bluuebunny/crossref_metadata_2025 huggingface dataset:
Unusual cardioactive peptide (CCAP) from pericardial organs of the shore crab
Carcinus maenas
Authors: Stangier J. , Hilbich C. , Beyreuther K. , Keller R.
Published: May 2006

An unusual crustacean cardioactive peptide (CCAP) from the pericardial organs of the shore crab
Carcinus maenas
has been purified to homogeneity by a two-step reversed-phase HPLC procedure. Manual microsequencing using the 4-(
N,N
-dimethylamino)azobenzene 4′-isothiocyanate/phenylisothiocyanate double-coupling technique and automated gas-phase sequencing of the oxidized peptide revealed that CCAP is a nonapeptide (
M
r
957) of the sequence Pro-Phe-[unk]Cys-Asn-Ala-Phe-Thr-Gly-Cys-NH
2
. We have confirmed the sequence by chemical synthesis of the C-terminally amidated and nonamidated forms of the peptide. The presence of the amide group was indicated by lack of susceptibility to carboxypeptidase A and Y treatment and was confirmed by the observation that the native CCAP comigrated with the amidated synthetic peptide on HPLC. Native and synthetic CCAP displayed high accelerating activity on a semi-isolated crab heart preparation, whereas the nonamidated synthetic peptide was of much lower potency. The effect of CCAP was both inoand chronotropic. The two pericardial organs of one animal yielded 30-40 pmol of extractable CCAP. Its sequence does not resemble that of any known neuropeptide. However, a “mirror-image” similarity to vasopressin is conspicuous.
Structure and expression of the gene encoding the vasoactive intestinal peptide precursor.
Authors: Linder S , Barkhem T , Norberg A , Persson H , Schalling M , Hökfelt T , Magnusson G
Published: May 2006

The gene encoding the human vasoactive intestinal peptide (VIP) and the histidine-methionine amide (PHM-27) peptide hormone was isolated from lambda phage libraries. The human gene was found to be composed of seven exons spanning approximately 9 kilobase pairs. The first exon codes for an untranslated leader sequence, and the second exon codes for a putative signal peptide. DNA sequences coding for the VIP and PHM-27 hormones are located in two different exons. Southern blot analysis with genomic DNA suggested that a single copy of the VIP/PHM-27 gene is present in the human haploid genome. The expression of VIP/PHM-27 precursor mRNA in various tissues in the rat was analyzed by RNA gel blot hybridization. In the organs examined, expression was only detected in the brain and duodenum. RNA isolated from various regions of the rat brain--including the cortex, hypothalamus, and hippocampus--hybridized to both VIP- and PHM-27-specific probes. The same pattern of hybridization was found when VIP- and PHM-27-specific probes were used, suggesting that possible differences in the localization of VIP and PHM-27 peptides between different brain regions cannot be accounted for by differential RNA processing.
On the possibility of dust condensation in the ejecta of supernova 1987a
Authors: Gehrz R. D. , Ney E. P.
Published: May 2006

We suggest that supernova 1987a may condense dust of substantial visual optical thickness as do many novae. The dust will act as a calorimeter of the photon luminosity of any central engine that is dominant at the time of dust formation. Observations of novae suggest that dust formation may occur when the expanding ejecta reach a temperature of 1000 K. The early luminosity of the supernova may be dominated by radioactivity that is unrelated to the central engine that determines the energy balance for the long-term development of the supernova. We discuss the possibility that a constant luminosity central power source such as a pulsar dominates the luminosity of the supernova ejecta by the time that dust can condense and argue that, if a shell mass of more than a few tenths of one solar mass was ejected, emission from dust may be observable in the thermal infrared spectral region. Maximum dust optical depth should occur by late 1987 or early 1988. If the dust becomes optically thick, the visual light from the supernova may drop precipitously. The characteristics of an optically thick dust shell as a calorimeter of the luminosity of the central engine are discussed and are related to previous observations of dust formation in type II supernovae. It is suggested that dust of several chemical compositions may form at different epochs.
Proposed mechanism for generation and localization of new cell division sites during the division cycle of Escherichia coli.
Authors: Cook W R , Kepes F , Joseleau-Petit D , MacAlister T J , Rothfield L I
Published: May 2006

The earliest detectable event at future sites of cell division in Escherichia coli is the appearance of paired periseptal annuli that flank the site of formation of the division septum. The development and localization of these structures were followed as the cell progressed through the division cycle. The data suggest that (i) new periseptal annuli are generated from annuli already in position at the midpoint of the newborn cell; (ii) the nascent annuli are then displaced laterally during cell elongation to positions at 1/4 and 3/4 cell length; and (iii) the annuli at 1/4 and 3/4 cell length are retained during division, becoming the midpoint annuli of the newborn cells at the sites of the forthcoming division septum. The results indicate that the sites of future divisions can be identified and committed to the division process prior to the division cycle in which these sites are utilized for septum formation, and they suggest a model in which preexisting sites of cell division generate future division sites by a replication/displacement mechanism.
Insulin exerts metabolic and growth-promoting effects by a direct action on the liver in vivo: clarification of the functional significance of the portal vascular link between the beta cells of the pancreatic islets and the liver.
Authors: Griffen S C , Russell S M , Katz L S , Nicoll C S
Published: May 2006

The functional significance of the portal vascular link between the beta cells of the pancreatic islets and the liver has not been established. Previous studies indicated that insulin does not acutely regulate glucose metabolism by a direct hepatic effect. More recent observations suggest that the role of insulin in regulating body growth may be mediated, at least in part, by the liver. Our experiments were designed to test whether insulin can promote body growth and regulate glucose metabolism by a direct hepatic action in vivo. Rats were made diabetic by injections of streptozotocin, and insulin or solvent was infused into the jugular vein (JV) or the hepatic portal vein (HPV) for 14 days using catheters that were attached to osmotic minipumps. Infusion of a low dose of insulin (2 units per kg per day) into the JV had no effects on the hyperglycemia, body weight gain, tail growth, tibial epiphysial cartilage plate thickness, or serum levels of somatomedin C in the diabetic rats. However, the same dose given into the HPV caused a 30% reduction of blood glucose and stimulated a significant degree of growth, as determined by all indices. Infusion of a higher dose of insulin (5 units per kg per day) into either vein caused full restoration of body weight gain and tail growth and it restored the glycemic status almost to normal. However, it did not increase the tibial epiphysial plate width or serum somatomedin C levels above those of the rats given the low dose of the hormone into the HPV. These results indicate that insulin can act directly on the liver to promote body growth and to regulate glucose metabolism. The significance of direct delivery of insulin from the pancreatic beta cells to the liver may be as much for growth control as for glucose homeostasis.
Genomic sequence of hst, a transforming gene encoding a protein homologous to fibroblast growth factors and the int-2-encoded protein.
Authors: Yoshida T , Miyagawa K , Odagiri H , Sakamoto H , Little P F , Terada M , Sugimura T
Published: May 2006

hst is a transforming gene first identified from transformed NIH 3T3 cells that were transfected with DNA of a human stomach cancer. A genomic fragment of hst obtained directly from a human genomic library also has transforming activity. This fragment has a coding sequence identical to that of the hst cDNA prepared from an NIH 3T3 transformant induced by DNA from a stomach cancer. The deduced amino acid sequence of the hst protein is 43%, 38%, and 40% homologous, respectively, to human basic fibroblast growth factor, human acidic fibroblast growth factor, and mouse int-2 protein in selected regions. This suggests that hst encodes a protein related to fibroblast growth factors, which are wide-spectrum mitogens, and to the int-2 protein, a potential oncogene product implicated in murine mammary carcinogenesis.
Discriminating figure from ground: the role of edge detection and region growing.
Authors: Mumford D , Kosslyn S M , Hillger L A , Herrnstein R J
Published: May 2006

Three general classes of algorithms have been proposed for figure/ground segregation. One class attempts to delineate figures by searching for edges, whereas another class attempts to grow homogeneous regions; the third class consists of hybrid algorithms, which combine both procedures in various ways. The experiment reported here demonstrated that humans use a hybrid algorithm that makes use of both kinds of processes simultaneously and interactively. This conclusion follows from the patterns of response times observed when humans tried to recognize degraded polygons. By blurring the edges, the edge-detection process was selectively impaired, and by imposing noise over the figure and background, the region-growing process was selectively impaired. By varying the amounts of both sorts of degradation independently, the interaction between the two processes was observed.
A relaxation model for memory with high storage density.
Authors: Bachmann C M , Cooper L N , Dembo A , Zeitouni O
Published: May 2006

We present a relaxation model for memory based on a generalized coulomb potential. The model has arbitrarily large storage capacity and, in addition, well-defined basins of attraction about stored memory states. The model is compared with the Hopfield relaxation model.
Scanning transmission electron microscopic examination of the hexagonal bilayer structures formed by the reassociation of three of the four subunits of the extracellular hemoglobin of Lumbricus terrestris.
Authors: Kapp O H , Mainwaring M G , Vinogradov S N , Crewe A V
Published: May 2006

A fraction obtained by gel filtration at neutral pH of the extracellular Hb of Lumbricus terrestris dissociated either at pH 9.8 or at pH 4.0, consisting of the three subunits D1 (31 kDa), D2 (37 kDa), and T (50 kDa), was found to produce two peaks when subjected to gel filtration on Superose 6 at pH 7. The first peak, which was eluted at a slightly greater volume than the native Hb, consisted of reassociated hexagonal bilayer structures when examined by scanning transmission electron microscopy. The dimensions of the two reassociated hexagonal bilayer structures were a vertex-to-vertex diameter of 25 nm and a height of 16 nm. The difference in size between the hexagonal bilayer structures and the native Hb is the contribution of subunit M, which consists of a single heme-containing chain I (16.75 kDa). Although the reassociated hexagonal bilayer structures have overall dimensions smaller than the 30 nm x 20 nm dimensions of the native Hb, the diameters of the central cavities are not substantially altered. Subtraction of the three-dimensional reconstructions of the reassociated hexagonal bilayer structures from those of the native Hb showed that subunit M was primarily localized at the periphery of Lumbricus Hb. The formation of hexagonal bilayer structures in the complete absence of subunit M provides additional support for the "bracelet" model of the quaternary structure of Lumbricus Hb proposed recently by us in which subunits D1 and D2 were assumed to act as linkers for complexes of subunits M and T or to form a "bracelet" decorated with 12 complexes of subunits M and T.
Close linkage of the locus for X chromosome-linked severe combined immunodeficiency to polymorphic DNA markers in Xq11-q13.
Authors: de Saint Basile G , Arveiler B , Oberlé I , Malcolm S , Levinsky R J , Lau Y L , Hofker M , Debre M , Fischer A , Griscelli C
Published: May 2006

The gene for X chromosome-linked severe combined immunodeficiency (SCID), a disease characterized by a block in early T-cell differentiation, has been mapped to the region Xq11-q13 by linkage analysis with restriction fragment length polymorphisms. High logarithm of odds (lod) scores were obtained with the marker 19.2 (DXS3) (z = 5.51 at a recombination fraction theta = 0.11) and with the marker cpX73 (DXS159) that showed complete cosegregation with the disease locus in the informative families analyzed (z = 5.27 at theta = 0.00). Other significant linkages were obtained with several markers from Xq11 to q22. With the help of a recently developed genetic map of the region, it was possible to perform multipoint linkage analysis, and the most likely genetic order is DXS1-(SCID, DXS159)-DXYS1-DXYS12-DXS3, with a maximum multipoint logarithm of odds score of 11.0. Our results demonstrate that the SCID locus (gene symbol IMD4) is not closely linked to the locus of Bruton's agammaglobulinemia (a defect in B-cell maturation). They also provide a way for a better estimation of risk for carrier and antenatal diagnosis.
Sex pheromone production and perception in European corn borer moths is determined by both autosomal and sex-linked genes
Authors: Roelofs Wendell , Glover Thomas , Tang Xian-Han , Sreng Isabelle , Robbins Paul , Eckenrode Charles , Löfstedt Christer , Hansson Bill S. , Bengtsson Bengt O.
Published: May 2006

Inheritance patterns for sex pheromone production in females, pheromone detection on male antennal olfactory receptor cells, and male pheromone behavioral responses were studied in pheromonally distinct populations of European corn borers from New York State. Gas chromatographic analyses of pheromone glands, single sensillum recordings, and flight tunnel behavioral analyses were carried out on progeny from reciprocal crosses, as well as on progeny from subsequent F
2
and maternal and paternal backcrosses. The data show that the production of the female pheromone blend primarily is controlled by a single autosomal factor, that pheromone-responding olfactory cells are controlled by another autosomal factor, and that behavioral response to pheromone is controlled by a sex-linked gene. F
1
males were found to possess olfactory receptor cells that give spike amplitudes to the two pheromone isomers that are intermediate to those of the high and low amplitude cells of the parent populations. Fifty-five percent of the F
1
males tested responded fully to pheromone sources ranging from the hybrid (
E
)-11-tetradecenyl acetate/(
Z
)-11-tetradecenyl acetate (E/Z) molar blend of 65:35 to the E/Z molar blend of 3:97 for the Z morph parents, but very few responded to the E/Z molar blend of 99:1 for the E morph parents. Data on the inheritance patterns support speculation that the Z morph is the ancestral and that the E morph is the derived European corn borer population.
Atypical mitochondrial DNA from the deep-sea scallop
Placopecten magellanicus
Authors: Snyder Marlene , Fraser Alan R. , LaRoche Julie , Gartner-Kepkay Katherine E. , Zouros Eleftherios
Published: May 2006

The mitochondrial DNA of most metazoan animals is highly conserved in size, averaging about 17 kilobase paris (kbp). The mitochondrial DNA from the deep-sea scallop
Placopecten magellanicus
, in contrast, has been found to be approximately 34 kbp long. It is also highly variable in size from individual to individual and is unusual in the extent of its size variation. Mitochondrial DNAs from individuals collected at the same site differ by as much as 7 kbp. The size variation is due largely to differences in the number of copies of a tandemly repeated 1.2-kbp element.
Spatial segregation of mRNA encoding myelin-specific proteins.
Authors: Trapp B D , Moench T , Pulley M , Barbosa E , Tennekoon G , Griffin J
Published: May 2006

The cellular and subcellular distributions of mRNAs encoding three myelin-specific proteins--myelin basic protein (MBP), proteolipid protein (PLP), and Po protein--were studied in tissue sections of developing rat nervous systems by in situ hybridization. The developmental appearance of these mRNAs closely paralleled the appearance of the proteins they encode as determined by immunocytochemistry. mRNA encoding the extrinsic membrane protein, MBP, was concentrated around oligodendrocyte and Schwann cell nuclei during initial stages of myelination; as myelination proceeded, MBP mRNA became distributed diffusely over myelinated fibers. In contrast, mRNAs encoding the intrinsic membrane proteins, PLP and Po, remained concentrated around oligodendrocyte (PLP) and Schwann cell (Po) nuclei at all stages of myelination. These results establish that myelinating cells spatially segregate certain myelin-specific mRNAs. The presence of MBP mRNA within the cytoplasmic domains of myelin internodes indicates that protein sorting during myelination involves transportation of mRNA to specific subcellular sites.
Molecular architecture of rabbit skeletal muscle aldolase at 2.7-A resolution.
Authors: Sygusch J , Beaudry D , Allaire M
Published: May 2006

The molecular architecture of the rabbit skeletal muscle aldolase (D-fructose-1,6-bisphosphate D-glyceraldehyde-3-phosphate-lyase, EC 4.1.2.13) tetramer has been determined to 2.7-A resolution. Solution of the three-dimensional structure of rabbit muscle aldolase utilized phase information from a single isomorphous Pt(CN)4(2-) derivative, which was combined with iterative-phase refinement based upon the noncrystallographic 222-fold symmetry exhibited by the tetramer subunits. The electron-density map calculated from the refined phases (mf = 0.72) was interpreted on the basis of the known amino acid sequence (363 amino acids per subunit). The molecular architecture of the aldolase subunit corresponds to a singly wound beta-barrel of the parallel alpha/beta class structures as has been observed in triose phosphate isomerase, pyruvate kinase, phosphogluconate aldolase, as well as others. Close contacts between tetramer subunits are virtually all between regions of hydrophobic residues. Contrary to other beta-barrel structures, the known active-site residues are located in the center of the beta-barrel and are accessible to substrate from the COOH side of the beta-barrel. Biochemical and crystallographic data suggest that the COOH-terminal region of aldolase covers the active-site pocket from the COOH side of the beta-barrel and mediates access to the active site. On the basis of sequence studies, active-site residues as well as residues lining the active-site pocket have been totally conserved throughout evolution. By comparison, homology in the COOH-terminal region is minimal. It is suggested that the amino acid sequence of the COOH-terminal region may be, in part, the basis for the variable specific activities aldolases exhibit toward their substrates.
Occurrence of beta-hydroxylated asparagine residues in non-vitamin K-dependent proteins containing epidermal growth factor-like domains.
Authors: Przysiecki C T , Staggers J E , Ramjit H G , Musson D G , Stern A M , Bennett C D , Friedman P A
Published: May 2006

Vitamin K-dependent bovine protein S has been shown to contain a posttranslationally hydroxylated asparagine within a conserved sequence in three of its epidermal growth factor (EGF)-like domains. In a review of amino acid sequences deduced from cDNA data, we have observed that a conserved sequence containing a potential asparagine hydroxylation site exists within EGF-like domains of a variety of functionally diverse proteins. We have studied a number of these and report the presence of erythro-beta-hydroxyasparagine (e-beta Hyn) in three non-vitamin K-dependent proteins: the plasma complement proteins C1r and C1s (where overbar indicates activated form) and the urinary protein uromodulin. For each protein, e-beta Hyn was identified in enzyme digests following the initial observation of erythro-beta-hydroxyaspartic acid (e-beta Hya) in acid hydrolysates of the proteins. e beta Hya and e-beta Hyn residues are detected by a postcolumn derivatization cation-exchange HPLC method herein described. HPLC isolation of the presumptive e-beta Hyn residue from enzyme digests of intact C1r allowed confirmation of its structure by GC/MS. Based upon available cDNA sequence data and observation of e-beta Hya in acid hydrolysates, we suggest other proteins in which e-beta Hyn may occur.
Selective elimination of mRNAs in vivo: complementary oligodeoxynucleotides promote RNA degradation by an RNase H-like activity.
Authors: Dash P , Lotan I , Knapp M , Kandel E R , Goelet P
Published: May 2006

Oligodeoxynucleotides lead to translation arrest of complementary mRNAs in the wheat germ translation system by a degradation of the mRNA. In an attempt to develop an effective reverse genetic approach in vivo, we demonstrate that injection of short (15- to 30-nucleotide) oligonucleotides into Xenopus oocytes leads to complete degradation of both injected and endogenous mRNAs by means of an RNase H-like activity.
Cloning of the cDNA for the human beta 1-adrenergic receptor.
Authors: Frielle T , Collins S , Daniel K W , Caron M G , Lefkowitz R J , Kobilka B K
Published: May 2006

Screening of a human placenta lambda gt11 library has led to the isolation of the cDNA for the human beta 1-adrenergic receptor (beta 1AR). Used as the probe was the human genomic clone termed G-21. This clone, which contains an intronless gene for a putative receptor, was previously isolated by virtue of its cross hybridization with the human beta 2-adrenergic receptor (beta 2AR). The 2.4-kilobase cDNA for the human beta 1AR encodes a protein of 477 amino acid residues that is 69% homologous with the avian beta AR but only 54% homologous with the human beta 2AR. This suggests that the avian gene encoding beta AR and the human gene encoding beta 1AR evolved from a common ancestral gene. RNA blot analysis indicates a message of 2.5 kilobases in rat tissues, with a pattern of tissue distribution consistent with beta 1AR binding. This pattern is quite distinct from the pattern obtained when the beta 2AR cDNA is used as a probe. Expression of receptor protein in Xenopus laevis oocytes conveys adenylate cyclase responsiveness to catecholamines with a typical beta 1AR specificity. This contrasts with the typical beta 2 subtype specificity observed when the human beta 2AR cDNA is expressed in this system. Mammalian beta 1AR and beta 2AR are thus products of distinct genes, both of which are apparently related to the putative G-21 receptor.
cDNA cloning for a bile canaliculus domain-specific membrane glycoprotein of rat hepatocytes.
Authors: Hong W , Doyle D
Published: May 2006

Hepatocytes are polarized cells with distinct sinusoidal, bile canalicular, and basolateral plasma membrane domains. Each domain contains proteins that are specific for it. We have isolated three cDNA clones encoding a rat liver bile canaliculus domain-specific glycoprotein with Mr 110,000 (gp110) by immunologically screening a rat kidney lambda gt11 cDNA library with a rabbit polyclonal antiserum directed against purified gp110. The authenticity of these clones was verified as follows. (i) The antiserum recognizes specifically isopropyl beta-D-thiogalactoside-induced fusion proteins on electrophoretic transfer blots of total lysogen lysates containing these cDNA clones. (ii) Antibodies epitope-selected by these clones are able to interact with gp110 on electrophoretic transfer blots. (iii) The amino acid sequencing derived from the DNA sequence was confirmed by amino acid sequencing of a tryptic peptide of gp110. Rescreening of the same library with the cDNA clones identified a full-length cDNA clone for this glycoprotein. Sequence analysis indicates that the N-linked carbohydrate chains are concentrated on the N-terminal part of this highly glycosylated protein.
Neurosteroids: oligodendrocyte mitochondria convert cholesterol to pregnenolone.
Authors: Hu Z Y , Bourreau E , Jung-Testas I , Robel P , Baulieu E E
Published: May 2006

Oligodendrocyte mitochondria from 21-day-old Sprague-Dawley male rats were incubated with 100 nM [3H]cholesterol. It yielded [3H]pregnenolone at a rate of 2.5 +/- 0.7 and 5-[3H]pregnene-3 beta, 20 alpha-diol at a rate of 2.5 +/- 1.1 pmol per mg of protein per hr. Cultures of glial cells from 19- to 21-day-old fetuses (a mixed population of astrocytes and oligodendrocytes) were incubated for 24 hr with [3H]mevalonolactone. [3H]Cholesterol, [3H]pregnenolone, and 5-[3H]pregnene-3 beta, 20 alpha-diol were characterized in cellular extracts. The formation of the 3H-labeled steroids was increased by dibutyryl cAMP (0.2 mM) added to the culture medium. The active cholesterol side-chain cleavage mechanism, recently suggested immunohistochemically and already observed in cultures of C6 glioma cells, reinforces the concept of "neurosteroids" applied to delta 5-3 beta-hydroxysteroids previously isolated from brain.
Azidothymidine: crystal structure and possible functional role of the azido group.
Authors: Camerman A , Mastropaolo D , Camerman N
Published: May 2006

The crystal and molecular structures of the anti-acquired immunodeficiency syndrome agent 3'-azido-3'-deoxythymidine (AZT) have been determined by x-ray diffraction. There are two crystallographically independent AZT molecules in the crystal asymmetric unit; they have similar conformations and differ primarily in the glycosyl torsion angle. Comparisons with a hydrated thymidylate structure indicate that the azido group does not significantly affect the gross conformational preference of the molecule. The comparisons also suggest possible functional roles for the azido group in enzyme binding.
Molecular cloning and sequence analysis of the Plasmodium falciparum dihydrofolate reductase-thymidylate synthase gene.
Authors: Bzik D J , Li W B , Horii T , Inselburg J
Published: May 2006

Genomic DNA clones that coded for the bifunctional dihydrofolate reductase (DHFR) and thymidylate synthase (TS) (DHFR-TS) activities from a pyrimethamine-sensitive strain of Plasmodium falciparum were isolated and sequenced. The deduced DHFR-TS protein contained 608 amino acids (71,682 Da). The coding region for DHFR-TS contained no intervening sequences and had a high A + T content (75%). The DHFR domain, in the amino-terminal portion of the protein, was joined by a 94-amino acid junction sequence to the TS domain in the carboxyl-terminal portion of the protein. The TS domain was more conserved than the DHFR domain and both P. falciparum domains were more homologous to eukaryotic than to prokaryotic forms of the enzymes. Predicted secondary structures of the DHFR and TS domains were nearly identical to the structures identified in other DHFR and TS enzymes.
Sec53, a protein required for an early step in secretory protein processing and transport in yeast, interacts with the cytoplasmic surface of the endoplasmic reticulum.
Authors: Ruohola H , Ferro-Novick S
Published: May 2006

The sec53 mutant is a conditional lethal yeast secretory mutant. At 37 degrees C, precursors to exported proteins become firmly attached to the endoplasmic reticulum membrane and are not released into the lumen in a soluble form. The accumulated precursors are insoluble in the detergent Triton X-100; however, urea, a known protein denaturant, solubilizes them. Using antibody directed against the Sec53 protein, we found that a substantial portion of the Sec53 protein is associated with the cytoplasmic surface of the endoplasmic reticulum membrane. Membrane-bound Sec53 protein is largely insoluble in Triton X-100, but the protein is effectively released from the membrane by urea. We propose that the Sec53 protein may be a member of a complex of proteins required for an early step in protein processing and transport.
Evolution of intrafamilial interactions.
Authors: Lynch M
Published: May 2006

A theory for the evolution of behavioral interactions among relatives is developed that allows for genetic correlations between the types of behavior that are expressed in different social contexts. Both theoretical and empirical considerations indicate that such genetic constraints will almost certainly be common in natural populations. It is shown that when genetic correlations between elements of social behavior exist, Hamilton's rule inaccurately describes the conditions for evolution by way of kin selection. The direction in which social organization evolves is a delicate function of the genetic covariance structure among behaviors expressed as an offspring, sibling, parent, etc. A change in this covariance structure caused by random genetic drift or by a change in environment for a population exhibiting genotype-environment interaction can cause the population to suddenly cross a threshold into a new selective domain. Consequently, radical changes in social organization may arise between closely related species without any major shift in selective pressures external to the population.
Characterization of pHV2 from Halobacterium volcanii and its use in demonstrating transformation of an archaebacterium.
Authors: Charlebois R L , Lam W L , Cline S W , Doolittle W F
Published: May 2006

We determined the complete nucleotide sequence of the 6354-base-pair plasmid pHV2 of the archaebacterium Halobacterium volcanii. This plasmid is present in approximately six copies per chromosome. We have generated a strain, H. volcanii WFD11, cured of pHV2 by treatment of liquid cultures with ethidium bromide. We describe PEG-mediated transformation of H. volcanii WFD11 with intact pHV2 and with a form of pHV2 marked by a 93-base-pair deletion generated in vitro.
Rhizobium meliloti has three functional copies of the nodD symbiotic regulatory gene.
Authors: Honma M A , Ausubel F M
Published: May 2006

We have identified two Rhizobium meliloti genes (nodD2 and nodD3) that are highly homologous and closely linked to the regulatory gene nodD (nodD1). R. meliloti strains containing mutations in the three nodD genes in all possible combinations were constructed and their nodulation phenotypes were assayed on Medicago sativa (alfalfa) and Melilotus alba (sweet clover). A triple nodD1-nodD2-nodD3 mutant exhibited a Nod- phenotype on alfalfa and sweet clover, indicating that nodD is an essential nodulation gene in R. meliloti. A nodD2 mutant exhibited no discernable defect in nodulation and nodD3 mutants exhibited a delayed nodulation phenotype of 2-3 days when inoculated onto either host. Alfalfa nodules elicited by a nodD1 mutant appeared 5-6 days after wild-type nodules, and sweet clover nodules elicited by a nodD1 mutant appeared 2-3 days after wild-type nodules. nodD1-nodD2 double mutants formed nodules with the same delay as single nodD1 mutants on both hosts. nodD2-nodD3 double mutants elicited sweet clover nodules at the same rate as single nodD3 mutants, but this same double mutant was slightly more delayed in alfalfa nodule formation than the nodD3 mutant. The nodD1-nodD3 mutant exhibited an extremely delayed nodulation phenotype on alfalfa and elicited no nodules on sweet clover. These experiments indicate that nodD1 and nodD3 have equivalent roles in nodulating sweet clover but that nodD1 plays a more important role than nodD3 in eliciting nodules on alfalfa. The nodD2 gene appears to have some effect on alfalfa nodulation and none on sweet clover. Our results indicate that R. meliloti has three functional nodD genes that modulate the nodulation process in a host-specific manner.
Serological responses in chimpanzees inoculated with human immunodeficiency virus glycoprotein (gp120) subunit vaccine.
Authors: Arthur L O , Pyle S W , Nara P L , Bess J W , Gonda M A , Kelliher J C , Gilden R V , Robey W G , Bolognesi D P , Gallo R C
Published: May 2006

The major envelope glycoprotein of a human immunodeficiency virus (HIV) has been purified and was utilized as a prototype vaccine in chimpanzees. The 120,000-dalton glycoprotein (gp120) was purified from membranes of human T-lymphotropic virus (HTLV)-IIIB-infected cells and the final preparation contained low levels to no detectable HTLV-IIIB core antigen (p24) and low levels of endotoxin. Chimpanzees inoculated with gp120 responded by developing antibodies that precipitated radiolabeled gp120 and neutralized in vitro infection of HTLV-IIIB. Antibodies to HTLV-IIIB p24 were not detected in the gp120-immunized chimpanzees. Peripheral blood leukocytes from the vaccinated animals were examined for T4+ and T8+ cells, and no decrease in the T4/T8 ratio was found, indicating that immunization with a ligand (gp120) that binds to T4 has no detectable adverse effect on the population of T4+ cells. The only current animal model that can be reproducibly infected with HIV is the chimpanzee. Immunization of chimpanzees with HIV proteins will provide an experimental system for testing the effectiveness of prototype vaccines for preventing HIV infection in vivo.
Chemical kinetic measurements of a mammalian acetylcholine receptor by a fast-reaction technique.
Authors: Udgaonkar J B , Hess G P
Published: May 2006

In the presence of acetylcholine, the nicotinic acetylcholine receptor undergoes two rapid conformational changes: one in the 1-ms time region, leading to the formation of a transmembrane channel and signal transmission between cells, and the other in the 100-ms time region, leading to an inactive "desensitized" form with altered ligand-binding properties. To determine the properties of the receptor that are relevant for channel opening and signal transmission, we have developed a cell-flow technique that allows measurements to be made with cells prior to receptor desensitization. Here we illustrate the usefulness of the technique. A wide concentration range of both a ligand that controls the opening of receptor channels (carbamoylcholine) and a receptor inhibitor (procaine) was used to measure the dissociation constant of the receptor site controlling channel opening (2.4 X 10(-4) M), the channel-opening equilibrium constant (5.5), the inhibition constant for procaine (5.8 X 10(-5) M), and the rate coefficients for two desensitization processes of 5 s-1 and 0.2 s-1. The cell-flow technique illustrated here is of interest because, by rapid-reaction techniques, it extends the chemical kinetic approach from investigations of reactions in solutions to investigations of many different receptors that exist in membranes of central nervous system cells and whose properties are not well known.
Comparative mutational analysis of wild-type and stretched tRNA3(Leu) gene promoters.
Authors: Fabrizio P , Coppo A , Fruscoloni P , Benedetti P , Di Segni G , Tocchini-Valentini G P
Published: May 2006

We demonstrate that, when the yeast tRNA(3Leu) gene is stretched so that the distance between the two portions of the intragenic promoter is increased to 365 base pairs, the A and B blocks remain functional. Mutations in the A block, which show a weak phenotype when inserted in the wild type, exert a dramatic effect when inserted into the stretched gene. Experiments with extensively purified transcription factor tau indicate that the tau B-B block interaction is not influenced by A-B distance; only the ability of tau A to interact with A block sequences is affected, possibly because of the additional free-energy cost of forming a large loop of the intervening DNA.
Expression of yeast DNA topoisomerase I can complement a conditional-lethal DNA topoisomerase I mutation in Escherichia coli.
Authors: Bjornsti M A , Wang J C
Published: May 2006

We show that, despite differences in primary structure, substrate preference, and mechanism of catalysis, yeast DNA topoisomerase I can functionally substitute for Escherichia coli DNA topoisomerase I. A family of plasmids expressing the yeast TOP1 gene or 5'-deletion mutations of it were used to complement the temperature-sensitive phenotype of an E. coli topA mutant. These plasmids were then isolated from the cells by a rapid lysis procedure and examined for their degrees of supercoiling. Functional complementation of a conditional-lethal mutation in topA, which encodes E. coli DNA topoisomerase I, correlates with the expression of a catalytically active yeast enzyme that reduces the degree of negative supercoiling of intracellular DNA. We also show that approximately 130 amino acids of the amino-terminal portion of the yeast enzyme can be deleted without affecting its activity in vitro; activity of the enzyme inside E. coli, however, is more sensitive to such deletions.
Release of soluble pilin antigen coupled with gene conversion in Neisseria gonorrhoeae.
Authors: Haas R , Schwarz H , Meyer T F
Published: May 2006

Gene conversion appears to be the frequent mechanism in Neisseria gonorrhoeae that leads to an altered expression of pilin, the subunit component of the pili. In this process segments of variable sequence information, the minicassettes, are transferred from silent storage loci into an expression locus. As a putative consequence of the rearrangement in the pilE gene, gonococci can enter a different phase of pilin production. Although the removal of a 7-amino acid leader peptide results in the production of typical P+ pilin used to form pili, the loss of an additional 39 amino acids yields S-pilin, a soluble form of pilin that is efficiently secreted into the extracellular environment. Both pilin types can coexist in an apparently homogeneous culture. Ps cells usually are piliated, although less extensively with regard to the length and the number of the pili when compared with P+ cells. Ps cells form T3/T4-type colonies also typical of nonpiliated cells (P-). The observations further suggest that the classical nonsecretory P- phenotype is not generated as a rule by precise gene conversion but rather by genetic changes that cause the production of an over-length pilin (L-pilin).
Genetic recombination within the human T-cell receptor alpha-chain gene complex.
Authors: Robinson M A , Kindt T J
Published: May 2006

Genetic analyses of the human T-cell receptor (TCR) alpha-chain genes indicate that recombination events may occur frequently within this gene complex. Examination of the inheritance of restriction fragment length polymorphisms (RFLP) detected by using probes for constant or variable region gene segments made it possible to assign TCR alpha haplotypes to the 16 parents and 43 offspring of eight families studied. A total of six RFLP, three for the constant region and three for variable region gene segments, were examined in the present studies. Most enzyme and probe combinations tested revealed no polymorphism and those finally selected for the study showed limited polymorphism in that only two or, in one case, three allelic forms of the gene were seen. In spite of limited variability at this level, extensive heterogeneity was observed for the combinations of markers present in haplotypes, suggesting that frequent recombination events have occurred. Most strikingly, multiple combinations of RFLP occurring in close proximity of the TCR alpha constant region gene were observed in this study. A high recombination frequency for the TCR alpha gene complex is further supported by the observation that two children, one in each of two families, inherited recombinant TCR alpha haplotypes.
A human major histocompatibility complex class I gene that encodes a protein with a shortened cytoplasmic segment.
Authors: Geraghty D E , Koller B H , Orr H T
Published: May 2006

We have cloned genomic DNA encoding a non-HLA-A, -B, -C class I gene located within a HindIII-generated restriction fragment of 6.0 kilobase pairs. This gene, designated HLA-6.0, is as homologous to HLA-A and HLA-B as they are to each other. The HLA class I protein encoded by HLA-6.0 is similar in organization to the HLA-A-, -B-, and -C-encoded proteins except that an in-frame termination codon prevents translation of a majority of the cytoplasmic region of the HLA-6.0 polypeptide. Moreover, the promoter region of HLA-6.0 resembles the promoter region of a Qa region gene. These structural features of HLA-6.0 suggest that this non-HLA-A, -B, -C gene is a structural homolog of a murine Qa region class I gene.
RAS gene mutations in acute and chronic myelocytic leukemias, chronic myeloproliferative disorders, and myelodysplastic syndromes.
Authors: Janssen J W , Steenvoorden A C , Lyons J , Anger B , Böhlke J U , Bos J L , Seliger H , Bartram C R
Published: May 2006

We report on investigations aimed at detecting mutated RAS genes in a variety of preleukemic disorders and leukemias of myeloid origin. DNA transfection analyses (tumorigenicity assay) and hybridization to mutation-specific oligonucleotide probes established NRAS mutations in codon 12 or 61 of 4/9 acute myelocytic leukemias (AML) and three AML lines. Leukemic cells of another AML patient showed HRAS gene activation. By using a rapid and sensitive dot-blot screening procedure based on the combination of in vitro amplification of RAS-specific sequences and oligonucleotide hybridization we additionally screened 15 myelodysplastic syndromes, 26 Philadelphia chromosome-positive chronic myelocytic leukemias in chronic or acute phase, and 19 other chronic myeloproliferative disorders. A mutation within NRAS codon 12 could thus be demonstrated in a patient with idiopathic myelofibrosis and in another with chronic myelomonocytic leukemia. Moreover, mutated NRAS sequences were detected in lymphocytes, in granulocytes, as well as in monocytes/macrophages of the latter case.
Specific expression of the pS2 gene in subclasses of breast cancers in comparison with expression of the estrogen and progesterone receptors and the oncogene ERBB2.
Authors: Rio M C , Bellocq J P , Gairard B , Rasmussen U B , Krust A , Koehl C , Calderoli H , Schiff V , Renaud R , Chambon P
Published: May 2006

The expression of the pS2 gene, which is induced by estrogen in the breast cancer cell line MCF-7, has been investigated in breast cancers by using pS2 mRNA determination in tumor specimens and immunocytochemistry to identify pS2 protein in paraffin-embedded sections. Using these assays we show that determination of pS2 gene expression allows the definition of subclasses of estrogen-receptor-containing breast cancers that may be used to more precisely identify estrogen-dependent tumors. Tumor specimens have also been analyzed for the presence of mRNAs for the estrogen receptor and for the ERBB2 oncogene. No evidence for the presence of truncated forms of estrogen-receptor mRNA has been found, and overexpression of the ERBB2 oncogene did not correlate with the steroid receptor status or pS2 gene expression.
Loss of heterozygosity on chromosomes 3, 13, and 17 in small-cell carcinoma and on chromosome 3 in adenocarcinoma of the lung.
Authors: Yokota J , Wada M , Shimosato Y , Terada M , Sugimura T
Published: May 2006

By a molecular genetic approach using polymorphic DNA markers that detect allelic deletion of specific chromosomal regions, we analyzed for possible loss of chromosomal heterozygosity in five different histological types of lung cancers obtained from 47 patients. In small-cell carcinomas, the incidence of allelic deletions at three different chromosomal loci was extremely high; loss of heterozygosity was detected on chromosomes 3p in 7 of 7 patients (100%), 13q in 10 of 11 patients (91%), and 17p in 5 of 5 patients (100%). The deletions at these loci in small-cell carcinomas were observed even in the tumors without any clinical evidence of metastasis. Furthermore, loss of heterozygosity on chromosomes 3p and 13q occurred prior to NMYC amplification and chromosome 11p deletion. Loss of heterozygosity on chromosome 3p was also detected with high frequency in adenocarcinomas [5 of 6 patients (83%)]. Heterozygosity of chromosomes 13q and 17p was lost in 10 of 31 patients (32%) and in 3 of 12 patients (25%), respectively, of lung cancers other than small-cell carcinomas. These results indicate that recessive genetic changes involving sequences on chromosomes 3p, 13q, and 17p may play important roles in the genesis of small-cell carcinoma, and those on chromosome 3p may play an important role in the genesis of adenocarcinoma.
RETRACTED: Loss of (2'-5')oligoadenylate synthetase activity by production of antisense RNA results in lack of protection by interferon from viral infections.
Authors: De Benedetti A , Pytel B A , Baglioni C
Published: May 2006

An expression vector was constructed that carries part of the human BK papovavirus with 0.5 kilobases of (2'-5')oligoadenylate (2-5A) synthetase cDNA inserted in inverted orientation downstream from the virion proteins (VP) promoter and the neomycin-resistance gene neo under the control of a simian virus 40 promoter. Cells transfected with this vector and selected for resistance to the neomycin derivative G418 synthesized RNA complementary to 2-5A synthetase mRNA. These cells lacked 2-5A synthetase activity, and the enzyme was not inducible by interferon. In contrast, 2-5A synthetase was induced in cells transfected with a control vector without the cDNA insert. Such cells were protected by interferon from RNA viruses, whereas cells lacking 2-5A synthetase were not protected from encephalomyocarditis virus, vesicular stomatitis virus, and Sindbis virus but were fully protected from influenza virus. These findings show that a high level of 2-5A synthetase is required for interferon-induced protection from the cytoplasmic RNA viruses tested.
Asymmetry and external noise-induced free energy transduction.
Authors: Chen Y D
Published: May 2006

The effect of fluctuations in rate constants on the kinetic behavior of cyclic reacting systems, caused by a fluctuating external parameter, is studied. It is shown (i) that the stochastic properties of the system can be analyzed analytically by using the usual master equation approach when the external parameter is fluctuating with discrete square pulses and (ii) that the system is equivalent to an expanded chemical kinetic system with no fluctuation in rate constants. When applied to a linear four-state cyclic enzyme system, the formalism can be used to prove analytically the finding that an enzyme can extract the free energy from an externally applied fluctuating membrane potential and perform active free energy transduction. The formalism also can be used to assess the asymmetry constraints imposed on the values of the rate constants in order for the model to work.
Phosphorylation of a high molecular weight DNA polymerase alpha.
Authors: Donaldson R W , Gerner E W
Published: May 2006

Anti-human DNA polymerase alpha murine IgG SJK-287-38 [Tanaka, S., Hu, S.-Z., Wang, T. S.-F. & Korn, D. (1982) J. Biol. Chem. 257, 8386-8390] neutralized DNA polymerase alpha activity from rat embryonic fibroblasts infected with a temperature-sensitive transformation mutant of Rous sarcoma virus (tsLA24). After centrifugation of a crude cytosol fraction from log-phase cells in a 5-20% linear sucrose gradient, polypeptides of Mr approximately equal to 185,000 and 220,000 were immunoprecipitated only from gradient fractions containing DNA polymerase alpha activity. When similar cultures were incubated in medium containing [32P]orthophosphate, it was found that the Mr 220,000 protein was phosphorylated but that the other peptides specific for polymerase alpha activity did not contain detectable amounts of phosphate. Phospho amino acid analysis of the high molecular weight immunoprecipitable proteins indicated that the labeled amino acid was phosphoserine. Incubation of 2.5 units of crude DNA polymerase alpha with 4 units of agarose-immobilized alkaline phosphatase resulted in a nearly complete inhibition of DNA polymerase alpha activity. Subsequent incubation of this preparation with 5 or 50 microM ATP, but not the nonhydrolyzable analog adenosine 5'-[gamma-thio]triphosphate, restored the in vitro DNA polymerizing activity. These results demonstrate that a high molecular weight DNA polymerase alpha (Mr approximately equal to 220,000) is phosphorylated in cultured cells and that this protein is a substrate for a serine kinase rather than the tyrosine-specific protein kinase of Rous sarcoma virus. The results suggest that phosphorylation/dephosphorylation reactions modulate the activity of this polymerase.