkage, 42,000:1). Furthermore, our findings strongly indicate that the beta-globin gene cluster lies between the PTH and insulin loci. Therefore, the gene order on 11p is centromere-PTH-beta-globin-insulin.
Selective cleavage in the avian retroviral long terminal repeat sequence by the endonuclease associated with the alpha beta form of avian reverse transcriptase.
Authors: Duyk G , Leis J , Longiaru M , Skalka A M
Published: May 2006

M13 recombinant DNA clones containing a 350-base sequence derived from the EcoRI fragment of two tandemly linked Rous-associated virus 2 (RAV-2) long terminal repeat (LTR) sequences have been used to map reverse transcriptase-associated endonuclease (RT-endonuclease) cleavage sites by primer extension studies. Under appropriate conditions, the alpha beta form of RT-endonuclease (composed of both the alpha and beta subunits) purified from Avian sarcoma virus (Pr-C and B-77 strains) introduces a specific break in the inverted complementary repeat sequence found at the junction of the LTRs. The cleavage sites occur in the same nucleotide sequence in (-) and (+) DNA strands; together they have the potential of generating a 6-base-pair staggered overlap that spans the junction. This supports the notion that the enzyme is involved in viral DNA integration. Other RT-endonuclease sites were analyzed. A second site, which occurs in the lac region of the M13 vector DNA upstream from the unique EcoRI cloning site, bears no apparent sequence homology to the site at the junction of the LTRs. However, it also lies within an inverted complementary repeat and, as is the case for the site in the LTR, the break occurs to the 5' side of the axis of symmetry. Cleavage at this second site is suppressed when the vector contains the RAV-2 LTR insert. Thus, the viral LTR appears to exert a cis effect that can influence a region over 300 base pairs away.
A specific phosphoprotein phosphatase acts on histone H1 phosphorylated by protein kinase C.
Authors: Sahyoun N , LeVine H , McConnell R , Bronson D , Cuatrecasas P
Published: May 2006

A phosphohistone phosphatase from rat liver cytosol acts specifically on histone H1 that is phosphorylated with the Ca2+-phospholipid-dependent protein kinase (protein kinase C). The apparent Km for 32P-labeled H1 is 1 microM; other histones or cytosolic proteins phosphorylated with protein kinase C or with cyclic AMP-dependent protein kinase are poor substrates for the phosphatase. The enzyme has been partially purified by gel-permeation chromatography and by utilizing a high-performance liquid chromatography ion-exchange column. The physical properties of this enzyme include a Stokes radius of 5.0 nm, a sedimentation coefficient (s20,w) of 7.0 S, and a Mr of 150,000. The detection of protein kinase as well as the specific phosphohistone phosphatase in purified rat liver nuclei suggests a physiologic role for a histone H1 phosphorylation-dephosphorylation cycle mediated by protein kinase C and the corresponding phosphohistone phosphatase.
A perfectly symmetric lac operator binds the lac repressor very tightly.
Authors: Sadler J R , Sasmor H , Betz J L
Published: May 2006

A completely symmetric DNA segment has been constructed that binds the lactose repressor of Escherichia coli 10-fold more tightly than does the natural lactose operator sequence. This tight-binding operator is an inverted repeat of a 15-base-pair segment from the left half of the natural operator sequence, the inversion being about the point indicated by the arrow shown below: (sequence in text) where the upper sequence is the natural operator and the lower sequence is the symmetric operator. The increased affinity of repressor for this symmetric sequence supports the idea that the tetrameric repressor is designed for a two-module binding to DNA, presumably via two (or two pairs) of its identical subunits. The natural operator is apparently "flawed" by "incorrect" base pairs in the right operator half and by an "incorrect" spacing between the operator halves with respect to maximal repressor binding.
Monoclonal antibodies that prevent adhesion of B 16 melanoma cells and reduce metastases in mice: crossreaction with human tumor cells.
Authors: Vollmers H P , Birchmeier W
Published: May 2006

Monoclonal antibodies raised against B 16 melanoma cells in syngeneic mice were functionally screened for their ability to inhibit cell adhesion in tissue culture. Three of these antibodies (16/43, 16/77, 16/82), when preinjected into C57BL/6 mice, markedly reduced the number of experimental lung metastases produced by B 16 cells, possibly by interference with their adhesion to the lung endothelia. We now report that these monoclonal antibodies block in vitro attachment of the majority of human melanoma cell lines tested and also of carcinoma, neuroblastoma, and glioblastoma cells from both mice and humans but untransformed cell lines such as 3T3 mouse or MRC-5 human fibroblasts are not affected. The antibodies also react with mouse teratocarcinoma stem cells (F9, PCC4) but not with differentiated teratocarcinoma lines (PYS-2, 944). Furthermore, the antiadhesion activity of the antibodies could be quantitatively absorbed by intact human and mouse tumor cells but not by untransformed cells, suggesting that the corresponding antigens may represent tumor-associated cell surface components. Correspondingly, the antigens were found on simian virus 40-transformed 3T3 mouse fibroblasts and are expressed in a temperature-sensitive fashion in chicken fibroblasts transformed with a temperature-sensitive Rous sarcoma virus. On "immunoblots" of NaDodSO4-containing gels the three selected antibodies (16/43, 16/82, 19/1) were absorbed by antigens with molecular weights of 40,000 and 50,000.
Transcriptional activation of an unrearranged and untranslocated c-myc oncogene by translocation of a C lambda locus in Burkitt.
Authors: Croce C M , Thierfelder W , Erikson J , Nishikura K , Finan J , Lenoir G M , Nowell P C
Published: May 2006

We have studied somatic cell hybrids between mouse myeloma cells and IARC-BL2 Burkitt lymphoma human cells carrying a t(8;22) chromosome translocation for the presence and expression of human immunoglobin lambda chains and for the c-myc oncogene. The results indicate that the c-myc oncogene remains on the 8q+ chromosome and that the excluded and rearranged C lambda allele translocates from chromosome 22 to this chromosome 8. As a result of the translocation, transcriptional activation of the c-myc oncogene on the rearranged chromosome 8 (8q+) occurs, while the c-myc oncogene in the normal chromosome 8 is transcriptionally silent. These findings suggest that the translocation of a rearranged immunoglobulin locus to the 3' side of an unrearranged c-myc oncogene may enhance its transcription and contribute to malignant transformation.
Spatial correlation between primary redox components in reaction centers of
Rhodopseudomonas sphaeroides
measured by two electrical methods in the nanosecond range
Authors: Trissl H.-W.
Published: May 2006

Relative distances between the the primary donor P, the intermediary pheophytin acceptor H, and the iron-quinone acceptor Q of bacterial reaction centers were determined by recording laser flash-induced photovoltages in two experimental systems with nanosecond time resolution. In one system a suspension of chromatophores was subjected to a light gradient and in the other system chromatophores were spread at a heptane/water interface. The 10-ns back reaction occurring in reaction centers with reduced Q could be time resolved. The initial photovoltage amplitude under conditions in which the charge separation proceeded up to the state [P
+
H
-
] was about ⅔ of that when it proceeded up to the state [P
+
HQ
-
]. If the amplitude of the photovoltage is considered to be proportional to the spatial displacement of charges, this result means that pheophytin lies closer to Q than to P.
Biochemical and immunochemical evidence that the "major postsynaptic density protein" is a subunit of a calmodulin-dependent protein kinase.
Authors: Kennedy M B , Bennett M K , Erondu N E
Published: May 2006

By three criteria, two biochemical and one immunochemical, the major postsynaptic density protein (mPSDp) is indistinguishable from the 50-kilodalton (kDa) alpha subunit of a brain calmodulin-dependent protein kinase. First, the two proteins comigrate on NaDodSO4/polyacrylamide gels. Second, iodinated tryptic peptide maps of the two are identical. Finally, a monoclonal antibody (6G9) that was raised against the protein kinase binds on immunoblots to a single 50 kDa band in crude brain homogenates and to both the alpha subunit of the purified kinase and the mPSDp from postsynaptic density fractions. The purified kinase holoenzyme also contains a 60-kDa subunit termed beta. A comparison of the peptide map of beta with the maps of 60-kDa proteins from the postsynaptic density fraction suggests that beta is present there but is not the only protein present in this molecular weight range. These results indicate that the calmodulin-dependent protein kinase is a major constituent of the postsynaptic density fraction and thus may be a component of type I postsynaptic densities.
Loss of unstably amplified dihydrofolate reductase genes from mouse cells is greatly accelerated by hydroxyurea.
Authors: Snapka R M , Varshavsky A
Published: May 2006

Previous work has shown that mammalian cells that carry unstably amplified genes for dihydrofolate reductase (DHFR) gradually lose the amplified DHFR genes when grown in the absence of the DHFR inhibitor methotrexate (MTX). Unstably amplified genes occur on small acentric chromosomes called double minutes (DMs) or even smaller chromatin fragments, in contrast to stably amplified genes, which reside in centromere-containing chromosomes. We have found that the rate of loss of the unstably amplified DHFR genes can be greatly oncreased by growing the cells in the presence of a nonlethal concentration of hydroxyurea. For example, in one MTX-resistant subline studied, approximately equal to 90% of the original DHFR gene dosage is lost in 25-30 cell doublings in the absence of MTX. The same degree of loss is achieved, however, in less than 4 doublings if cells are grown in the presence of 50 microM hydroxyurea. This new effect of hydroxyurea does not appear to be due to changes in plating efficiency or selective cytotoxicity. In particular, no increase in cell death occurs at 50 microM hydroxyurea, and cells continue to multiply, albeit 1/2 to 2/3 as fast as in the absence of hydroxyurea. The ability to selectively accelerate the loss of amplified genes from mammalian cells as shown in the present work may have important implications both for the problem of drug resistance in cancer chemotherapy and for curing mammalian cells of extrachromosomally maintained DNA genomes of pathogenic viruses.
Passive transfer of Lambert-Eaton myasthenic syndrome with IgG from man to mouse depletes the presynaptic membrane active zones.
Authors: Fukunaga H , Engel A G , Lang B , Newsom-Davis J , Vincent A
Published: May 2006

In the Lambert-Eaton myasthenic syndrome (LEMS), there is a decreased release of acetylcholine quanta from the nerve terminal by nerve impulse. Recently, an autoimmune origin of LEMS was documented by passive transfer of its electrophysiologic features from man to mouse with IgG. Freeze-fracture electron microscopy of LEMS neuromuscular junctions has revealed a paucity of presynaptic membrane active zones. Thus, the active zones might be the targets of the pathogenic autoantibodies in LEMS. To test this assumption, freeze-fracture electron microscopic studies were done in mice injected with 10 mg of IgG daily from each of three LEMS patients and in control mice treated with normal human IgG or no IgG. IgG from patients 1 and 2 impaired neuromuscular transmission in mice, but IgG from patient 3 failed to do so. After 52-69 days of treatment, diaphragm or anterior tibial muscles were removed and coded. Paired muscles from control mice and mice receiving LEMS IgG were studied "blindly." Satisfactory freeze-fracture replicas of 185 presynaptic membrane P-faces were analyzed by stereometric methods. In mice treated with LEMS IgG that was pathogenic by electrophysiologic criteria, there was a selective depletion of active zones and active-zone particles but not of other membrane particles and there was a concomitant increase of large membrane particles aggregated into clusters. These findings provide additional evidence that the active zones facilitate quantal transmitter release by nerve impulse, lend further support to the assumption that the active-zone particles are Ca2+ channels, and establish mediation of the membrane lesions in LEMS by IgG.
Estradiol binds to a receptor-like cytosol binding protein and initiates a biological response in Paracoccidioides brasiliensis.
Authors: Loose D S , Stover E P , Restrepo A , Stevens D A , Feldman D
Published: May 2006

Paracoccidioidomycosis, a disease caused by Paracoccidioides brasiliensis, which is endemic to Latin America, is much more common in men than women, suggesting a role for hormonal factors. We recently showed that two other yeasts possess steroid binding proteins and postulated that these receptor-like molecules represented a mechanism by which the hormonal milieu of the host might influence an infecting pathogen. Therefore, we examined P. brasiliensis for a sex steroid binding protein. Because tritiated steroids rapidly dissociated from the other fungal binding proteins, we developed a fast binding method with Sephadex G-50 microcolumns speeded by centrifugation. This method detected specific binding of [3H]estradiol in P. brasiliensis cytosol. Other tritiated steroid hormones, including testosterone and corticosterone, failed to exhibit specific binding. Scatchard analysis of [3H]estradiol binding showed an apparent dissociation constant (Kd) of 1.7 X 10(-8) M and a maximal binding capacity (Nmax) of 235 fmol/mg of protein. Susceptibility to trypsin indicated the binding site was protein in nature. The protein had a Stokes radius of approximately equal to 32 A by HPLC exclusion column and a sedimentation coefficient of 4.4 S by sucrose gradient, consistent with an apparent Mr of approximately equal to 60,000. Competition experiments revealed that estrone, estriol, and progesterone had 25% of the affinity of estradiol, whereas diethylstilbestrol, androgens, and corticosteroids had low affinity. Investigation of steroid hormone actions in P. brasiliensis indicated that estradiol inhibited the fungal transformation from mycelial form to yeast form, the initial step of infection. This suppressive effect was dose-dependent and not found with testosterone. We hypothesize that endogenous estrogens in the host, acting through the cytosol binding protein in the fungus, inhibit mycelial-to-yeast transformation, thus explaining the resistance of women to paracoccidioidomycosis.
Thermodynamics of aqueous sodium chloride to 823 K and 1 kilobar (100 MPa)
Authors: Pitzer Kenneth S. , Li Yi-gui
Published: May 2006

It is shown that a very simple semiempirical equation represents quite accurately the thermodynamic properties of aqueous sodium chloride from 373 to 823 K. The equation comprises one Margules term and a Debye—Hückel term. Just the one Margules parameter is freely adjustable because the Debye—Hückel parameter is determined by the properties of water. The equation is valid from the saturation composition down to infinite dilution for solvent density above 0.75 g·cm
-3
but at lower density only above a solute mole fraction of about 0.1 on an ionized basis. Both solute and solvent activity coefficients are fitted from the lowest pressure of solution existence up to 1 kilobar (1 bar = 100 kPA). Derivation of enthalpy and other related functions is discussed.