bluuebunny/crossref_metadata_2025 huggingface dataset:
Enhanced synthesis and stabilization of Mr 68,000 protein in transformed BALB/c-3T3 cells: candidate for restriction point control of cell growth.
Authors: Croy R G , Pardee A B
Published: May 2006

We have proposed that transformation of cells to tumorigenicity by chemical carcinogens can depend upon stabilization of a protein responsible for growth regulation. Cell kinetic experiments in which normal and benzo[a]pyrene-transformed BALB/c-3T3 cells were pulsed with cycloheximide indicated this protein should have a half-life of a few hours in normal cells and be considerably more stable in transformed cells [Campisi, J., Medrano, E. E., Morreo, G. & Pardee, A. B. (1982) Proc. Natl. Acad. Sci. USA 79, 436-440]. A protein with these properties has not yet been reported. We have searched for such a protein using two-dimensional electrophoresis to resolve protein from cells labeled with [35S]methionine. Among approximately 1,000 proteins that were resolved in these gels, we have found one that has a greater rate of synthesis and stability in benzo[a]pyrene-transformed than in untransformed cells. This result satisfies a necessary prediction of our labile protein hypothesis. We suggest that this protein could be important in determining the loss of growth regulation in these tumor cells.
Sequence of the
nifD
gene coding for the α subunit of dinitrogenase from the cyanobacterium
Anabaena
Authors: Lammers Peter J. , Haselkorn Robert
Published: May 2006

The nucleotide sequence of
nifD
, the structural gene for the α subunit of dinitrogenase from
Anabaena
7120, has been determined. The coding sequence contains 1,440 nucleotides, which predict an amino acid sequence of 480 residues and
M
r
of 54,283. The predicted sequence contains eight cysteines, of which five are conserved with respect to adjoining sequences and position relative to the α subunits of dinitrogenase from
Azotobacter, Clostridium
, and
Klebsiella
. Because there are also five conserved cysteines in the β subunit of
Anabaena
dinitrogenase [Mazur, B. J. & Chiu, C.-F. (1982)
Proc. Natl. Acad. Sci. USA
79, 6782-6786], the number of cysteine residues participating as ligands to FeS clusters is likely to be 20 per α
2
β
2
tetramer. This number is sufficient to accommodate the known four Fe
4
S
4
clusters, leaving at least four cysteines to be shared among the two FeMo cofactors and the more poorly characterized two-iron center. Although the α- and β-subunit gene sequences are not recognizably homologous, their secondary structures, predicted from the sequences, indicate similar domains around three of the conserved cysteine residues.
Differential expression of the normal and of the translocated human c-myc oncogenes in B cells.
Authors: Nishikura K , ar-Rushdi A , Erikson J , Watt R , Rovera G , Croce C M
Published: May 2006

We have investigated whether the translocated and the untranslocated human c-myc oncogenes of Burkitt lymphoma cells are equally or differentially expressed in host mouse B cells. The human c-myc mRNA levels in somatic cell hybrids between mouse plasmacytoma cells and Burkitt lymphoma cells with either the t(8;14) or the t(2;8) chromosome translocation were determined by using the nuclease S1 protection procedure. Although both the human parental lines and the hybrid cells carrying the translocated c-muc oncogene expressed high levels of human specific c-myc transcripts, the hybrid cells carrying the untranslocated c-myc gene on normal chromosome 8 did not contain human specific c-myc mRNA. These results suggest that the translocated human c-myc oncogene has escaped the normal transcriptional control to which the untranslocated c-myc gene remains subjected. This interpretation is also supported by the finding that the expression of the c-myc genes of lymphoblastoid cells and of HL-60 promyelocytic leukemia cells are repressed when they are transferred into a mouse plasmacytoma background. The ability of the translocated c-myc oncogene to escape the normal transcriptional control occurring in B cells may be important for the expression of B cell neoplasia in mouse and man. We have also transferred the Burkitt 14q+ chromosome carrying a translocated c-myc oncogene into mouse LM-TK- fibroblasts and studied the levels of human c-myc transcripts in the hybrids. Because the levels of human c-myc transcripts in the fibroblast hybrids are dramatically decreased in comparison to the plasmacytoma hybrids, we conclude that the levels of transcripts of the translocated c-myc oncogene depend on the differentiated state of the cells harboring the translocated chromosome.
Treatment of sickle cell anemia with 5-azacytidine results in increased fetal hemoglobin production and is associated with nonrandom hypomethylation of DNA around the gamma-delta-beta-globin gene complex.
Authors: Charache S , Dover G , Smith K , Talbot C C , Moyer M , Boyer S
Published: May 2006

Increased production of fetal hemoglobin (HbF) was observed in a patient with sickle cell anemia treated with 5-azacytidine. Each of four courses of therapy resulted in a rapid and prolonged increase in the percentage of HbF containing reticulocytes (F reticulocytes) and HbF containing erythrocytes (F cells). The percentage of HbF in peripheral blood rose from 1.8 to 8.9%. The rise in HbF production was accompanied by an increase in peripheral blood hemoglobin concentration from 8 to 12 g/dl and an increase in mean erythrocyte volume. Treatment with 5-azacytidine resulted in hypomethylation of total genomic and a Y-chromosome-specific DNA fragment isolated from both peripheral blood and bone marrow. Of 15 restriction enzyme sites around the gamma-delta-beta-globin gene complex, only 2 became hypomethylated: one 107 bases 5' to the gamma G and the other 107 bases 5' to the gamma A globin genes.
Association of 16S and 23S ribosomal RNAs to form a bimolecular complex.
Authors: Burma D P , Nag B , Tewari D S
Published: May 2006

Association of the 30S and 50S subunits to generate the 70S ribosomes of Escherichia coli has long been known but the mechanism of this interaction remains obscure. Light-scattering studies indicate that naked 16S and 23S RNAs can also associate under conditions similar to those required for the assembly of ribosomes from the constituent RNAs and proteins. The RNA-RNA association also takes place in the presence of ethanol, which promotes folding of 16S and 23S RNAs into specific compact structures with the morphological features of 30S and 50S ribosomes, respectively. Equimolar amounts of the two RNAs are involved in the association. The formation of a stoichiometric complex was shown by light scattering, sucrose density gradient centrifugation, and composite polyacrylamide/agarose gel electrophoresis. The presence of the two species of RNA in the complex was also shown by gel electrophoresis. The association of naked 16S and 23S RNAs suggests that RNA-RNA interaction may play an important role in the association of 30S and 50S subunits.
High-resolution 1H nuclear magnetic resonance study of cerebral hypoxia in vivo.
Authors: Behar K L , den Hollander J A , Stromski M E , Ogino T , Shulman R G , Petroff O A , Prichard J W
Published: May 2006

1H NMR spectra at 360.13 MHz were obtained of the rat brain in vivo by using a surface coil placed over the skull. Resonances of numerous metabolites were identified by comparison with the 1H NMR spectra of excised rat brain tissue and acid extracts of the tissue. Changes in cerebral lactate levels resulting from the administration of gas mixtures low in oxygen were monitored in the in vivo brain spectra with a time resolution of 2.3 min. The electroencephalogram and electrocardiogram were recorded simultaneously during the NMR experiment. Reversibility of the hypoxic response was documented when, upon oxygen administration, cerebral lactate returned to its prehypoxic level. These experiments demonstrate the applicability of high-resolution 1H NMR to monitor pathophysiology of brain metabolism in real time.
Generation of protein-reactive antibodies by short peptides is an event of high frequency: implications for the structural basis of immune recognition.
Authors: Niman H L , Houghten R A , Walker L E , Reisfeld R A , Wilson I A , Hogle J M , Lerner R A
Published: May 2006

Recent studies have shown that chemically synthesized small peptides can induce antibodies that often react with intact proteins regardless of their position in the folded molecule. These findings are difficult to explain in view of the experimental and theoretical data which suggest that in the absence of forces provided by the folded protein, small peptides in aqueous solution do not readily adopt stable structures. In order to rationalize the two findings, there has been general acceptance of a stochastic model which suggests that the multiple conformers of a peptide in solution induce sets of antibodies with a small percentage reactive with conformations shared by the folded protein. This stochastic model has become less tenable as the success rate for the generation of protein-reactive anti-peptide antibodies has grown. To test the stochastic model, we have used monoclonal anti-peptide antibodies as a way of estimating the frequency with which small peptides induce antibodies that react with folded proteins. We have made monoclonal antibodies to six chemically synthesized peptides from three proteins. The frequency with which the peptides induce protein-reactive antibodies is at least 4 orders of magnitude greater than expected from previous experimental work and vastly different from what would be predicted by calculating the possible number of peptide conformers in solution. These findings make the stochastic model less likely and lead to consideration of other models. Aside from their practical significance for generation of highly specific reagents, these findings may have important implications for the protein folding problem.
Granulocytes without degranulation: neutrophil function in granule-depleted cytoplasts.
Authors: Korchak H M , Roos D , Giedd K N , Wynkoop E M , Vienne K , Rutherford L E , Buyon J P , Rich A M , Weissmann G
Published: May 2006

Neutrophils respond to a variety of stimuli by generating superoxide anion, degranulating, and aggregating. Because it has been suggested that fusion of granules with the plasmalemma (degranulation) is necessary for aggregation and superoxide anion generation, we have tested whether these responses can be demonstrated in "neutrophilic cytoplasts" (granule-free vesicles of cytoplasm enclosed by plasmalemma). When examined by electron microscopy, cytoplasts were found to be approximately 4 microns in diameter and essentially granule free. Cytoplasts exposed to fMet-Leu-Phe (0.1 microM) generated superoxide anion after a lag of 16 sec but released no detectable beta-glucuronidase, lysozyme, or elastase. Aggregation of cytoplasts, as measured by changes in light transmission, was also activated by fMet-Leu-Phe; no lag period was observed. Electron microscopy of the aggregates demonstrated clusters of cytoplasts with a scalloped appearance. Superoxide anion generation and aggregation of cytoplasts were also activated by phorbol 12-myristate 13-acetate, concanavalin A, and leukotriene B4. Exposure of cytoplasts to the dye 3,3'-dihexyloxacarbocyanine iodide (DiOC6(3)] led to dye uptake and enhancement of fluorescence, implying that the vesicles were sealed and maintained a membrane potential across the plasmalemma. Exposure of DiOC6(3)-loaded cytoplasts to fMet-Leu-Phe and PMA caused a rapid loss of dye fluorescence that was not inhibited by CN-, compatible with their lack of mitochondria. Exposure of dye-loaded cytoplasts to concanavalin A or leukotriene B4 caused an increase in fluorescence--i.e., a hyperpolarization. These results demonstrate that degranulation is not a prerequisite for aggregation or superoxide anion generation. The retention of ionic gradients and changes in membrane potential, as measured by DiOC6(3) fluorescence changes, suggest a fundamental role for ionic movements in activating superoxide anion generation and aggregation.
Two-dimensional electrophoresis of plasma polypeptides reveals "high" heterozygosity indices.
Authors: Rosenblum B B , Neel J V , Hanash S M
Published: May 2006

A series of 62 plasma samples have been examined for genetic variation by the technique of two-dimensional polyacrylamide gel electrophoresis followed by silver-staining of the gels. Twenty polypeptides chosen without respect to variability were considered suitable for scoring. Of the total of 1,240 polypeptides, 29 could not be scored unambiguously. Seventy-five of the remaining 1,211 exhibited the combination of a normal and a variant polypeptide. All variants were present in either the father or the mother of the subject. This index of heterozygosity (6.2 +/- 0.7%) is substantially higher than those reported by others in similar studies of human fibroblasts, lymphocytes, kidney, or brain cells.
Complete nucleotide sequence of an infectious clone of Friend spleen focus-forming provirus: gp55 is an envelope fusion glycoprotein.
Authors: Clark S P , Mak T W
Published: May 2006

The Friend spleen focus-forming provirus is 6,296 base pairs (bp) in length. Compared to Moloney murine leukemia virus, it has undergone five major deletions, three substitutions, and a number of minor alterations. Otherwise, these viruses are about 90% homologous. A 16-bp palindrome is found in the region thought to be involved in packaging and dimerization of the RNA genome. Premature termination of translation of the gag polyprotein is attributed to a 13-bp deletion in the p12 region. A substitution of xenotropic env sequences was identified in the 5' region of the env gene; 150 nucleotides 3' to this substitution, a deletion of 585 bp removes the site where the normal env precursor protein is cleaved to form gp70 and p15(E), resulting in a fusion protein of Mr 44,725. Due to these changes, the env product gp55 is expected to have a substantially different conformation on the cell surface compared to either a xenotropic or ecotropic gp70 protein, and may be responsible for the rapid erythroleukemic potential of spleen focus-forming virus.
Deficiency in ubiquinone cytochrome c reductase in a patient with mitochondrial myopathy and lactic acidosis.
Authors: Darley-Usmar V M , Kennaway N G , Buist N R , Capaldi R A
Published: May 2006

The skeletal muscle of a patient with a mitochondrial myopathy was examined. A defect in the electron transport chain was identified at the position of complex III by activity measurements and the low levels of reducible cytochrome b. The polypeptide composition of complex III in the patient's mitochondria was determined by antibody binding experiments. The method allowed detection of individual polypeptides at a lower limit of 10-40 ng of protein. Characterization of protein composition is thus possible by using a biopsy sample of 1 g of tissue. The level of core proteins, FeS protein, and subunit VI was greatly diminished in the patient's mitochondria. Cytochrome c1 polypeptide was found at normal levels but was sensitive to proteolysis by trypsin. These results show that complex III is not assembled in the patient's mitochondria. The possible role of cytochrome b as the site of the primary lesion is discussed.
L-glutamic acid: a neurotransmitter candidate for cone photoreceptors in human and rat retinas.
Authors: Brandon C , Lam D M
Published: May 2006

We have combined immunocytochemical localization of L-aspartate aminotransferase (L-aspartate:2-oxoglutarate aminotransferase, EC 2.6.1.1; glutamic-oxaloacetic transaminase) with autoradiographic localization of high-affinity uptake sites for L-glutamate or L-aspartate to identify the neurotransmitters of mammalian photoreceptors. In both human and rat retinas, high aspartate aminotransferase immunoreactivity is found in cones but not in rods; certain putative bipolar and amacrine cells are also heavily stained. In the human retina, and perhaps also in the rat retina, cones possess a high-affinity uptake mechanism for L-glutamate but not L-aspartate, whereas rods and Müller (glial) cells take up both L-glutamate and L-aspartate. Taken together, our results indicate that (i) L-glutamate is much more likely than L-aspartate to be the transmitter for human cones, and possibly for cones of other mammalian species as well, and (ii) major differences exist between mammalian cones and rods in the transport and metabolism or utilization of L-aspartate and L-glutamate.
Patch-recorded single-channel currents of the purified and reconstituted Torpedo acetylcholine receptor.
Authors: Tank D W , Huganir R L , Greengard P , Webb W W
Published: May 2006

Small unilamellar vesicles containing purified and reconstituted nicotinic acetylcholine receptors from Torpedo electroplax have been fused by a simple freeze-thaw procedure to form large liposomes. Giga-seal patch-recording techniques were used to form isolated patches of liposome-membrane and to measure single-channel properties of the reconstituted receptor-ion channel complex. The observed properties are quantitatively similar to those reported for vertebrate muscle nicotinic acetylcholine receptor species recorded in situ. The results demonstrate that the pentameric complex consisting of the alpha 2 beta gamma delta subunits is fully functional. The methods used in these experiments should be useful in studying the effects of chemical alterations on the properties of acetylcholine receptor channels as well as other types of purified and reconstituted ion channels.
Marked increase of human platelet phospholipase A2 activity in vitro and demonstration of an endogenous inhibitor.
Authors: Ballou L R , Cheung W Y
Published: May 2006

When soluble and particulate fractions of human platelet homogenate were chromatographed on DEAE-cellulose and hydroxylapatite columns, total phospholipase A2 (PLA2; phosphatide 2-acylhydrolase, EC 3.1.1.4) activity increased sharply. PLA2 activity detected in these partially purified preparations was approximately 12 times greater than that associated with the original homogenate. Chromatography of the particulate enzyme on a DEAE-cellulose column yielded one activity peak. Fractions eluted near the activity peak showed a trace of PLA2 activity but inhibited purified PLA2 stoichiometrically, suggesting the presence of an endogenous "inhibitor" in the homogenate. The inhibitor activity was heat-stable, trypsin insensitive, and extractable by chloroform/methanol, and thus appears to be associated with a lipid(s). PLA2 activity was Ca2+ dependent. The crude enzyme was variably stimulated by calmodulin, whereas the purified enzyme was not, suggesting that the effect of calmodulin on the crude enzyme is indirect. Our results suggest that human platelet PLA2 activity reported in the literature may have been underestimated, apparently due to the presence of an endogenous inhibitor.
Molecular cloning of the 3'-proximal third of Sendai virus genome.
Authors: Dowling P C , Giorgi C , Roux L , Dethlefsen L A , Galantowicz M E , Blumberg B M , Kolakofsky D
Published: May 2006

Portions of the Sendai virus genome were randomly cloned by using virion 50S RNA and calf thymus DNA pentanucleotides as primers. The recombinant clones were probed first with radiolabeled products of an in vitro virion RNA polymerase reaction to locate early message clones and then with a probe from the viral genome 3' end to locate the most 3'-proximal clones. Clones were then ordered from the 3' end of the genome and used to construct a genetic map of the 3'-proximal third of the genome by hybrid-selection of mRNAs. We report that the gene order for this region is 3'-NP - P + C - M-5' and that the genetic loci of the viral P and C proteins cannot be separated by these techniques.
Prestalk and prespore differentiation in
Dictyostelium
as detected by cell type-specific monoclonal antibodies
Authors: Tasaka Masao , Noce Toshiaki , Takeuchi Ikuo
Published: May 2006

Monoclonal antibodies specifically reactive against prestalk and prespore cells of the cellular slime mold
Dictyostelium discoideum
were obtained. By the use of these antibodies, we examined processes of differentiation of the two cell types during development. Cells stained with prespore-specific antibodies first appeared after 12-14 hr of starvation within cell aggregates with tips, coincidentally with the appearance of other prespore markers. The number of prespore cells then increased to a level of 70-80% of total cells at the slug stage. By contrast, cells stained with prestalk-specific antibodies began to appear after 3 hr of starvation and thereafter increased in number to a maximum of
ca
. 80% after 12 hr of starvation. Stained cells appeared at random in the aggregation field and were not morphologically distinguishable from unstained cells. Furthermore, cells showed a considerable heterogeneity in the amount of antigen they contain. Concomitantly with the increase in prespore cells, the number of cells stained by the prestalk antibodies decreased to a level of
ca
. 20% by the slug stage. From these experiments, we suggest that the prestalk antigen is synthesized in the majority of cells during the early period of aggregation. Within tight cell aggregates, some of these cells lose the antigen to become prespore cells and the normal proportion between the two cell types will eventually result within slugs.
Mutations affect storage and use of memory differentially in
Drosophila
Authors: Dudai Yadin
Published: May 2006

The behavior of normal
Drosophila
and of X-linked olfactory conditioning mutants was analyzed by using an olfactory memory choice task. Memory in normal flies was optimal at 5-12 min after training and then decayed slowly. Memory in the mutants was 40-80% of normal when tested 30 sec after training but it decayed rapidly and was very low after 7 min. Successive conditioning experiments indicated that some types of mutants can behave as though they store information for a longer period than that revealed by straightforward memory tests, but this information reveals itself only by interaction with newly acquired information in a conflict situation. It thus seems that the mutations can either prevent the formation of normal memory or prevent memory from evoking a normal behavioral response.
Transcribed chromatin exhibits an altered nucleosomal spacing.
Authors: Smith R D , Seale R L , Yu J
Published: May 2006

The nucleosomal repeat lengths of bulk chromatin and the chromatin of transcriptionally active and inactive genes were analyzed in two mouse cell lines and adult mouse spleens. The adult beta-globin gene exhibits a nucleosomal repeat length approximately 11 base pairs longer than (i) an inactive embryonic globin gene, epsilon y3; (ii) an immunoglobulin heavy chain gene, Cmu; and (iii) the bulk chromatin in murine erythroleukemia cell line DS19. The repeat length of the Cmu gene was approximately 14 base pairs longer than that of the adult beta-globin or epsilon y3 genes in the IgM-producing cell line M104E. The chromatin of several inactive genes had repeat lengths less than or equal to bulk chromatin. Individual genes were shown to vary in repeat length among the cell types examined. In addition, genes that exhibited an increased nucleosomal spacing were digested to mononucleosomes more rapidly than bulk chromatin or inactive genes with shorter repeats. Increased repeat length was also correlated with an increased sensitivity to DNase I. Thus, increased nucleosomal spacing may be a property of transcriptionally active genes or genes with the potential for transcription.
Luteinizing hormone receptor appearance in cultured porcine granulosa cells requires continual presence of follicle-stimulating hormone.
Authors: Segaloff D L , Limbird L E
Published: May 2006

During the differentiation of ovarian granulosa cells, follicle-stimulating hormone (follitropin; FSH) mediates the induction of cell surface receptors for luteinizing hormone (lutropin; LH). Using primary cultures of porcine granulosa cells, we demonstrate that both the induction and maintenance of LH receptors are critically dependent upon the continual presence of FSH. The termination of FSH-promoted LH receptor induction is paralleled by the termination of FSH-induced intracellular cAMP accumulation. Changing the medium is without effect on FSH-induced appearance of LH receptors. Furthermore, 1 mM aminoglutethimide, which completely blocks FSH-stimulated progesterone biosynthesis, does not decrease the induction of LH receptors by FSH. Thus, the induction of LH receptors by FSH does not appear to require the accumulation of a factor in the medium, nor does it appear to be mediated via FSH-stimulated progesterone synthesis. These data suggest that intracellular cAMP, produced while FSH remains bound to the cell surface, mediates the induction of LH receptors and that the continual presence of FSH is required for both the induction and maintenance of cell surface LH receptors.
One of two Epstein-Barr virus nuclear antigens contains a glycine-alanine copolymer domain.
Authors: Hennessy K , Kieff E
Published: May 2006

A gene fusion between an Epstein-Barr virus (EBV) triplet nucleotide repeat array (IR3), which has homology to host DNA, and lacZ was used to demonstrate that this EBV sequence encodes part of the Epstein-Barr nuclear antigen (EBNA). The IR3 sequence is translated into a glycine-alanine copolymer that reacts with anti-EBNA human sera. Some EBV-immune human antisera recognize a second intranuclear protein that is also specific for latently infected cells and is designated EBNA2. EBNA2 is not related to EBNA1 because the molecular mass of EBNA2 is 82 kilodaltons, whereas that of EBNA1 varies from 68 to 85 kilodaltons among cells transformed by different EBV isolates; also EBNA2 does not contain the copolymer domain of EBNA1.
Efficacy of combinations of difluoromethylornithine and bleomycin in a mouse model of central nervous system African trypanosomiasis.
Authors: Clarkson A B , Bacchi C J , Mellow G H , Nathan H C , McCann P P , Sjoerdsma A
Published: May 2006

DL-alpha-Difluoromethylornithine, a polyamine biosynthesis inhibitor, and bleomycin, a currently used antineoplastic agent, have each previously been shown to be curative for acute short-term infections of mice with Trypanosoma brucei brucei, an African trypanosome closely related to those that cause the human disease African sleeping sickness. These agents were tested singly and in combination in a previously described mouse model of sleeping sickness with demonstrable brain involvement. The original model is extended by using two additional strains of outbred mice and by demonstrating that melarsoprol, an arsenical and currently the only drug used for human African trypanosomiasis involving the brain, was also curative for these brain infections. Neither difluoromethylornithine nor bleomycin alone was curative for the brain infections; however, many combinations of the two drugs were found to be 100% curative with no evidence of immediate toxicity.
Primary structure and neuronal effects of α-bag cell peptide, a second candidate neurotransmitter encoded by a single gene in bag cell neurons of
Aplysia
Authors: Rothman Barry S. , Mayeri Earl , Brown Richard O. , Yuan Pau-Miau , Shively John E.
Published: May 2006

A discharge of impulse activity in a group of neuroendocrine cells, the bag cells, produces several types of prolonged responses in various identified neurons of the abdominal ganglion of
Aplysia
. Two excitatory responses are almost certainly mediated by egg-laying hormone, but this peptide cannot account for other responses, such as inhibition of left upper quadrant neurons. We report here the isolation from bag cell clusters of three structurally similar peptides, seven, eight, and nine residues long, that are candidate transmitters for mediating bag cell-induced inhibition. They may also serve as autoexcitatory transmitters since the seven-residue peptide produces a slow depolarization of the bag cells similar to that which occurs during bag cell discharge. The amino acid sequence of the largest peptide, termed α-bag cell peptide[1-9], is H-Ala-Pro-Arg-Leu-Arg-Phe-Tyr-Ser-Leu-OH. The other two peptides are identical to α-BCP[1-9] except that they lack the COOH-terminal Ser-Leu or leucine residues. The three peptides inhibit left upper quadrant neurons at relative potencies of 10:30:1 (seven-, eight-, and nine-residue peptides, respectively). Recent molecular genetic analysis shows that both α-BCP[1-9] and egg-laying hormone are encoded by the same bag cell-specific gene. The multiple neuronal effects of bag cells are therefore likely to be mediated by at least two transmitters that are cleaved from a common precursor molecule.
X-ray diffraction by phospholipid monolayers on single-crystal silicon substrates
Authors: Seul M. , Eisenberger P. , McConnell Harden M.
Published: May 2006

Monolayers of dipalmitoyl phosphatidylcholine have been transferred from an air-water interface to single-crystal silicon wafers previously alkylated with octadecyltrichlorosilane. By using synchrotron radiation it has been found possible to observe diffraction by single-crystal regions of these monolayers with negligible background scattering from the solid substrate. In one series of experiments, diffraction signals at Bragg spacings of 0.4247 ± 0.0002 nm and 0.4253 ± 0.0002 nm were observed. The supported phospholipid monolayer crystals show remarkably high in-plane order: the positional coherence length is at least 500 nm and the orientational order is better than 0.01°. Preliminary temperature scans were carried out. The data reveal the existence of a phase transition at about 65°C.
Secondary structure of the lac repressor DNA-binding domain by two-dimensional 1H nuclear magnetic resonance in solution.
Authors: Zuiderweg E R , Kaptein R , Wüthrich K
Published: May 2006

A recently proposed approach for spatial structure determination in noncrystalline proteins by nuclear magnetic resonance was applied to the lac repressor DNA-binding domain. On the basis of sequence-specific 1H NMR assignments, the location of alpha-helices in the amino acid sequence was determined from nuclear Overhauser enhancement data and from amide proton exchange studies. These investigations provide detailed experimental data on the structure of a noncrystalline DNA-binding protein. The results support the hypothesis advanced by others that sequence-specific interactions between lac repressor and DNA are mediated by a particular spatial arrangement of two alpha-helices common to various different DNA-binding proteins.
13C NMR studies of acetate metabolism during sporulation of Saccharomyces cerevisiae.
Authors: Dickinson J R , Dawes I W , Boyd A S , Baxter R L
Published: May 2006

The sporulation of Saccharomyces cerevisiae in the presence of [2-13C]acetate was studied by 13C NMR spectroscopy. The fate of 13C label was analyzed in vivo and in cell extracts. During the first 4 hr of sporulation the major metabolite produced from [2-13C]acetate utilization was glutamate. From the labeling pattern observed it is concluded that both the tricarboxylic acid cycle and the glyoxylate cycle are operating. After about 4 hr trehalose is made. Comparison of the doublet/singlet ratios for C-1,1(1) and C-6,6(1) of trehalose shows a steady drop in the ratio of C-1, C-2-coupled species over trehalose labeled only at C-1 in the C-1, 2 segment of the molecule. The negative correlation of this ratio with that for the C-5, 6 segment indicates a cycling of glucose through the hexose monophosphate shunt. Subsequently fatty acid biosynthesis commences. Large amounts of saturated fatty acid were made. There were conspicuous differences observed in the metabolism of [2-13C]acetate between sporulating and vegetatively growing cells.
Structural basis for Robertsonian translocations in man: association of ribosomal genes in the nucleolar fibrillar center in meiotic spermatocytes and oocytes.
Authors: Stahl A , Luciani J M , Hartung M , Devictor M , Bergé-Lefranc J L , Guichaoua M
Published: May 2006

The spatial relationships of acrocentric chromosomes were studied during prophase I of meiosis in human oocytes and spermatocytes by using cytogenetic techniques, electron microscopy, and in situ hybridization. Ultrastructural investigations revealed an ordered arrangement of nucleolar bivalents at the zygotene and pachytene stages. The end of the bivalent corresponding to the cytological satellite was consistently attached to the nuclear envelope. The fibrillar center of the nucleolus always contained rDNA chromatin fibers emanating from the secondary constriction region. Association of ribosomal genes from two bivalents in the same fibrillar center was frequently observed. Ultrastructural studies demonstrated the close proximity of chromatids in the short arm region of the involved nonhomologous acrocentrics. A breakage/reunion model based on our data can explain the formation of all observed types of Robertsonian translocations: monocentrics and dicentrics with or without rDNA.
Murine monoclonal anti-DNA antibodies with an absolute specificity for DNA have a large amount of idiotypic diversity.
Authors: Tron F , Jacob L , Bach J F
Published: May 2006

The clonal heterogeneity of nine monoclonal antibodies with absolute specificities for deoxyribonucleic acid (DNA) was analyzed. These monoclonal anti-DNA antibodies were generated in three different fusion experiments using autoimmune (NZB X NZW)F1 mouse spleen cells. Isoelectric focusing analyses demonstrated different isoelectric points within the IgG2a and IgG2b subclasses. Three anti-idiotypic antisera were prepared (one in a rabbit and two in mice) against two monoclonal anti-DNA antibodies. These antisera detected idiotypic determinants uniquely associated with homologous hybridoma anti-DNA antibodies. Two of these idiotypes could be detected at low levels in the sera of (NZB X NZW)F1 mice. Anti-PME77 idiotypic antiserum had no effect in vitro on the total binding capacity of (NZB X NZW)F1 sera. Taken together these results demonstrate that, in (NZB X NZW)F1 mice, the anti-DNA antibody repertoire contains molecules that show similar antigen binding characteristics but are not structurally uniform.
Dendritic cells are critical accessory cells for thymus-dependent antibody responses in mouse and in man.
Authors: Inaba K , Steinman R M , Van Voorhis W C , Muramatsu S
Published: May 2006

We report that dendritic cells (DC) are necessary and potent accessory cells for anti-sheep erythrocyte responses in both mouse and man. In mice, a small number of DC (0.3-1% of the culture) restores the response of B/T-lymphocyte mixtures to that observed in unfractionated spleen. An even lower dose (0.03-0.1% DC) is needed if the T cells have been primed to antigen. Responses are both antigen and T cell dependent. Selective depletion of DC from unfractionated spleen with the monoclonal antibody 33D1 and complement ablates the antibody response. In contrast to DC, purified spleen macrophages are weak or inactive stimulators. However, when mixed with DC, macrophages can increase the yield of antibody-secreting cells about 2-fold. In man, small numbers (0.3-1%) of blood DC stimulate antibody formation in vitro. Purified human monocytes do not stimulate but in low doses (1% of the culture) inhibit the antibody response. Likewise, selective removal of human monocytes with antibody and complement enhances or accelerates the development of antibody-secreting cells. We conclude that DC are required for the development of T-dependent antibody responses by mouse and human lymphocytes in vitro.
Amiloride reduces the taste intensity of Na+ and Li+ salts and sweeteners.
Authors: Schiffman S S , Lockhead E , Maes F W
Published: May 2006

The diuretic amiloride, a potent inhibitor of sodium transport in a variety of epithelial systems, was applied to the human tongue. Application of amiloride reduced the taste intensity of sodium and lithium salts and of sweeteners ranging widely in chemical structure. The sweeteners included saccharides, glycosides, dipeptides, proteins, and amino acids. Amiloride did not affect perception of potassium or calcium salts, bitter and sour tastes, or amino acids without a sweet or salty component. These findings were supported by neurophysiological studies in rat, which showed that amiloride diminished the NaCl response relative to KCl. The results are consistent with the position that an amiloride-sensitive transport mechanism is involved in taste perception of sodium and lithium salts and of sweeteners.
In vivo
31
P NMR study of the metabolism of murine mammary 16/C adenocarcinoma and its response to chemotherapy, x-radiation, and hyperthermia
Authors: Evanochko W. T. , Ng T. C. , Lilly M. B. , Lawson A. J. , Corbett T. H. , Durant J. R. , Glickson J. D.
Published: May 2006

31
P NMR spectroscopy with surface coils has been used to monitor,
in vivo
, the phosphate metabolism of subcutaneously implanted mammary 16/C adenocarcinoma in C3H/He mice. This model tumor was studied during untreated tumor growth and after treatment with adriamycin, hyperthermia, and x-radiation. The mammary 16/C tumor exhibited a Gompertzian growth pattern. Levels of high-energy phosphate metabolites—phosphocreatine and ATP—decreased with increases in tumor mass. There was a concomitant increase in the level of P
i
and a decrease in the apparent pH of the tumor. These spectral changes appear to reflect changes in tumor vascularization that accompany tumor growth, the tumor becoming progressively more hypoxic. Partial response of this tumor to chemotherapy with adriamycin was reflected in a small but measurable increase in the phosphocreatine resonance, a decrease in P
i
, and a return of the intra-tumor pH to neutral. Hyperthermia resulted in progressive conversion of the
31
P NMR spectrum to that of a dead tumor (high levels of P
i
, small levels of residual sugar phosphates and pyridine dinucleotides, and acidic pH). X-irradiation (14.0 Gy) led to disappearance of the phosphocreatine peak within 15 min of treatment. Subsequently, this resonance grew back beyond its pretreatment level. As the tumor receded, its spectrum reflected the characteristics of aerobically metabolizing tissue (high levels of phosphocreatine and ATP and low levels of P
i
and sugar phosphates).
Paramagnetic centers in the nickel-containing, deazaflavin-reducing hydrogenase from
Methanobacterium thermoautotrophicum
Authors: Kojima Nakao , Fox Judith A. , Hausinger Robert P. , Daniels Lacy , Orme-Johnson William H. , Walsh Christopher
Published: May 2006

Two hydrogenases from the methanogenic bacterium
Methanobacterium thermoautotrophicum
strain ΔH have been purified and contain tightly bound nickel as well as the anticipated iron/sulfur atoms with a fixed ratio of 15-20 iron atoms per nickel. One hydrogenase reduces the 8-hydroxy-5-deazaflavin coenzyme factor 420 (F
420
), whereas the other has been purified as a methyl viologen-reducing hydrogenase. Both enzymes possess an EPR signal attributed to paramagnetic nickel as demonstrated by hyperfine coupling in
61
Ni-containing hydrogenases. Comparison to model compounds suggests a nickel(III) oxidation state in the inactive forms of these aerobically purified enzymes. Loss of the nickel(III) signal accompanies reductive activation but is not kinetically correlated with regain of high specific activity. On replacement of H
2
by argon in the gas phase over reduced, active, F
420
-reducing enzyme, several EPR signals appear, including a signal at
g
= 2.004 that is probably enzyme-bound FADH semiquinone, two signals at
g
= 2.140 and 2.196 that reflect a new form of paramagnetic nickel(III), and also a signal at
g
= 2.036 that may be an iron signal. The F
420
-reducing hydrogenase in the second paramagnetic nickel form is either itself active or in facile equilibrium with active enzyme. The size of the signal at
g
= 2.036 may correlate with the degree of activation of the enzyme. In contrast to the hydrogenase of
Clostridium pasteurianum
[Erbes, D. L., Burris, R. H. & Orme-Johnson, W. H. (1975)
Proc. Natl. Acad. Sci. USA
72, 4795-4799], which appears to use only iron/sulfur prosthetic groups and which reacts with one-electron-transfer agents, this methanogen hydrogenase seems to utilize iron, nickel, and flavin redox sites and to reduce obligate one-electron (viologen) and two-electron (deazaflavin) oxidants.
Cloned restriction/modification system from Pseudomonas aeruginosa.
Authors: Gingeras T R , Brooks J E
Published: May 2006

DNA fragments from Pseudomonas aeruginosa carrying the PaeR7 restriction/modification genes have been cloned in the plasmid vector pBR322 and propagated in Escherichia coli. A subclone (pPAORM3.8) has been constructed that contains the complete restriction/modification system on a 3.8-kilobase DNA fragment. Digestion of the pPAORM3.8 plasmid with nuclease BAL-31 has yielded two types of clones. One type contains an active methylase gene but no active endonuclease gene; such clones will modify the DNA but not restrict the growth of incoming phage in vivo. The second type contains an active endonuclease gene but no active methylase gene, as judged both by in vivo tests and by the activity of the cell extracts in vitro. Although extracts of cells containing these plasmids display restriction endonuclease activity, these bacteria are unable to restrict the growth of incoming phage. Furthermore, chromosomal and phage DNA isolated from these host cells are not protected against cleavage by PaeR7 in vitro. The properties of PaeR7 endonuclease and methylase enzymes have also been examined. The PaeR7 restriction endonuclease recognizes and cleaves the sequence C decreased T-C-G-A-G, as does Xho I. However, there exists a canonical Xho I site at 26.5% on the adenovirus 2 genome which is totally refractory to PaeR7 cleavage but is cut by Xho I. Under conditions of low salt, high glycerol, and high enzyme concentrations, a "PaeR7" activity is found that is similar to that observed for EcoRI. Finally, evidence is presented that the PaeR7 methylase modifies the adenine residue within the recognition sequence.
Primary structure of an amyloid prealbumin variant in familial polyneuropathy of Jewish origin.
Authors: Pras M , Prelli F , Franklin E C , Frangione B
Published: May 2006

The complete amino acid sequence of three related amyloid proteins (Mr 14,000, 10,000, and 5,000) derived from tissues of a Jewish patient who suffered from a variant of familial polyneuropathic amyloidosis was determined. The protein, which contains 127 residues, is identical to a human serum prealbumin subunit. Only one amino acid substitution, glycine for threonine, was detected at position 49, where enzymatic cleavage occurred, yielding Mr 5,000 and 10,000 fragments which represent the amino terminus (residues 1-48) and carboxyl terminus (residues 49-127) of the molecule, respectively. Thus, a prealbumin variant and its fragments constitute the amyloid fibrils in a heredofamilial amyloidosis syndrome of dominant inheritance.
Some rat sensory neurons in culture express characteristics of differentiated pain sensory cells.
Authors: Baccaglini P I , Hogan P G
Published: May 2006

Sensory neurons were dissociated from trigeminal ganglia or from dorsal root ganglia of rats, grown in culture, and examined for expression of properties of pain sensory cells. Many sensory neurons in culture are excited by low concentrations of capsaicin, reportedly a selective stimulus for pain sensory neurons. Many are excited by bradykinin, sensitized by prostaglandin E2, or specifically stained by an antiserum against substance P. These experiments provide a basis for the study of pain mechanisms in cell culture.
A pteroylpolyglutamate binds to tetramers in deoxyhemoglobin but to dimers in oxyhemoglobin.
Authors: Benesch R E , Benesch R , Kwong S , Baugh C M
Published: May 2006

The binding of a physiological concentration of pteroylhepta(glutamate) to oxy- and deoxyhemoglobin in large excess was measured by ultrafiltration. The variation of free to bound folate with hemoglobin concentration showed that a single molecule of the pteroylpolyglutamate is bound by deoxyhemoglobin tetramers and by alpha beta dimers in oxyhemoglobin. Although the binding sites are different, the affinity constants are the same and very similar to the 2,3-bisphosphoglycerate binding energy. Nevertheless, in view of the small proportion of dimers in oxyhemoglobin much more pteroylhepta(glutamate) is bound by deoxyhemoglobin over a wide range of hemoglobin concentrations. Because even 2% deoxyhemoglobin is enough to bind all of the erythrocyte folate as polyglutamate, the bulk of it will be bound at physiological oxygen pressures. Free folate could only be expected in fully oxygenated erythrocytes. Therefore, the reaction of pteroylpolyglutamates with hemoglobin represents an oxygenation-dependent storage mechanism that can account for the 40-fold excess of the vitamin in the erythrocyte over the amounts in the serum. Because methotrexate is also converted to polyglutamate derivatives in the erythrocyte, this drug is likely to be concentrated and stored there by the same mechanism.
Chromosomal mapping of the mink cell focus-inducing and xenotropic env gene family in the mouse.
Authors: Blatt C , Mileham K , Haas M , Nesbitt M N , Harper M E , Simon M I
Published: May 2006

Chromosomal locations of members of the xenotropic-related env gene family in the mouse genome have been determined. Endonuclease restriction site polymorphisms detected by molecular hybridization were used to study the inheritance of mink cell-focus inducing and xenotropic env gene-related sequences in recombinant inbred strains of mice. Some of the endogenous env sequences appear to be closely linked to genes determining leukemia virus induction and to genes involved in the immune response, such as the heavy and light chains of the immunoglobulin molecules or allotypic determinants on B and T lymphocytes. The use of probes that detect restriction fragment length polymorphisms in a small family of dispersed sequences promises to yield a large number of markers that can be used together with recombinant inbred strains for efficient mapping of the mouse genome.
Membership mutation of the genetic code: loss of fitness by tryptophan.
Authors: Wong J T
Published: May 2006

Bacillus subtilis strain QB928, a tryptophan-auxotroph, was serially mutated to yield strain HR15. For QB928, tryptophan functioned as a competent amino acid and 4-fluorotryptophan as merely an inferior analogue. For HR15, these roles were reversed. The tryptophan/4-fluorotryptophan growth ratio decreased by a factor of 2 X 10(4) in the transition from QB928 to HR15.
Studies on mutant human insulin genes: identification and sequence analysis of a gene encoding [SerB24]insulin.
Authors: Haneda M , Chan S J , Kwok S C , Rubenstein A H , Steiner D F
Published: May 2006

Both alleles of the insulin gene of a patient with mild diabetes [maturity-onset-diabetes-of-the-young (MODY)-type syndrome] associated with hyperinsulinemia have been cloned, and the sequences have been determined. One allele contained a mutation (single nucleotide transition) in the coding sequence for the B chain at position 24 (TTC leads to TCC), resulting in the loss of a restriction enzyme (Mbo II) cleavage site in the gene. This mutation results in the substitution of serine for phenylalanine in a critically important region of the insulin molecule that is intimately involved in receptor binding. Both insulin alleles were of the alpha type and, aside from a single nucleotide deletion in the 5' region of the normal allele, their sequences were identical to those previously determined.