bluuebunny/crossref_metadata_2025 huggingface dataset:
Human adult T-cell leukemia virus: complete nucleotide sequence of the provirus genome integrated in leukemia cell DNA.
Authors: Seiki M , Hattori S , Hirayama Y , Yoshida M
Published: May 2006

Human retrovirus adult T-cell leukemia virus (ATLV) has been shown to be closely associated with human adult T-cell leukemia (ATL) [Yoshida, M., Miyoshi, I. & Hinuma, Y. (1982) Proc. Natl. Acad. Sci. USA 79, 2031-2035]. The provirus of ATLV integrated in DNA of leukemia T cells from a patient with ATL was molecularly cloned and the complete nucleotide sequence of 9,032 bases of the proviral genome was determined. The provirus DNA contains two long terminal repeats (LTRs) consisting of 755 bases, one at each end, which are flanked by a 6-base direct repeat of the cellular DNA sequence. The nucleotides in the LTR could be arranged into a unique secondary structure, which could explain transcriptional termination within the 3' LTR but not in the 5' LTR. The nucleotide sequence of the provirus contains three large open reading frames, which are capable of coding for proteins of 48,000, 99,000, and 54,000 daltons. The three open frames are in this order from the 5' end of the viral genome and the predicted 48,000-dalton polypeptide is a precursor of gag proteins, because it has an identical amino acid sequence to that of the NH2 terminus of human T-cell leukemia virus (HTLV) p24. The open frames coding for 99,000- and 54,000-dalton polypeptides are thought to be the pol and env genes, respectively. On the 3' side of these three open frames, the ATLV sequence has four smaller open frames in various phases; these frames may code for 10,000-, 11,000-, 12,000-, and 27,000-dalton polypeptides. Although one or some of these open frames could be the transforming gene of this virus, in preliminary analysis, DNA of this region has no homology with the normal human genome.
Corticotropin/lipotropin common precursor-like material in normal rat extrapituitary tissues.
Authors: Saito E , Odell W D
Published: May 2006

A Mr 26,000 corticotropin (ACTH)-like material is present in glacial acetic acid extracts of all normal rat extrapituitary tissues. In the present study, beta-melanotropin (beta-MSH) immunoactivity was detected in glacial acetic acid extracts of normal rat extrapituitary tissues. beta-MSH immunoactivity was also present in all extracts (mean +/- SEM, fmol/mg of protein): brain, 71.0 +/- 16.3; stomach, 11.5 +/- 1.6; kidney, 8.9 +/- 0.8; colon, 8.2 +/- 1.1; small intestine, 6.5 +/- 1.1; liver, 4.3 +/- 0.5; and heart, 3.2 +/- 0.5. Except in brain extracts, beta-MSH and ACTH immunoactivities of tissue extracts were strongly correlated to each other (r = 0.79; n = 42). When tissue extracts (except brain) were passed through a Sephadex G-75 (superfine) column, ACTH and beta-MSH immunoactivities were eluted in a single peak corresponding to Mr 26,000. In contrast, for brain extracts, the MrS of major peaks of ACTH and beta-MSH immunoactivities were 4,500 and 8,000, respectively; a smaller peak of Mr 26,000 ACTH/beta-MSH-like material was also eluted. Specific anti-ACTH immunocolumns, which did not bind purified synthetic beta-MSH, adsorbed both ACTH and beta-MSH immunoactivities of all tissue extracts except those of brain. One-third of the beta-MSH immunoactivity in brain extracts adsorbed to the anti-ACTH immunocolumn, but two-thirds of beta-MSH immunoactivity passed through the column. We conclude that ACTH and beta-MSH immunoactivities are present in all normal rat extrapituitary tissues and exist in most tissues on the same molecule. This Mr 26,000 molecule is closely related to the pituitary ACTH/beta-lipotropin common precursor.
Structure of iron superoxide dismutase from Pseudomonas ovalis at 2.9-A resolution.
Authors: Ringe D , Petsko G A , Yamakura F , Suzuki K , Ohmori D
Published: May 2006

The three-dimensional structure of the iron-containing superoxide dismutase (EC 1.15.1.1) from Pseudomonas ovalis has been determined at 2.9-A resolution by the method of multiple isomorphous replacement. The molecule is a dimer of two identical subunits with the iron atom per monomer. The conformation of the enzyme is completely different from that of the eukaryotic copper-zinc superoxide dismutase. Each subunit consists of about 50% alpha-helix plus three strands of antiparallel pleated sheet. The iron atoms are coordinated by four protein ligands, one of which is the side-chain of histidine-26. Crystals of complexes with the inhibitors azide or fluoride are considerably more resistant to irradiation than those of the free enzyme. The structure of the apoprotein is identical to that of the iron-containing molecule.
Occurrence of a hormone-sensitive inhibitory coupling component of the adenylate cyclase in S49 lymphoma cyc- variants.
Authors: Jakobs K H , Schultz G
Published: May 2006

The influence of somatostatin was studied on cyclic AMP levels and adenylate cyclase activity in cyc- variants of S49 lymphoma cells. These cells are deficient in the guanine nucleotide site that mediates hormone-induced adenylate cyclase stimulation, but their cyclase can be stimulated by forskolin. Somatostatin maximally decreased the 30 microM forskolin-stimulated cyclic AMP levels by 35%. Half-maximal suppression occurred at about 0.1 nM somatostatin. Somatostatin (up to 1 microM) had no effect on the 100 microM forskolin-stimulated adenylate cyclase activity in cyc- membrane preparations when guanine nucleotides were not present. In the presence of GTP, however, which by itself caused a small decrease in activity, somatostatin maximally inhibited the enzyme by 20-25%. GTP was half-maximally effective at 0.1 microM, and half-maximal inhibition by somatostatin was observed at 0.1- 1 nM. In the presence of the stable GTP analog guanosine 5'-O-(3-thiotriphosphate) (1 microM), which decreased the stimulated activity by about 40% after a short lag period, somatostatin (1 microM) did not cause a further decrease in final activity but reduced the lag period by about 50%. The data indicate that membranes of cyc- variants contain a regulatory site that mediates both guanine nucleotide and hormone-induced inhibition of the adenylate cyclase and suggest that the mechanisms of activation and inactivation of this inhibitory site are similar to those of the stimulatory component missing in cyc-membranes.
Two levels of regulation of beta-interferon gene expression in human cells.
Authors: Raj N B , Pitha P M
Published: May 2006

We cloned alpha- and beta-interferon cDNA and used them as specific probes to determine the relative levels of interferon mRNA in human fibroblasts cells induced with poly(rI).poly(rC) or Newcastle disease virus to synthesize interferon. Both inducers activated only the beta-interferon gene; however, the half life of beta-interferon mRNA in cells induced with virus was substantially longer than in poly(rI).poly(rC)-induced cells. The transcription rate of beta-interferon RNA sequences was examined in nuclei isolated from poly(rI).poly(rC)-induced cells; it was found that the induction leads to transcriptional activation of the beta-interferon gene and that the shutoff period when no interferon synthesis or cytoplasmic betamRNA are detected. Thus, the synthesis of beta interferon in poly(rI).poly-(rC)-induced human fibroblasts is controlled both by activation of transcription of the beta-interferon gene and by alteration of the beta-interferon mRNA stability.
Comparison of maxicircle DNAs of Leishmania tarentolae and Trypanosoma brucei.
Authors: Muhich M L , Simpson L , Simpson A M
Published: May 2006

The conserved portions of the maxicircle DNAs of Leishmania tarentolae and Trypanosoma brucei are organized in a basically colinear manner over a 15- to 17-kilobase region that is interrupted by two small less-homologous sequences. The most highly conserved regions are those encoding the 9S and 12S genes. An approximately 12-kilobase region directly upstream of the 12S gene in the L. tarentolae maxicircle showed no sequence homology with the T. brucei maxicircle and also was not transcribed. An approximately 6-kilobase region in the T. brucei maxicircle in the same relative location also showed no sequence homology with the L. tarentolae maxicircle. We propose that evolution of maxicircle DNA occurs mainly within this "divergent region."
Binding of factors IX and IXa to cultured vascular endothelial cells.
Authors: Stern D M , Drillings M , Nossel H L , Hurlet-Jensen A , LaGamma K S , Owen J
Published: May 2006

Factor IX and its activated form IXa have been found to bind to confluent cultured bovine aortic and human umbilical vein endothelial cells. Binding of bovine factors IX and IXa to the bovine endothelial cells was saturable and specific and reached a plateau in 75 min at 4 degrees C and 30 min at 37 degrees C. Binding was half-maximal at a total factor IX or IXa concentration of 2.3 +/- 0.2 nM. At 4 degrees C, a maximum of 42 fmol of tritiated factor IX or IXa bound to 10(6) cells (an average of 20,000 molecules per cell). The binding of tritiated factor IX or IXa was inhibited by excess unlabeled factor IX or IXa but not by factor X, prothrombin, or thrombin. Competition studies indicated that factors IX and IXa interacted with the same site. Binding was reversible, with 50% of the specifically bound factor IX or IXa eluted in 40 min by a 400-fold excess of unlabeled protein. Specific binding required Ca2+ with half-maximal binding at 1.2 mM CaCl2. Factor IXa bound to the cells was tested for procoagulant activity in a clotting assay with factor IX-deficient plasma, cephalin, and CaCl2. Cell-bound factor IXa was at least 3-fold more active than was factor IXa in solution. The retention of procoagulant activity by cell surface-bound factor IXa provides a mechanism for the localization of clot-promoting activity.
Mevinolin and colestipol stimulate receptor-mediated clearance of low density lipoprotein from plasma in familial hypercholesterolemia heterozygotes.
Authors: Bilheimer D W , Grundy S M , Brown M S , Goldstein J L
Published: May 2006

In subject with heterozygous familial hypercholesterolemia (FH), a 50% deficiency of receptors for plasma low density lipoprotein (LDL) impairs the removal of LDL from plasma and produces hypercholesterolemia. In these patients mevinolin, an inhibitor of 3-hydroxy-3-methylglutaryl-CoA reductase, blocks cholesterol synthesis and lowers plasma LDL levels. To determine the mechanism for the LDL-lowering effect, we administered 131I-labeled LDL intravenously to six FH heterozygotes before and during treatment with mevinolin and calculated the apparent fractional catabolic rate (FCR) and synthetic rate for LDL. After mevinolin treatment, the mean plasma LDL-cholesterol level declined from 262 to 191 mg/dl (27% decrease), the mean FCR for 131I-labeled LDL increased from 0.30 to 0.41 pools per day (37% increase), and the mean calculated synthetic rate for LDL-protein did not change significantly. In one of FH heterozygote with an ileal bypass and in another who received colestipol, the addition of mevinolin caused, respectively, a 41% and 60% decrease in plasma LDL levels and a 60% and 100% increase in the FCR for plasma LDL. The contribution of receptor-dependent pathways to the FCR for plasma LDL was estimated in three FH heterozygotes by simultaneous measurements of the FCR for native 131I-labeled LDL and 125I-labeled glucosylated LDL, which does not bind to LDL receptors. Whereas the removal rate for native LDL increased after mevinolin treatment, the removal rate for glucosylated LDL did not change. The current data suggest that mevinolin alone or mevinolin plus bile acid depletion (i.e., ileal bypass or colestipol therapy) decreases plasma LDL levels primarily by raising the number of LDL receptors and, thus, enhancing the removal of LDL from plasma.
Antigenic variation of Plasmodium knowlesi malaria: identification of the variant antigen on infected erythrocytes.
Authors: Howard R J , Barnwell J W , Kao V
Published: May 2006

Erythrocytes infected with mature asexual stages of Plasmodium knowlesi express a new surface antigen such that rhesus monkey antisera specifically agglutinate these cells. Cloned parasites can express different antigenic variants of this antigen. The variant antigen has been identified by comparison of the surface membrane antigens of a clone and of an antigenic variant of that clone of different agglutination phenotype. After lactoperoxidase labeling, 125I-labeled proteins of Mrs 210,000 and 190,000 with one clone and of Mrs 205,000 with the antigenic variant were only immunoprecipitated by antisera containing homologous anti-variant antibody. Antigens of the same Mr from each clone were labeled by [35S]methionine incorporation during parasite growth and also specifically immunoprecipitated only by agglutinating antisera. Therefore, the variant antigens are malarial proteins rather than modified host proteins.
Pertussis toxin substrate, the putative Ni component of adenylyl cyclases, is an alpha beta heterodimer regulated by guanine nucleotide and magnesium.
Authors: Codina J , Hildebrandt J , Iyengar R , Birnbaumer L , Sekura R D , Manclark C R
Published: May 2006

The final step in a scheme for the purification of the guanine nucleotide- and Mg2+-binding stimulatory regulatory component (Ns) of adenylyl cyclase [adenylate cyclase; ATP pyrophosphate-lyase (cyclizing), EC 4.6.1.1] from human erythrocyte membranes involves chromatography over hydroxylapatite (HAP) which yields two fractions. The first fraction (HAP I) contains predominantly two peptides that, upon sodium dodecyl sulfate/polyacrylamide gel electrophoresis, migrate with Mr values of 39,000 and 35,000. The second fraction (HAP II) contains predominantly Ns formed of two peptides of Mr 42,000 and 35,000. The HAP I, Mr 39,000 peptide is shown to be a substrate for the ADP-ribosylating toxin of Bordetella pertussis (pertussis toxin). Upon sucrose density gradient centrifugation, both the Mr 39,000 and the Mr 35,000 peptides of HAP I migrate at about 4 S. Treatment of HAP I with guanine nucleotide and Mg2+ prior to centrifugation results in a coordinated change in the migration of both peptides to 2 S. It is postulated that HAP I contains an alpha beta heterodimeric protein composed of an alpha subunit of Mr 39,000 and a beta subunit of Mr 35,000. Further, this protein dissociates under the influence of guanine nucleotides and Mg2+ into its individual alpha and beta subunits. Because previous studies have shown that treatment of cells and cell membranes with pertussis toxin results in attenuation of the effects of hormones that inhibit adenylyl cyclase activity, and because this effect correlates with the ADP-ribosylation of a Mr approximately equal to 40,000 peptide, we believe that we have purified a guanine nucleotide- and Mg2+-binding inhibitory regulatory component of adenylyl cyclases--i.e., the Ni.
Activation of calcineurin by limited proteolysis.
Authors: Manalan A S , Klee C B
Published: May 2006

Calcineurin, a heterodimer of calcineurin B, a 19,000 Mr Ca2+-binding subunit, and calcineurin A, a 61,000 Mr calmodulin-binding subunit, was previously proposed to be a calmodulin- and Ca2+-regulated protein phosphatase. Like other calmodulin-stimulated enzymes, calcineurin can be activated and rendered calmodulin- and Ca2+-independent by limited proteolysis. By glycerol gradient centrifugation, the native enzyme has a s20,w of 4.5 S in EGTA and 5 S in the presence of Ca2+-calmodulin. Under the same conditions, the s20,w of the trypsin-activated enzyme (4.3 S) is not affected by Ca2+ and calmodulin. The trypsin-treated enzyme is a heterodimer of calcineurin B and a 45,000 Mr fragment of calcineurin A that has lost its ability to interact with calmodulin. Phosphatase activity sediments with calcineurin or its proteolytic fragments, providing further evidence that calcineurin is indeed a protein phosphatase. Calmodulin protects calcineurin against tryptic digestion; proteolysis occurs more slowly, yielding fragments with Mr 57,000, 55,000, and 54,000 that have preserved their ability to interact with calmodulin. After trypsin treatment in the presence of calmodulin, the protein phosphatase activity of calcineurin is still regulated by calmodulin. Prolonged trypsin treatment in the presence of calmodulin produces a 46,000 Mr fragment. Unlike the fragments generated in the absence of calmodulin, this 46,000 Mr fragment still interacts weakly with calmodulin. Thus, calcineurin, like other calmodulin-regulated enzymes, consists of a catalytic domain resistant to proteolysis and a calmodulin-binding regulatory domain susceptible to protease action in the absence of calmodulin but not in its presence. In the absence of calmodulin, the regulatory domain exerts an inhibitory effect on the catalytic domain; the inhibition is relieved upon calmodulin binding to or tryptic degradation of the regulatory domain.
Expression of the large T protein of polyoma virus promotes the establishment in culture of "normal" rodent fibroblast cell lines.
Authors: Rassoulzadegan M , Naghashfar Z , Cowie A , Carr A , Grisoni M , Kamen R , Cuzin F
Published: May 2006

Transfer into mouse and rat embryo fibroblasts in primary culture of cloned polyoma virus genes encoding only the large T protein led to the establishment of flat colonies in sparse subcultures at a frequency equal to that of transformation by wild-type virus. Cell lines could be derived from such colonies and maintained in culture for large numbers of generations without entering crisis. They exhibited a normal phenotype, by the criteria of growth on plastic to a low saturation density and of anchorage dependency. However, they required a lower serum concentration for growth than spontaneously established 3T3 cells. Similar results were obtained after transfer of recombinant DNA molecules encoding only the amino-terminal 40% of the large T protein, suggesting that this "immortalization" function corresponds to the activity of an amino-terminal domain of the protein. Immunoprecipitation analysis of T antigens in cell lines established after transfer of the full-size and of the truncated large T genes demonstrated the expression of the full-size large T protein and of a Mr 40,000 antigen expressed from the amino-terminal part of the gene, respectively. After transfer of a "large T only" plasmid that carries a tsa mutation, cell lines were established at 33 degrees C with the same efficiency as with the wild-type large T gene, but their growth was arrested after a shift to 40 degrees C, with a progressive loss in cell viability. This result indicates a continuous requirement for a large T function in the maintenance of "immortality."
Idiotypic manipulation in mice: BALB/c mice can express the crossreactive idiotype of A/J mice.
Authors: Moser M , Leo O , Hiernaux J , Urbain J
Published: May 2006

The response of A/J mice to arsonate-coupled keyhole limpet hemocyanin is characterized by a crossreactive idiotype (CRIA). CRIA+ antibodies are restricted to the Igh-Ic haplotype and are never expressed in BALB/c mice after immunization with antigen. Studies at the DNA level suggest that the gene encoding the CRIA heavy chain in A/J mice is probably absent in the genome of BALB/c mice. Despite this, using the immunization cascade tool, we have been able to induce the expression of CRIA+ antibodies in BALB/c mice. These studies led to an apparent paradox, whose understanding will provide new insights into the regulatory mechanisms of the immune system. We suggest that clones secreting CRIA-like Igs in BALB/c mice are "somatic variants" that could arise from gene conversion events.
Mammalian nitrate biosynthesis: incorporation of 15NH3 into nitrate is enhanced by endotoxin treatment.
Authors: Wagner D A , Young V R , Tannenbaum S R
Published: May 2006

Incorporation of an oral dose of [15N]ammonium acetate into urinary [15N]nitrate has been demonstrated in the rat. Investigation of the regulation of nitrate synthesis has shown that Escherichia coli lipopolysaccharide potently stimulates urinary nitrate excretion (9-fold increase). It was further shown that the enhanced rate of nitrate excretion by lipopolysaccharide was due not to a reduction in nitrate metabolic loss but rather to an increased rate of synthesis. This conclusion was based on finding a proportionally increased incorporation of [15N]ammonium into nitrate nitrogen with lipopolysaccharide treatment. Nitrate biosynthesis was also increased by intraperitoneal injection of carrageenan and subcutaneous injection of turpentine. It is proposed that the pathway of nitrate biosynthesis may be the result of oxidation of reduced nitrogen compounds by oxygen radicals generated by an activated reticuloendothelial system.
Attenuation of Mach bands by adjacent stimuli.
Authors: Ratliff F , Milkman N , Rennert N
Published: May 2006

Pronounced bright and dark bands are seen at the bright and dark edges of half-shadows and similar distributions of illumination. These are the so-called Mach bands. A pair of vertical Mach bands was generated with a ramp pattern in the central strip of a horizontal tripartite oscilloscope display. This pattern consisted of two uniform fields (one of low luminance, one of high luminance) joined by a gradient of uniform slope. The upper and lower strips were uniform throughout. A coupled pair of pointers could be displayed in these two strips and adjusted by the observer to match the apparent location and width of either of the Mach bands in the central strip. Insertion of a vertical bar in the central strip nearby and on either side of the ramp attenuates the corresponding Mach band. The closer the bar is to the Mach band, the stronger the attenuation. The attenuation is nearly independent of the sign of the contrast of the bar, but it does depend upon the magnitude and sharpness of the contrast. Also, the attenuation is independent of the width of the bar; a narrow line is as effective as a broad bar of the same contrast. No net luminance change is required; a bipolar stimulus with equal parts above and below the mean is as effective as a monopolar stimulus. These results point to two competing physiological mechanisms with different spatial sensitivities--one that generates Mach bands and one that attenuates them.
The second law as a selection principle: The microscopic theory of dissipative processes in quantum systems
Authors: Prigogine I. , George Cl.
Published: May 2006

The second law of thermodynamics, for quantum systems, is formulated, on the microscopic level. As for classical systems, such a formulation is only possible when specific conditions are satisfied (continuous spectrum, nonvanishing of the collision operator, etc.). The unitary dynamical group can then be mapped into two contractive semigroups, reaching equilibrium either for
t
→ +∞ or for
t
→ -∞. The second law appears as a symmetry-breaking selection principle, limiting the observables and density functions to the class that tends to thermodynamic equilibrium in the
future
(for
t
→ +∞). The physical content of the dynamical structure is now displayed in terms of the appropriate semigroup, which is realized through a nonunitary transformation. The superposition principle of quantum mechanics has to be reconsidered as irreversible processes transform pure states into mixtures and unitary transformations are limited by the requirement that entropy remains invariant. In the semigroup representation, interacting fields lead to units that behave incoherently at equilibrium. Inversely, nonequilibrium constraints introduce correlations between these units.
Molecular requirements for B-lymphocyte activation by Escherichia coli lipopolysaccharide.
Authors: Raetz C R , Purcell S , Takayama K
Published: May 2006

Certain Escherichia coli mutants altered in phosphatidylglycerol metabolism accumulate fatty acyl derivatives of glucosamine 1-phosphate. Especially prominent is 2,3-diacylglucosamine 1-phosphate (previously designated lipid X), which may be an early precursor of lipid A. We have examined the activity of lipid X (Mr = 711.9) and several related compounds as mitogens towards mouse lymphocytes. As judged by labeling with [methyl-3H]thymidine, lipid X is mitogenic, and it mimics the properties of lipopolysaccharide and lipid A. The following evidence suggests that lipid X exerts its effects by a route similar to that of lipopolysaccharide: (i) lymphocytes from C3H/HeJ mice, which are unresponsive to lipopolysaccharide, are also not stimulated by lipid X; (ii) polymyxin B abrogates lymphocyte stimulation by lipid X; and (iii) lipid X induces the proliferation and maturation of lymphocytes to antibody-producing plaque-forming cells. Selective removal of the ester-linked hydroxymyristate moiety at position 3 totally abolishes mitogenic activity. Other phospholipids, such as phosphatidic acid, CDP-diglyceride, phosphatidylcholine, and lysophosphatidylcholine, have no activity as mitogens. If lipid X and lipid A induce by common mechanism(s) B-lymphocyte proliferation, then it follows from structural comparison that the reducing-end subunit of lipid A is the minimal structural requirement for this activity. Because the structure of lipid X is completely defined, biochemical and pharmacological dissection of B-cell activation by lipopolysaccharide should now be possible.
Radioprotection of human lymphoid cells by exogenously supplied glutathione is mediated by gamma-glutamyl transpeptidase.
Authors: Jensen G L , Meister A
Published: May 2006

Human lymphoid cells depleted of glutathione by treatment with buthionine sulfoximine, a specific inhibitor of gamma-glutamylcysteine synthetase, may be partially repleted by adding glutathione in the medium. The mechanism of repletion involves the action of gamma-glutamyl transpeptidase on exogenous glutathione, transport of products of glutathione metabolism, and intracellular synthesis of glutathione. Lymphoid cells, previously shown to export glutathione at rates proportional to intracellular glutathione levels, do not take up intact glutathione to an appreciable extent, even under conditions of marked glutathione deficiency. The role of glutathione in radioprotection was examined by subjecting cells to gamma-radiation after modification of cellular glutathione levels. Glutathione-depleted cells exhibited increased radiosensitivity under aerobic conditions, as compared to the nondepleted controls. Partial repletion of cellular glutathione prior to irradiation led to radiosensitivity comparable to nondepleted controls. Cells were not protected by suspension in media containing glutathione just prior to irradiation; thus, protection appears to require intracellular glutathione.
Intrinsic resistance to the lethal effects of x-irradiation in insect and arachnid cells
Authors: Koval Thomas M.
Published: May 2006

Twelve cell lines representing 10 genera of three orders (Diptera, Lepidoptera, and Orthoptera) of the class Insecta and one cell line (Acarina) from the class Arachnida were examined to discern their sensitivity to the lethal effects of x-irradiation. Radiosensitivity was measured by a combination of colony formation and population growth curve techniques. Each of these arthropod cell lines is significantly more radioresistant than mammalian cells, though the degree of resistance varies greatly with order. Dipteran cells are 3 to 9 times and lepidopteran cells 52 to 104 times more radioresistant than mammalian cells. Orthopteran and acarine cells are intermediate in radiosensitivity between dipteran and lepidopteran cells. These cells, especially the lepidopteran, should be valuable in determining the molecular nature of repair mechanisms that result in resistance to ionizing radiation.
Two small RNAs encoded by Epstein-Barr virus can functionally substitute for the virus-associated RNAs in the lytic growth of adenovirus 5.
Authors: Bhat R A , Thimmappaya B
Published: May 2006

Adenovirus (Ad) serotypes 2 and 5 synthesize large amounts of two low molecular weight RNAs designated virus-associated (VA) 1 and 2. Recently, genetic and biochemical approaches have been used to show that Ad2 VA1 RNA is required for efficient translation of viral mRNAs produced at late times after infection. Primate cells harboring the Epstein-Barr virus genome (EBV) synthesize large amounts of two low molecular weight RNAs: EBER1 and EBER2. Striking similarities of gene organization have been noted between the genes coding for Ad5 VA RNAs and the EBV EBERs [Rosa, M. D., Gottlieb, E., Lerner, M. R. & Steitz, J. A. (1981) Mol. Cell. Biol. 1, 785-796]. To examine whether the EBRs can functionally substitute for the VA RNAs for the lytic growth of Ad5, we have constructed an Ad5 substitution mutant in which the two VA RNA genes have been deleted and replaced by an EBV DNA segment coding for the two EBERs. The resulting Ad5 mutant synthesizes large amounts of the EBERs and is viable. Thus, two small RNAs of EBV origin, having primary and secondary structures different from those of the VA RNAs, can functionally substitute for the VA RNAs in the lytic growth of Ad5. These results are discussed in the context of mechanism of function of the VA RNAs.
Pancreatic epithelium is permeable to sucrose and inulin across secretory cells.
Authors: Mélèse T , Rothman S S
Published: May 2006

The pancreatic epithelium is permeable to both sucrose and inulin and becomes more permeable when protein secretion by the gland is stimulated. Because these molecules are not thought to enter cells, it has been assumed that their movement across the epithelium from interstitial to ductal fluid, as well as the increase in that flux that is observed during augmented protein secretion, is due to their passage through paracellular shunts. In the present experiments we have considered the alternative possibility that sucrose and inulin travel through the cells of the secretory epithelium instead of, or in addition to, their passage through paracellular shunts. The data support this view and suggest that the pancreas is unusually permeable to water soluble molecules of substantial size by means of a transcellular pathway.
Energy depletion retards the loss of membrane transport during reticulocyte maturation.
Authors: Weigensberg A M , Blostein R
Published: May 2006

The effect of metabolic depletion on the maturation-associated loss of membrane functions has been studied by using sheep reticulocytes incubated in vitro at 37 degrees C for periods up to 41 hr. ATP was either maintained with glucose, adenosine plus inosine, or depleted with 2-deoxyglucose plus arsenate. Two membrane transport systems were studied: Na+-dependent glycine transport activity and the sodium pump, estimated from measurements of the number of [3H]ouabain binding sites per cell. Both transport systems were decreased during maturation. However, the decrease was much less in ATP-depleted cells compared to ATP-replete cells. It is concluded that the loss of certain functions during reticulocyte maturation is retarded by metabolic depletion.
Cultural versus genetic adaptation.
Authors: Cavalli-Sforza L L , Feldman M W
Published: May 2006

We analyze the problem of the competition between genetic and cultural adaptation, testing various types of cultural transmission models versus a genetic (haploid) transmission model. With a vertical (parent-to-child) plus an infectious (oblique) cultural transmission, genetic adaptation always prevails, although its relative increase may be slow and polymorphism may persist for some time. Only under very special conditions of cultural transmission may a permanent polymorphism in which the two types of adaptation are represented be reached. There may, however, be an overall evolutionary advantage to a flexible mechanism of cultural transmission that allows adaptation to new situations for which no genetic mutants are available.
cDNA clone spanning the alpha-gamma subunit junction in the precursor of the murine fourth complement component (C4).
Authors: Ogata R T , Shreffler D C , Sepich D S , Lilly S P
Published: May 2006

cDNA clones carrying parts of murine fourth complement component (C4, serum substance protein) mRNA sequences have been identified by differential hybridization to mRNA from a high C4-producing strain, B10.WR, and a congeneic low C4 strain, B10.BR, followed by hybrid-selected translation and DNA sequence analysis. One clone, pMLC4/w7-2, encodes an open amino acid reading frame that includes four tandem arginine residues immediately preceding a sequence 85% homologous with the NH2-terminal sequence of the human C4 gamma-chain. The amino acid composition of the predicted sequence upstream of the tandem arginines matches quite closely with the composition of a similar sized peptide at the COOH terminus of the human C4 alpha chain. The latter result raises questions regarding the nature and extent of plasma-mediated postsynthetic processing of the C4 alpha-chain COOH terminus. The results also demonstrate that strain differences in plasma C4 levels (low C4 vs. high C4) reflect differences in steady-state levels of liver C4 mRNA in these strains.
Monoclonal antibodies to rat Na+,K+-ATPase block enzymatic activity.
Authors: Schenk D B , Leffert H L
Published: May 2006

A panel of nine mouse monoclonal antibodies has been prepared against purified preparations of rat kidney Na+,K+-ATPase (EC 3.6.1.3). Selection for specific antibody was based upon the ability of crude hybridoma fluids to inhibit Na+-ATPase activity (using luciferase-linked ATPase assays) and upon antibody binding to both the purified kidney membrane enzyme and to glutaraldehyde-fixed hepatocytes by using standard enzyme-linked immunoadsorbent assays. After immunoaffinity purification, two of the antibodies (both of the IgG1 subclass) fully inhibit kidney and liver membrane Na+,K+-ATPase activity with Ki (apparent) values of 30 nM ("9-A5") and 600 nM ("9-B1"). Immunoblots demonstrate directly that three different 125I-labeled antibodies (6-4, 9-A5, and 9-B1) bind predominantly to a 94,000 Mr protein that comigrates in NaDodSO4/polyacrylamide gels with the fluorescein isothiocyanate-labeled alpha subunit of the Na+,K+-ATPase. Indirect immunofluorescence studies with these antibodies on paraformaldehyde-fixed liver slices reveal staining patterns congruent with bile canalicular membrane domains. These results together suggest that the antibodies exert inhibitory effects by recognizing alpha subunits of both liver and kidney Na+ pumps in their native conformations.
Crystal structures of the active site in specifically metal-depleted and cobalt-substituted horse liver alcohol dehydrogenase derivatives.
Authors: Schneider G , Eklund H , Cedergren-Zeppezauer E , Zeppezauer M
Published: May 2006

Two derivatives of horse liver alcohol dehydrogenase (LADH) in which the active site is specifically metal-depleted [H4Zn(n)2LADH] or specifically Co-substituted [Co(c)2-Zn(n)2LADH] have been studied by crystallographic methods. (In these formulae, "n" identifies the noncatalytic zinc ion and "c" identifies the catalytic metal ion.) X-ray data were collected for H4Zn(n)2LADH to 2.7-A resolution and for Co(c)2Zn(n)2LADH to 2.4-A resolution. Difference Fourier maps demonstrate clearly that the catalytic zinc ions are removed in H4Zn(n)2LADH, whereas the noncatalytic zinc ions are still present. A 2.5-A shift in the sulphur position of cysteine-46 and a slight torsion of the imidazole ring of histidine-67 are the only changes in the protein structure that could be detected when compared to the native zinc enzyme. The structure of Co(c)2Zn(n)2LADH is essentially the same as that of the native enzyme. Each cobalt ion is bound to the ligands cysteine-46, cysteine-174, and histidine-67 and to a water molecule in a distorted tetrahedral geometry. A slight change in the position of histidine-67 was found. No further structural changes could be observed in the protein.
Transformation of human umbilical cord blood T cells by human T-cell leukemia/lymphoma virus.
Authors: Popovic M , Lange-Wantzin G , Sarin P S , Mann D , Gallo R C
Published: May 2006

Several isolates of human T-cell leukemia/lymphoma virus (HTLV) were transmitted to normal human T cells obtained from the umbilical cord blood of newborns. T cells from seven specimens were immortalized by infection with different HTLV isolates and their properties were compared with those of activated uninfected normal T cells grown in the presence of T-cell growth factor (TCGF) and with those of HTLV-positive neoplastic T-cell lines derived from patients with T-cell malignancies. The HTLV-infected cells generally belonged to a class of mature T cells (OKT4+ and Leu 3A+) and differed from the normal uninfected cells in that they could be propagated in culture indefinitely; possessed altered morphology, including convoluted nuclei and some bi- and multinucleated giant cells; formed large clumps in culture; demonstrated a diminished requirement for TCGF; had an increased density of TCGF receptors; often became completely independent of exogenous TCGF; and expressed HLA-DR determinants. These properties of the HTLV-infected cord blood T cells contrasted to those of uncultured cord blood T cells and of cord blood cells stimulated with mitogen and grown with TCGF but resembled the characteristics of T-cell lines established previously from patients with HTLV-associated T-cell malignancies. This in vitro system offers a unique opportunity to study the basic mechanism involved in abnormal growth and neoplastic transformation of a specific class of human T cells.
Inducible transcription of five globin genes in K562 human leukemia cells.
Authors: Dean A , Ley T J , Humphries R K , Fordis M , Schechter A N
Published: May 2006

We studied the abundance and structure of globin mRNAs present in K562 cells both before and after induction of hemoglobin synthesis by hemin. In vitro translation of poly(A)+ RNA from K562 cells generated protein products corresponding to alpha, A gamma-, G gamma-, epsilon-, and zeta-globin mRNAs. Individual globin mRNAs increased 1.5- to 3-fold after induction. Similar results were obtained by measuring steady-state mRNA concentrations of induced and uninduced cells by using S1 nuclease mapping. Globin gene transcripts were correctly initiated and processed. In addition, S1 nuclease analysis revealed the presence of delta-globin mRNA in both control and induced cells. A small percentage of delta-globin transcripts appeared to be initiated upstream from the normal initiation site. beta-Globin mRNA was not detected in any studies. The results (i) suggest that hemin induction of K562 cells is mediated at a transcriptional level and (ii) reveal the dissociation of delta- and beta-globin gene expression in K562 cells compared with normal erythroid cells.
Rapid selective effects by a growth inhibitor and epidermal growth factor on the incorporation of [35S]methionine into proteins secreted by African green monkey (BSC-1) cells.
Authors: Nilsen-Hamilton M , Holley R W
Published: May 2006

Confluent African green monkey kidney (BSC-1) cells secrete a protein (Mr approximately equal to 24,000) that inhibits DNA synthesis and growth of the same cells. Using [35S]methionine to metabolically label proteins, we have found that this growth inhibitor selectively induces the BSC-1 cells to synthesize and secrete another protein with a relative Mr of 48,000 on NaDodSO4/polyacrylamide gels. We have called this protein "inhibitor-inducible protein" (IIP48). The maximal increase in rate of labeling of IIP48 due to treatment with the growth inhibitor averages 12-fold over the control. IIP48 is an N-glycosidically linked glycoprotein, and it is not a major intracellular protein. This protein is maximally induced within 4 to 6 hr of adding the growth inhibitor to the cells. This is an early response of these cells to the growth inhibitor and may represent a primary response to the growth inhibitor. Epidermal growth factor (EGF) increases the rate of labeling of three other secreted proteins (MrS 28,000, 59,000, and 61,000), which we have called "mitogen-inducible proteins" (MIP28, MIP59, and MIP61). The specific effects of both EGF and the growth inhibitor on the secreted levels of these proteins are inhibited if actinomycin D is added with the growth effectors. Thus, RNA synthesis appears necessary for the inductions. EGF and the growth inhibitor induce these secreted proteins by independent and noninteracting pathways.
Molecular evolution of the human adult alpha-globin-like gene region: insertion and deletion of Alu family repeats and non-Alu DNA sequences.
Authors: Hess J F , Fox M , Schmid C , Shen C K
Published: May 2006

Previous heteroduplex studies have revealed extensive sequence homology between the two human adult alpha-globin-like genes (alpha 2 and alpha 1) and their flanking regions. These homologous regions, which are interrupted by two blocks of nonhomology, each span approximately 4 kilobases [Lauer, J., Shen, C.-K. J. & Maniatis, T. (1980) Cell 20, 119-130]. We have determined 3 kilobases of DNA sequences within and flanking the nonhomologous blocks of these two tandem duplication units. A total of three Alu family repeats has been identified. Two of them are approximately 300 base pairs long and define the 3' ends of the first homology blocks. The third Alu family member is a 600-base-pair-long sequence consisting of two monomeric Alu members arranged in a head-to-tail fashion. It is located in the 3' portion of the first block of nonhomology in alpha 2-gene-containing unit. We present direct evidence that this dimeric Alu sequence was inserted at a staggered break. The second nonhomology block is the result of insertion or deletion of a 224-base-pair sequence. From these data and the calculation of sequence divergence, we propose a history for the evolution of the human adult alpha-globin-like gene region. We also suggest that DNA insertion elements may disrupt gene correction processes in the two duplication units containing alpha 2- and alpha 1-globin genes.
Distribution and mobility of murine histocompatibility H-2Kk antigen in the cytoplasmic membrane.
Authors: Damjanovich S , Trón L , Szöllösi J , Zidovetzki R , Vaz W L , Regateiro F , Arndt-Jovin D J , Jovin T M
Published: May 2006

The topographical distributions and mobilities of the murine histocompatibility antigen H-2Kk and of concanavalin A (Con A) binding sites have been studied on a murine lymphoma cell line. The spatial distribution of H-2Kk antigens, the average distance between H-2Kk antigens and Con A binding sites, and the separation of different determinants on the H-2Kk antigen itself were determined by using fluorescence resonance energy-transfer measurements with a dual-laser flow sorter. From the lack of energy transfer between bound monoclonal anti-H-2Kk antibodies conjugated with fluorescein (donor) and rhodamine (acceptor), we conclude that the H-2Kk antigen exists without appreciable clustering on the cell surface. Substantial energy transfer between appropriately labeled Con A and antibodies bound to the H-2Kk antigen shows that the two populations are interspersed. Donor/acceptor pairs of monoclonal antibodies binding to different determinants on the same H-2Kk antigen exhibited a degree of energy transfer indicative of a mean separation of 8.6 nm between the sites. Time-resolved phosphorescence anisotropy measurements with anti-H-2Kk antibodies labeled with eosin or erythrosin yielded rotational mobility information for the antigen-antibody complexes on the cell membrane. The rotational correlation time of 10-20 mus and the finite residual anisotropy are compatible with an uniaxial mode of rotation of monomeric antigen around its transmembrane portion and, thus, provide additional evidence for an unclustered distribution. Capping by rabbit anti-mouse IgG immobilized the antigen-antibody complex. Fluorescence recovery after photobleaching was used to calculate an apparent lateral diffusion coefficient of 5 +/- 3 X 10(-10) cm2 . s-1 for the H-2Kk antigen labeled with fluoresceinated IgG or its corresponding Fab fragment.
Acquired alpha-thalassemia in preleukemia is due to decreased expression of all four alpha-globin genes.
Authors: Anagnou N P , Ley T J , Chesbro B , Wright G , Kitchens C , Liebhaber S , Nienhuis A W , Deisseroth A B
Published: May 2006

A somatic mutation(s), acquired during the evolution of preleukemia in a 75-year-old Caucasian male of North European origin, resulted in a marked decrease in alpha-globin mRNA. The small amount of alpha-globin mRNA present in bone marrow cells was normally processed, had a normal (alpha 1/alpha 2)-globin mRNA ratio, and was translated normally. No detectable zeta-globin mRNA was found. The alpha- and zeta-globin genes were both hypomethylated and restriction endonuclease maps of the alpha- and zeta-globin genes were comparable in the patient's marrow and fibroblast DNA. The data are most consistent with the acquisition of a mutation(s) that resulted in decreased expression of all four alpha-globin genes.
Familial cutaneous malignant melanoma: autosomal dominant trait possibly linked to the Rh locus.
Authors: Greene M H , Goldin L R , Clark W H , Lovrien E , Kraemer K H , Tucker M A , Elder D E , Fraser M C , Rowe S
Published: May 2006

Segregation and linkage analyses were undertaken in families with multiple cases of cutaneous malignant melanoma (CMM) and a recently-described melanoma precursor, the dysplastic nevus syndrome (DNS). Clinical and laboratory data, including 23 genetic markers, were collected on 401 members of 14 high-risk kindreds. Pedigree analysis was compatible with an autosomal dominant mode of inheritance for the familial CMM trait. Although a similar model probably applies to the DNS trait as well, segregation analysis could not confirm the presence of a major locus. However, linkage analysis suggested that an autosomal dominant model was appropriate for the DNS, and that a DNS/CMM susceptibility gene may be located on the short arm of chromosome 1, within 30 map units of the Rh locus [maximum logarithm of odds (lod) score = 2.00].
Diffusely distributed acetylcholine receptors can participate in cluster formation on cultured rat myotubes.
Authors: Stya M , Axelrod D
Published: May 2006

On aneurally cultured rat primary myotubes, 10% of the acetylcholine receptors (AcChoR) are found to be aggregated and immobilized in endogenous clusters while the remainder are diffusely distributed and partially mobile. This paper reports that AcChoR in clusters can be gathered from AcChoR in diffuse areas during the course of normal myotube development. AcChoR were fluorescently labeled with rhodamine-conjugated alpha-bungarotoxin, and all existing clusters in a circumscribed region of the culture dish were irreversibly photobleached by a slightly defocused laser beam, the movement of which was controlled by a lens mounted on a joystick translator. This procedure leaves intact only the fluorescent label on the diffusely distributed AcChoR. Observation of the myotubes after several hours of incubation revealed cluster fluorescence redevelopment. This cluster fluorescence must have consisted of AcChoR that previously were diffusely distributed. The majority (but not all) of cluster fluorescence redevelopment occurred in the location of a previously bleached cluster. About half of the redeveloped clusters have an annular shape. The major conclusions of this study are (i) diffusely distributed AcChoR can become clustered; (ii) endogenous clusters appear to form, at least in part, by "trapping" receptors as they diffuse in from surrounding regions; (iii) cluster formation is an ongoing process in cultured rat myotubes; and (iv) colchicine (a microtubule-disrupting agent) inhibits cluster formation.