bluuebunny/crossref_metadata_2025 huggingface dataset:
Molecular Dynamics of a Cytochrome c-Cytochrome b
5
Electron Transfer Complex
Authors: Wendoloski J. J. , Matthew James B. , Weber P. C. , Salemme F. R.
Published: October 2006

Cytochrome c and cytochrome b
5
form an electrostatically associated electron transfer complex. Computer models of this and related complexes that were generated by docking the x-ray structures of the individual proteins have provided insight into the specificity and mechanism of electron transfer reactions. Previous static modeling studies were extended by molecular dynamics simulations of a cytochrome c-cytochrome b
5
intermolecular complex. The simulations indicate that electrostatic interactions at the molecular interface result in a flexible association complex that samples alternative interheme geometries and molecular conformations. Many of these transient geometries appear to be more favorable for electron transfer than those formed in the initial model complex. Of particular interest is a conformational change that occurred in phenylalanine 82 of cytochrome c that allowed the phenyl side chain to bridge the two cytochrome heme groups.
Transposon Tagging and Molecular Analysis of the Maize Regulatory Locus
opaque-2
Authors: Schmidt Robert J. , Burr Frances A. , Burr Benjamin
Published: October 2006

Genetic analyses suggested that the
opaque-2
(
o2
) locus in maize acts as a positive, transacting, transcriptional activator of the zein seed storage-protein genes. Because isolation of the gene is requisite to understanding the molecular details of this regulation, transposon mutagenesis with the transposable element
suppressor-mutator
(
Spm
) was carried out, and three mutable
o2
alleles were obtained. One of these alleles contained an 8.3-kilobase autonomous
Spm
, another a 6.8-kilobase nonautonomous
Spm
, and the third an unidentified transposon that is unrelated to
Spm
. A DNA sequence flanking the autonomous
Spm
insertion was verified to be
o2
-specific and provided a probe to clone a wild-type allele. Northern blots indicated that the gene is expressed in wild-type endosperm but not in leaf tissues or in endosperms homozygous for a mutant allele of the
O2
gene. A transcript was detected in endosperms homozygous for mutations at
opaque-7
and
floury-2
, an indication that
O2
expression is independent of these two other putative regulators of zein synthesis.
A Gene for Dihydrofolate Reductase in a Herpesvirus
Authors: Trimble John J. , Murthy Shridhara C. S. , Bakker Anke , Grassmann Ralph , Desrosiers Ronald C.
Published: October 2006

The enzyme dihydrofolate reductase (DHFR) is found ubiquitously in both prokaryotes and eukaryotes. It is essential for de novo synthesis of purines and of deoxythymidine monophosphate for DNA synthesis. Among viruses, however, only the T-even and T5 bacteriophage have been found to encode their own DHFR. In this study a gene for DHFR was found in a specific subgroup of the gamma or lymphotropic class of herpesviruses. DNA sequences for DHFR were found in herpesvirus saimiri and herpesvirus ateles but not in Epstein-Barr virus, Marek's disease virus, herpes simplex virus, varicella-zoster virus, herpesvirus tamarinus, or human cytomegalovirus. The predicted sequence of herpesvirus saimiri DHFR is 186 amino acids in length, the same length as human, murine, and bovine DHFR. The human and herpesvirus saimiri DHFRs share 83 percent positional identity in amino acid sequence. The herpesvirus saimiri DHFR gene is devoid of intron sequences, suggesting that it was acquired by some process involving reverse transcription. This is to our knowledge the first example of a mammalian virus with a gene for DHFR.
Biologic Features of HIV-1 That Correlate with Virulence in the Host
Authors: Cheng-Mayer Cecilia , Seto Deborah , Tateno Masatoshi , Levy Jay A.
Published: October 2006

Individuals infected with the human immunodeficiency virus type 1 (HIV-1) may be asymptomatic or have AIDS-related complex or the acquired immuno deficiency syndrome (AIDS). Little is known about the factors that influence progression of infection to AIDS. In this study of isolates of HIV-1 obtained at intervals during the infection of four individuals, the development of disease was found to be correlated with the emergence of HIV-1 variants that were more cytopathic in vitro as the disease progressed and that replicated more efficiently in a wide variety of different human cells. The biologic properties of HIV-1 in vitro thus appear to reflect its virulence in the host. Further studies of such sequentially isolated viruses may lead to the identification of viral genes that govern pathogenesis.
Genetic and Crystallographic Studies of the 3′,5′-Exonucleolytic Site of DNA Polymerase I
Authors: Derbyshire Victoria , Freemont Paul S. , Sanderson Mark R. , Beese Lorena , Friedman Jonathan M. , Joyce Catherine M. , Steitz Thomas A.
Published: October 2006

Site-directed mutagenesis of the large fragment of DNA polymerase I (Klenow fragment) yielded two mutant proteins lacking 3′,5′-exonuclease activity but having normal polymerase activity. Crystallographic analysis of the mutant proteins showed that neither had any alteration in protein structure other than the expected changes at the mutation sites. These results confirmed the presumed location of the exonuclease active site on the small domain of Klenow fragment and its physical separation from the polymerase active site. An anomalous scattering difference Fourier of a complex of the wild-type enzyme with divalent manganese ion and deoxythymidine monophosphate showed that the exonuclease active site has binding sites for two divalent metal ions. The properties of the mutant proteins suggest that one metal ion plays a role in substrate binding while the other is involved in catalysis of the exonuclease reaction.
DNAs of the Two Mating-Type Alleles of
Neurospora crassa
Are Highly Dissimilar
Authors: Glass N. Louise , Vollmer Steven J. , Staben Chuck , Grotelueschen Jeff , Metzenberg Robert L. , Yanofsky Charles
Published: October 2006

The mating-type alleles
A
and
a
of
Neurospora crassa
control mating in the sexual cycle and function in establishing heterokaryon incompatibility in the vegetative cycle. The
A
and
a
alleles were cloned, and they were shown to encode both the sexual functions and vegetative incompatibility. The mating-type clones contain nonhomologous DNA segments that are flanked by common DNA sequences.
Neurospora crassa
and all heterothallic and pseudohomothallic
Neurospora
species contain a single copy of one mating-type sequence or the other within each haploid genome. The six known self-fertile homothallic isolates contain an
A
homolog, but only one species also contains
a
homologous sequences. Homothallism in these species is not due to mating-type switching, as it is in
Saccharomyces cerevisiae
.
DNA Binding by Proteins
Authors: Schleif Robert
Published: October 2006

Study of proteins that recognize specific DNA sequences has yielded much information, but the field is still in its infancy. Already two major structural motifs have been discovered, the helix-turn-helix and zinc finger, and numerous examples of DNA-binding proteins containing either of them are known. The restriction enzyme Eco RI uses yet a different motif. Additional motifs are likely to be found as well. There is a growing understanding of some of the physical chemistry involved in protein-DNA binding, but much remains to be learned before it becomes possible to engineer a protein that binds to a specific DNA sequence.
Insulin-Stimulated Release of Lipoprotein Lipase by Metabolism of Its Phosphatidylinositol Anchor
Authors: Chan Betty Liwah , Lisanti Michael P. , Rodriguez-Boulan Enrique , Saltiel Alan R.
Published: October 2006

Lipoprotein lipase (LPL) plays a critical role in the metabolism of plasma lipoproteins. In 3T3-L1 adipocytes, insulin elicits the rapid release of LPL through mechanisms that are independent of energy metabolism and protein synthesis. Some of the metabolic actions of insulin may be mediated by the activation of a specific phospholipase that hydrolyzes a glycosyl phosphatidylinositol (PI) molecule. The insulin-sensitive glycosyl-PI is structurally similar to the glycolipid membrane anchor of a number of proteins. LPL appears to be anchored to the 3T3-L1 cell surface by glycosyl-PI, and its rapid release by insulin may be due to activation of a glycosyl-PI-specific phospholipase C.
Heterogeneity of Glycine Receptors and Their Messenger RNAs in Rat Brain and Spinal Cord
Authors: Akagi Hiroyuki , Miledi Ricardo
Published: October 2006

Messenger RNAs isolated from adult or newborn rat spinal cord were fractionated in a sucrose gradient. The fractions were injected into
Xenopus
oocytes to determine their potencies for expression of glycine receptors (GlyRs), which were then examined electrophysiologically. The sedimentation profiles disclosed two classes of GlyR mRNAs, one heavy and the other light. The adult spinal cord was rich in heavy GlyR mRNA, whereas the light GlyR mRNA was more abundant in neonatal spinal cord and in adult cerebral cortex. Glycine receptors encoded by heavy and light mRNAs of adult spinal cord showed some electrophysiological differences. Thus there are two types of GlyRs encoded by mRNAs of different sizes, and the expression of these mRNAs is developmentally regulated. A tissue- and age-dependent distribution of heterogeneous GlyR mRNAs may imply diverse roles of the GlyRs in neuronal function in the central nervous system.
Two Cytosolic Neutrophil Oxidase Components Absent in Autosomal Chronic Granulomatous Disease
Authors: Volpp Bryan D. , Nauseef William M. , Clark Robert A.
Published: October 2006

Neutrophils kill microorganisms with oxygen radicals generated by an oxidase that uses the reduced form of nicotinamide adenine dinucleotide phosphate (NADPH) as substrate. This system requires both membrane and cytosolic components and is defective in patients with chronic granulomatous disease. A cytosolic complex capable of activating latent membrane oxidase was eluted from guanosine triphosphate-agarose and was used to raise polyclonal antiserum that recognized 47- and 67-kilodalton proteins. These proteins were restricted to the cytosol of myeloid cells. Both proteins were associated with NADPH oxidase-activating capacity when neutrophil cytosol was purified on nucleotide affinity matrices or molecular sizing columns. Neutrophils from patients with two different forms of autosomal chronic granulomatous disease lacked either the 47- or 67-kilodalton protein.
Lineage-Specific Development of Calcium Currents During Embryogenesis
Authors: Simoncini Luciana , Block Melodye L. , Moody William J.
Published: October 2006

The development of electrophysiological properties of isolated, identified ascidian blastomeres was followed from the fertilized egg to the neurula, and the stage at which cells of different lineages first express different functional ion channel populations was determined. Little has been known about such events because of the difficulties of making voltage-clamp recordings from small embryonic cells and of identifying their developmental fates in dissociated preparations. The problem of small cell size was circumvented by using the whole-cell patch clamp, and identification was facilitated by the use of a species of ascidian,
Boltenia villosa
, in which endogenous pigment marks cells of specific developmental fates. Within approximately 3 hours after gastrulation, muscle-lineage blastomeres in these embryos developed a voltage-dependent calcium current while surrounding blastomeres of other lineages did not. At about the same time, all cells developed delayed outward potassium currents and lost the inwardly rectifying potassium currents present at earlier stages.
Restoration of Torque in Defective Flagellar Motors
Authors: Blair David F. , Berg Howard C.
Published: October 2006

Paralyzed motors of
motA
and
motB
point and deletion mutants of
Escherichia coli
were repaired by synthesis of wild-type protein. As found earlier with a point mutant of
motB
, torque was restored in a series of equally spaced steps. The size of the steps was the same for both MotA and MotB. Motors with one torque generator spent more time spinning counterclockwise than did motors with two or more generators. In deletion mutants, stepwise decreases in torque, rare in point mutants, were common. Several cells stopped accelerating after eight steps, suggesting that the maximum complement of torque generators is eight. Each generator appears to contain both MotA and MotB.
Atriopeptin-Immunoreactive Neurons in the Brain: Presence in Cardiovascular Regulatory Areas
Authors: Saper Clifford B. , Standaert David G. , Currie Mark G. , Schwartz David , Geller David M. , Needleman Philip
Published: October 2006

Antisera to atriopeptin III and to a cyanogen bromide fragment of the precursor molecule atriopeptigen were prepared and used to examine the distribution of atriopeptin-like immunoreactive material in the heart and brain of the rat. Granules of this material were seen in myocytes throughout the right and left atria and were densest in the perinuclear region. The distribution of atriopeptin-like immunoreactive material in the heart is consistent with previous reports of atrial secretory granules. In the brain neurons containing the material were observed in the hypothalamus and the pontine tegmentum. Atriopeptin in the brain may serve as a neurotransmitter in neural systems controlling blood volume and composition, the same physiological functions regulated by blood-borne atriopeptin.
Homeo Boxes in the Study of Development
Authors: Gehring Walter J.
Published: October 2006

The body plan of
Drosophila
is determined to a large extent by homeotic genes, which specify the identity and spatial arrangement of the body segments. Homeotic genes share a characteristic DNA segment, the homeo box, which encodes a defined domain of the homeotic proteins. The homeo domain seems to mediate the binding to specific DNA sequences, whereby the homeotic proteins exert a gene regulatory function. By isolating the normal
Antennapedia
gene, fusing its protein-coding sequences to an inducible promoter, and reintroducing this fusion gene into the germline of flies, it has been possible to transform head structures into thoracic structures and to alter the body plan in a predicted way. Sequence homologies suggest that similar genetic mechanisms may control development in higher organisms.
Homozygosity Mapping: A Way to Map Human Recessive Traits with the DNA of Inbred Children
Authors: Lander Eric S. , Botstein David
Published: October 2006

An efficient strategy for mapping human genes that cause recessive traits has been devised that uses mapped restriction fragment length polymorphisms (RFLPs) and the DNA of affected children from consanguineous marriages. The method involves detection of the disease locus by virtue of the fact that the adjacent region will preferentially be homozygous by descent in such inbred children. A single affected child of a first-cousin marriage is shown to contain the same total information about linkage as a nuclear family with three affected children. Calculations show that it should be practical to map a recessive disease gene by studying DNA from fewer than a dozen unrelated, affected inbred children, given a complete RFLP linkage map. The method should make it possible to map many recessive diseases for which it is impractical or impossible to collect adequate numbers of families with multiple affected offspring.
Three Recessive Loci Required for Insulin-Dependent Diabetes in Nonobese Diabetic Mice
Authors: Prochazka Michal , Leiter Edward H. , Serreze David V. , Coleman Douglas L.
Published: October 2006

A polygenic basis for susceptibility to insulin-dependent diabetes in nonobese diabetic (NOD) mice has been established by outcross to a related inbred strain, nonobese normal (NON). Analysis of first and second backcross progeny has shown that at least three recessive genes are required for development of overt diabetes. One,
Idd-1
s
, is tightly linked to the H-2K locus on chromosome 17; another,
Idd-2
s
, is localized proximal to the
Thy-1
/
Alp-1
cluster on chromosome 9. Segregation of a third,
Idd-3
s
, could be shown in a second backcross. Neither
Idd-1
s
nor
Idd-2
s
could individually be identified as the locus controlling insulitis; leukocytic infiltrates in pancreas were common in most asymptomatic BC1 mice. Both F1 and BC1 mice exhibited the unusually high percentage of splenic T lymphocytes characteristic of NOD, suggesting dominant inheritance of this trait. The polygenic control of diabetogenesis in NOD mice, in which a recessive gene linked to the major histocompatibility complex is but one of several controlling loci, suggests that similar polygenic interactions underlie this type of diabetes in humans.
Absence of TGF-β Receptors and Growth Inhibitory Responses in Retinoblastoma Cells
Authors: Kimchi Adi , Wang Xiao-Fan , Weinberg Robert A. , Cheifetz Sela , Massagué Joan
Published: October 2006

The responses of retinoblastoma tumor cells and normal retinal cells to various growth inhibitory factors were examined. Whereas fetal retinal cells were highly sensitive to the antimitogenic effects of transforming growth factor β1 (TGF-β1), retinoblastoma tumor cell lines were all resistant to this factor. Binding assays and affinity labeling of these cells with radioiodinated TGF-β1 revealed that the cells did not have TGF-β receptors. The retinoblastoma cells lacked the three affinity-labeled proteins of 65, 95, and 300 kilodaltons typically seen in human cell lines and thus differed from normal retinal cells and from other types of neuroectodermal tumors that display the normal pattern of receptors. Loss of TGF-β receptors, which is a rare event among tumor cells, may represent one mechanism through which these cells escape from negative control and form retinoblastomas.
Living with Clathrin: Its Role in Intracellular Membrane Traffic
Authors: Brodsky Frances M.
Published: October 2006

Clathrin polymerization at the cytoplasmic side of the plasma membrane forms coated pits and vesicles that mediate uptake of cell surface receptors. Clathrin-coated vesicles have also been implicated in protein export but definition of their precise role has been controversial. Recent advances in characterization of the clathrin subunits and additional coated vesicle components have identified molecular interactions involved in clathrin polymerization and coated vesicle formation, and have provided new approaches to investigating its function. These studies suggest that clathrin's role, in both inward and outward membrane traffic, is to facilitate receptor transport by a concentration and sorting process that initiates targeting to specific intracellular compartments.
Mental Rotation of the Neuronal Population Vector
Authors: Georgopoulos Apostolos P. , Lurito Joseph T. , Petrides Michael , Schwartz Andrew B. , Massey Joe T.
Published: October 2006

A rhesus monkey was trained to move its arm in a direction that was perpendicular to and counterclockwise from the direction of a target light that changed in position from trial to trial. Solution of this problem was hypothesized to involve the creation and mental rotation of an imagined movement vector from the direction of the light to the direction of the movement. This hypothesis was tested directly by recording the activity of cells in the motor cortex during performance of the task and computing the neuronal population vector in successive time intervals during the reaction time. The population vector rotated gradually counterclockwise from the direction of the light to the direction of the movement at an average rate of 732° per second. These results provide direct, neural evidence for the mental rotation hypothesis and indicate that the neuronal population vector is a useful tool for "reading out" and identifying cognitive operations of neuronal ensembles.
Direct Observation of Native DNA Structures with the Scanning Tunneling Microscope
Authors: Beebe Thomas P. , Wilson Troy E. , Ogletree D. Frank , Katz Joseph E. , Balhorn Rod , Salmeron Miquel B. , Siekhaus Wigbert J.
Published: October 2006

Uncoated double-stranded DNA dissolved in a salt solution was deposited on graphite and imaged in air with the scanning tunneling microscope (STM). The resolution was such that the major and minor grooves could be distinguished. The pitch of the helix varied between 27 and 63 angstroms in the images obtained. Thus the STM can be useful for structural studies of a variety of uncoated and isolated biomolecules.
Amyloid β Protein Precursor Is Possibly a Heparan Sulfate Proteoglycan Core Protein
Authors: Schubert David , Schroeder Rolly , LaCorbiere Monique , Saitoh Tsunao , Cole Greg
Published: October 2006

The amyloid β protein peptide is a major constituent of amyloid plaque cores in Alzheimer's disease and is apparently derived from a higher molecular weight precursor. It is now shown that the core protein of a heparan sulfate proteoglycan secreted from a nerve cell line (PC12) has an amino acid sequence and a size very similar to those of the amyloid β protein precursor and that these molecules are antigenically related. This amyloid β protein precursor-related protein is not found in the conditioned medium of a variant cell line (F3 PC12) that does not secrete heparan sulfate proteoglycan. The synaptic localization and metabolism of this class of proteoglycans are consistent with its potential involvement in central nervous system dysfunction.
Sequence Homology and Morphologic Similarity of HTLV-III and Visna Virus, a Pathogenic Lentivirus
Authors: Gonda Matthew A. , Wong-Staal Flossie , Gallo Robert C. , Clements Janice E. , Narayan Opendra , Gilden Raymond V.
Published: October 2006

A study was conducted of the genetic relation between human T-cell lymphotropic retroviruses and visna virus. The human T-cell lymphotropic viruses include those associated with T-cell malignancies (HTLV-I and HTLV-II) as well as the etiologic agent of the acquired immune deficiency syndrome (HTLV-III). Visna virus, a slowly replicating and pathogenic but nononcogenic retrovirus of sheep, is a member of the subfamily Lentivirinae. Results obtained by molecular hybridization and heteroduplex analysis indicated that a greater extent of nucleotide sequence homology exists between HTLV-III and visna virus than between HTLV-III and any of the other viruses. The homology observed under conditions of low stringency spanned the entire genome, but was strongest in the
gag
/
pol
region. The morphogenesis and fine structure of HTLV-III and visna virus also demonstrated striking similarities. The data provide strong evidence for a close taxonomic and thus evolutionary relation between HTLV-III and the Lentivirinae subfamily.
HTLV-III Infection in Brains of Children and Adults with AIDS Encephalopathy
Authors: Shaw George M. , Harper Mary E. , Hahn Beatrice H. , Epstein Leon G. , Gajdusek D. Carleton , Price Richard W. , Navia Bradford A. , Petito Carol K. , O'Hara Carl J. , Groopman Jerome E. , Cho Eun-Sook , Oleske James M. , Wong-Staal Flossie , Gallo Robert C.
Published: October 2006

Unexplained debilitating dementia or encephalopathy occurs frequently in adults and children with the acquired immune deficiency syndrome (AIDS). Brains from 15 individuals with AIDS and encephalopathy were examined by Southern analysis and in situ hybridization for the presence of human T-cell leukemia (lymphotropic) virus type III (HTLV-III), the virus believed to be the causative agent of AIDS. HTLV-III DNA was detected in the brains of five patients, and viral-specific RNA was detected in four of these. In view of these findings and the recent demonstration of morphologic and genetic relatedness between HTLV-III and visna virus, a lentivirus that causes a chronic degenerative neurologic disease in sheep, HTLV-III should be evaluated further as a possible cause of AIDS encephalopathy.
Suppression of Gamma Interferon Production by Inactivated Feline Leukemia Virus
Authors: Engelman R. W. , Fulton R. W. , Good R. A. , Day N. K.
Published: October 2006

Supernatants from cultures of normal feline lymphocytes stimulated with
Staphylococcus
enterotoxin A showed antiviral activity, characterized as a gamma-like interferon. With the addition of inactivated feline leukemia virus, markedly less interferon was produced. The reduction in interferon production was not attributable to lowered lymphocyte viability or reduced mitogenic properties of
Staphylococcus
enterotoxin A and appears to be a direct retroviral effect. This finding may reflect clinically relevant events that may contribute to the development of the feline or human states of acquired immunodeficiency.
Bond Order and Charge Localization in Nucleoside Phosphorothioates
Authors: Frey Perry A. , Sammons R. Douglas
Published: October 2006

In the recent literature on nucleoside phosphorothioate anions the structural formulas show a double bond between phosphorus and sulfur and a single bond between phosphorus and oxygen with a negative charge localized on oxygen. However, a review of physical data on these compounds shows the reverse to be the case; that is, in phosphorothioate anions the P-S bond is a single bond with a negative charge localized on sulfur, while the P-O bond order for exocyclic and nonbridging oxygens is greater than 1, approaching 2 in O-alkyl phosphorothioate monoanions and O,O-dialkyl phosphorothioates. The P-O bond orders in phosphorothioate dianions and trianions approach 1 ½ and 1 ⅓, respectively, owing to delocalization of negative charge among two or three oxygens. These conclusions are based on bond lengths obtained from x-ray crystallographic data and electron diffraction, the magnitudes of the effects of
18
O on the
31
P-nuclear magnetic resonance chemical shifts of phosphorus in nucleoside [
18
O]phosphorothioates, the
p
H -dependence of
17
O-NMR chemical shifts in [
17
O]phosphate and [
17
O]thiophosphate, the vibrational spectra of thiophosphate di- and trianions, and the
pK
a
(dissociation constant) values for phosphoric and thiophosphoric acids.
Transcription of Novel Open Reading Frames of AIDS Retrovirus During Infection of Lymphocytes
Authors: Rabson Arnold B. , Daugherty Daryl F. , Venkatesan Sundararajan , Boulukos Kim E. , Benn Steven I. , Folks Thomas M. , Feorino Paul , Martin Malcolm A.
Published: October 2006

The retrovirus frequently isolated from patients with the acquired immune deficiency syndrome (AIDS) has two novel open reading frames previously designated "A" and "B." The "A" region was found to be specifically expressed as polyadenylated RNA's of 5.5 and 5.0 kilobases in infected cells. The "B" region was expressed as 1.8- to 2.0-kilobase RNA species. Additional full-length and spliced messenger RNA's of the
env
region were also identified.
Wounding and Its Role in RSV-Mediated Tumor Formation
Authors: Dolberg David S. , Hollingsworth Robert , Hertle Mark , Bissell Mina J.
Published: October 2006

Tumors induced in chickens by Rous sarcoma virus remain localized at the site of injection even though the animals become viremic. Tumors have now been shown to be inducible at other sites if a wound is inflicted or if the tissue is injured by administration of tumor promoters. These findings indicate that local wounding plays a role in the spread of tumorigenicity of Rous sarcoma virus.
Iron(II) EDTA Used to Measure the Helical Twist Along Any DNA Molecule
Authors: Tullius Thomas D. , Dombroski Beth A.
Published: October 2006

A new method has been devised to measure the number of base pairs per helical turn along any DNA molecule in solution. A DNA restriction fragment is adsorbed onto crystalline calcium phosphate, fragmented by reaction with iron(II) EDTA, and subjected to electrophoresis on a denaturing polyacrylamide gel. A modulated cutting pattern results, which gives directly the helical periodicity of the DNA molecule. A 150-base pair sequence directly upstream of the thymidine kinase gene of the type 1 herpes simplex virus was found to have an overall helical twist of 10.5 base pairs per turn, which is characteristic of the
B
conformation of DNA. In addition, purines 3′ to pyrimidines showed lower than expected reactivity toward the iron cutting reagent, which is evidence for sequence-dependent variability in DNA conformation.
Hypoglycemia-Induced Neuronal Damage Prevented by an
N
-Methyl-D-Aspartate Antagonist
Authors: Wieloch Tadeusz
Published: October 2006

The possibility that neuronal damage due to hypoglycemia is induced by agonists acting on the
N
-methyl-D-aspartate (NMDA) receptor was investigated in the rat caudate nucleus. Local injections of an NMDA receptor antagonist, 2-amino-7-phosphonoheptanoic acid, were performed before induction of 30 minutes of reversible, insulin-induced, hypoglycemic coma. Neuronal necrosis in these animals after 1 week of recovery was reduced 90 percent compared to that in saline-injected animals. The results suggest that hypoglycemic neuronal damage is induced by NMDA receptor agonists, such as the excitatory amino acids or related compounds.
Sodium-Specific Membrane Channels of Frog Skin Are Pores: Current Fluctuations Reveal High Turnover
Authors: Lindemann B. , Van Driessche W.
Published: October 2006

The reversible sodium transport blocker amiloride causes current fluctuations at the apical membrane of the outer stratum granulosum of frog skin. Their power density spectra reveal that single transport sites translocate more than 10
6
sodium ions per second, which indicates a pore mechanism. The density of open plus amiloride-blocked pores is in the order of 10
8
pores per square centimeter of skin area with 60 millimolar sodium and 18 micromolar amiloride in the outer solution.
Expression of the
Escherichia coli
lacZ
Gene on a Plasmid Vector in a Cyanobacterium
Authors: Buzby Jeffrey S. , Porter Ronald D. , Stevens S. Edward
Published: October 2006

A biphasic plasmid vector was used to introduce the
Escherichia coli
K-12 lac operon into the unicellular cyanobacterium Agmenellum quadruplicatum PR-6. The PR-6 transformants expressed β-galactosidase at nearly as high a level as did Escherichia coli transformants. In order to accomplish this, it was necessary to obtain PR-6 mutants that could be transformed by plasmids with unmodified recognition sites for the endogenous PR-6 restriction endonuclease Aqu I. These mutants were generated by a variation of the ectopic mutagenesis techniques that have been used in other naturally transforming bacteria. The ability to assay the expression of lacZ in PR-6 paves the way for the construction of gene fusions with various PR-6 promoters and quantitation of their expression under specific in vivo conditions.
The Product of the Human c-
erb
B-2 Gene: a 185-Kilodalton Glycoprotein with Tyrosine Kinase Activity
Authors: Akiyama Tetsu , Sudo Chie , Ogawara Hiroshi , Toyoshima Kumao , Yamamoto Tadashi
Published: October 2006

Antibodies were raised against a synthetic peptide corresponding to 14 amino acid residues at the COOH-terminus of a protein deduced from the human c-
erb
B-2 nucleotide sequence. These antibodies immunoprecipitated a 185-kilodalton glycoprotein from MKN-7 adenocarcinoma cells. Incubation of the immunoprecipitates with (γ-
32
)ATP resulted in the phosphorylation of this protein on tyrosine residues. These results indicate that the human c-
erb
B-2 gene product is the 185-kilodalton glycoprotein that is associated with tyrosine kinase activity. Although the c-
erb
B-2 protein was predicted to encode a protein very similar to epidermal growth factor (EGF) receptor, EGF did not stimulate this kinase activity either in vivo or in vitro.
Tandem Regions of Yeast DNA Topoisomerase II Share Homology with Different Subunits of Bacterial Gyrase
Authors: Lynn Richard , Giaever Guri , Swanberg Stephen L. , Wang James C.
Published: October 2006

The nucleotide sequence for the
Saccharomyces cerevisiae
gene TOP2, which encodes DNA topoisomerase II, was compared with the sequence for bacterial DNA gyrase. The amino and carboxyl terminal halves of the single-subunit yeast enzyme showed homologies with the B and A subunits of bacterial gyrase, respectively, at corresponding positions along the polypeptide chains. Although the two enzymes differ in both quaternary structure and activity, the homology between the two proteins indicates mechanistic as well as structural similarities, and a probable evolutionary relationship.
Metals and DNA: Molecular Left-Handed Complements
Authors: Barton Jacqueline K.
Published: October 2006

Chiral metal complexes provide unique molecular probes for DNA. Chiral reagents that "recognize" different local structures along the DNA strand have been designed by a process in which the asymmetry in shape and size of the complex is matched to that of the DNA helical groove. As a result, the chiral metal complexes provide very sensitive probes for local helical structure, both left- and right-handed. Direct coordination of chiral complexes to the DNA bases adds an element of sequence selectivity to the probe design. With a suitable reactive metal center, reagents that target chemically specific sites along the strand may be developed. One such chiral reagent, which cleaves left-handed DNA sites with photoactivation, has been useful in mapping this distinct conformation and examining its biological role. The conformation-specific molecular cleaver, much like a DNA-binding enzyme, recognizes and reacts at discrete sites along the DNA strand. These site-specific chiral metal complexes provide exciting new tools for probing the local variations in DNA structure and its role in the regulation of gene expression.
Control of a Cell Surface Major Glycoprotein by Epidermal Growth Factor
Authors: Chen Lan Bo , Gudor Robert C. , Sun Tung-Tien , Chen Anthony B. , Mosesson Michael W.
Published: October 2006

When the serum concentration of the culture medium is below 0.7 percent, 3T3 mouse cells lose most of their large external transformation sensitive (LETS) protein at the cell surface, Subsequent addition of epidermal growth factor results in the reappearance of massive fibrillar LETS protein networks on the surface of confluent 3T3 cells. Thirteen other hormones tested do not have this effect. It appears that epidermal growth factor can control the expression of LETS protein.
The Metabolism of Phosphoinositide-Derived Messenger Molecules
Authors: Majerus Philip W. , Connolly Thomas M. , Deckmyn Hans , Ross Theodora S. , Bross Teresa E. , Ishii Hidemi , Bansal Vinay S. , Wilson David B.
Published: October 2006

The phosphoinositides are minor phospholipids present in all eukaryotic cells. They are storage forms for messenger molecules that transmit signals across the cell membrane and evoke responses to extracellular agonists. The phosphoinositides break down to liberate messenger molecules or precursors of messenger molecules. Many different compounds are formed, although the functions of only a few are understood. Recent studies elaborating the pathways for formation of products from phosphoinositides and the factors controlling their metabolism are summarized here.
The Regulation of Natural Anticoagulant Pathways
Authors: Esmon Charles T.
Published: October 2006

Vascular endothelium plays an active role in preventing blood clot formation in vivo. One mechanism by which prevention is achieved involves a cell surface thrombin-binding protein, thrombomodulin, which converts thrombin into a protein C activator. Activated protein C then functions as an anticoagulant by inactivating two regulatory proteins of the coagulation system, factors Va and VIIIa. The physiological relevance of the protein C anticoagulant pathway is demonstrated by the identification of homozygous protein C-deficient infants with severe thrombotic complications. Recent studies suggest that this pathway provides a link between inflammation and coagulation.
Transformation of Rat Fibroblasts by FSV Rapidly Increases Glucose Transporter Gene Transcription
Authors: Birnbaum Morris J. , Haspel Howard C. , Rosen Ora M.
Published: October 2006

Elevation of glucose transport is an alteration common to most virally induced tumors. Rat fibroblasts transformed with wild-type or a temperature-sensitive Fujinami sarcoma virus (FSV) were studied in order to determine the mechanisms underlying the increased transport. Five- to tenfold increases in total cellular glucose transporter protein in response to transformation were accompanied by similar increases in transporter messenger RNA levels. This, in turn, was preceded by an absolute increase in the rate of glucose transporter gene transcription within 30 minutes after shift of the temperature-sensitive FSV-transformed cells to the permissive temperature. The transporter messenger RNA levels in transformed fibroblasts were higher than those found in proliferating cells maintained at the nonpermissive temperature. The activation of transporter gene transcription by transformation represents one of the earliest known effects of oncogenesis on the expression of a gene encoding a protein of well-defined function.
Ubiquitin Is a Component of Paired Helical Filaments in Alzheimer's Disease
Authors: Mori Hiroshi , Kondo Jun , Ihara Yasuo
Published: October 2006

Paired helical filaments (PHF), which constitute a distinct type of pathological neuronal fiber, are the principal constituent of neurofibrillary tangles that occur in the brain of patients with Alzheimer's disease. Their insolubility in sodium dodecyl sulfate and urea has prevented the analysis of their subunit composition by gel electrophoresis. A monoclonal antibody (DF2) was isolated that specifically labeled PHF at both the light and electron microscopic levels. It labeled a small polypeptide (5 kilodaltons) that was shown to be ubiquitin in immunoblots of the soluble fraction of brain homogenates. To obtain direct evidence that ubiquitin is a component of PHF, PHF were treated with concentrated formic acid and digested with lysylendopeptidase; ubiquitin-derived peptides were then identified by reversed-phase high-performance liquid chromatography. Two fragments in the PHF digest were identified as derived from ubiquitin by protein sequencing. This procedure should make possible definitive identification of other PHF components.
Visualization of Viral Clearance in the Living Animal
Authors: Verdin Eric M. , Maratos-Flier Eleftheria , Kahn C. Ronald , Sodoyez Jean-Claude , Sodoyez-Goffaux Francoise , De Vos Claudine J. , Lynn Steven P. , Fields Bernard N.
Published: October 2006

The early events in viral dissemination via the bloodstream were identified by monitoring the fate of
123
I-radiolabeled reovirus after it was injected intravenously in rats. Continuous scintillation camera imaging showed that reovirus serotypes 1 and 3 were cleared from the circulation in less than 10 minutes by specific and distinct target organs. Reovirus serotype 1 accumulated predominantly in the lungs and the liver, whereas serotype 3 accumulated in the liver and the spleen with very little virus uptake by the lungs. Incubation of reovirus serotype 1 with a monoclonal antibody directed against the viral hemagglutinin before injection totally inhibited the clearance of the virus by the lungs. Similar results were obtained when viruses biolabeled with
35
S were used. These results demonstrate that viruses can be rapidly transported through the bloodstream to specific target organs and that the localization of the viruses depends on the interaction between specific viral surface components and the target organ.
Divalent Cations Directly Affect the Conductance of Excised Patches of Rod Photoreceptor Membrane
Authors: Stern J. H. , Knutsson Hans , MacLeish P. R.
Published: October 2006

Phototransduction in rod cells is likely to involve an intracellular messenger system that links the absorption of light by rhodopsin to a change in membrane conductance. The direct effect of guanosine 3′,5′-monophosphate (cGMP) on excised patches of rod outer segment membrane strongly supports a role for cGMP as an intracellular messenger in phototransduction. It is reported here that magnesium and calcium directly affect the conductance of excised patches of rod membrane in the absence of cGMP and that magnesium, applied to intact rod cells, blocks a component of the cellular light response. The divalent cation-suppressed conductance in excised patches showed outward rectification and cation-selective permeability resembling those of the light-suppressed conductance measured from the intact rod cell. The divalent cation-suppressed conductance was partly blocked by a concentration of the pharmacological agent L-
cis
-diltiazem that blocked all of the cGMP-activated conductance. Divalent cations may act, together with cGMP, as an intracellular messenger system that mediates the light response of the rod photoreceptor cell.
Selective Disruption of Gap Junctional Communication Interferes with a Patterning Process in Hydra
Authors: Fraser Scott E. , Green Colin R. , Bode Hans R. , Gilula Norton B.
Published: October 2006

The cells that make up the body column of hydra are extensively joined by gap junctions, capable of mediating the rapid exchange of small hydrophilic molecules between the cytoplasms of neighboring cells. Both the rate of transfer of small molecules through the gap junctions and the rate of return of gap junction coupling after grafting experiments are sufficiently rapid to mediate events in the patterning of hydra tissue. Antibodies to the major rat liver gap junction protein (27,000 daltons) recognize a gap junction antigen in hydra and are effective in eliminating junctional communication between hydra cells. The antibodies perturb the head inhibition gradient in grafting operations, suggesting that cell-cell communication via gap junctions is important in this defined tissue patterning process.
Identification of a Family of Muscarinic Acetylcholine Receptor Genes
Authors: Bonner T. I. , Buckley N. J. , Young A. C. , Brann M. R.
Published: October 2006

Complementary DNAs for three different muscarinic acetylcholine receptors were isolated from a rat cerebral cortex library, and the cloned receptors were expressed in mammalian cells. Analysis of human and rat genomic clones indicates that there are at least four functional muscarinic receptor genes and that these genes lack introns in the coding sequence. This gene family provides a new basis for evaluating the diversity of muscarinic mechanisms in the nervous system.
Mammalian Glucocorticoid Receptor Derivatives Enhance Transcription in Yeast
Authors: Schena Mark , Yamamoto Keith R.
Published: October 2006

In mammalian cells, the glucocorticoid receptor binds specifically to glucocorticoid response element (GRE) DNA sequences and enhances transcription from linked promoters. It is shown here that derivatives of the glucocorticoid receptor also enhance transcription when expressed in yeast. Receptor-mediated enhancement in yeast was observed in fusions of GRE sequences to the yeast cytochrome c
1
(
CYC1
) promoter; the
CYC1
upstream activator sequences were not essential, since enhancement was observed in fusions of GREs to mutant
CYC1
promoters retaining only the TATA region and transcription startpoints. It is concluded that the receptor operates by a common, highly conserved mechanism in yeast and mammalian cells.
Irreversible Inhibition of Sodium Entry Sites in Frog Skin by a Photosensitive Amiloride Analog
Authors: Benos D. J. , Mandel L. J.
Published: October 2006

A photosensitive binding reaction is described in which an analog of amiloride is bound to sites that control sodium entry into frog skin. This reaction results in irreversible inhibition of net sodium transport.
Human IL-3 and GM-CSF Act Synergistically in Stimulating Hematopoiesis in Primates
Authors: Donahue Robert E. , Seehra Jasbir , Metzger Mark , Lefebvre Denise , Rock Brian , Carbone Suzanne , Nathan David G. , Garnick Marc , Sehgal Prabhat K. , Laston David , LaVallie Edward , McCoy John , Schendel Paul F. , Norton Christine , Turner Katherine , Yang Yu-Chang , Clark Steven C.
Published: October 2006

Interleukin-3 (IL-3) is a member of a family of growth factors, each of which supports the proliferation and development of hematopoietic precursors in culture. Although the biologic effects of the different hematopoietic growth factors have been well documented in different culture systems, it has only recently become possible to study the activities of these molecules in vivo. In comparison with the later acting hematopoietic growth factors granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor, IL-3 elicited a delayed and relatively modest leukocytosis when continuously infused intravenously in primates. The IL-3 infusion, however, greatly potentiated the responsiveness of the animal to subsequent administration of a low dose of GM-CSF. These results suggest that IL-3 expands an early cell population in vivo that subsequently requires the action of a later acting factor such as GM-CSF to complete its development. Optimal stimulation of hematopoiesis may be achieved with combinations of hematopoietic growth factors.
The Human Immune Response to the OKT3 Monoclonal Antibody Is Oligoclonal
Authors: Chatenoud Lucienne , Jonker Margreet , Villemain Françoise , Goldstein Gideon , Bach Jean-François
Published: October 2006

The availability of highly specific and homogeneous antibodies to human T cells by the hybridoma technique has elicited new interest in the clinical use of antibodies to lymphocytes as immunosuppressive agents. OKT3 is the murine monoclonal antibody that has been the most widely used in clinical transplantation to induce immunosuppression. This antibody recognizes a membrane molecular complex, exclusively present on mature human T lymphocytes, which is tightly linked to the T-cell antigen receptor. The long-term therapeutic use of murine monoclonal antibodies in vivo is hampered by the intense antibody response that occurs in most human patients. Thus, when administered alone, OKT3 manifests its immunosuppressive activity only during the 10 to 15 days that precede the onset of sensitization. The results presented here show, by use of isoelectrofocusing, that the antibody response to OKT3, already reported to be restricted in its specificity (only anti-isotypic and anti-idiotypic antibodies are produced), is in addition oligoclonal. This restriction of the anti-monoclonal response may suggest that an efficient way to circumvent the sensitization problem would be to administer consecutively different monoclonal antibodies presenting the same specificity but distinct idiotypes.
Cytotoxicity of Human
p
I 7 Interleukin-1 for Pancreatic Islets of Langerhans
Authors: Bendtzen Klaus , Mandrup-Poulsen Thomas , Nerup Jørn , Nielsen Jens H. , Dinarello Charles A. , Svenson Morten
Published: October 2006

Activated mononuclear cells appear to be important effector cells in autoimmune beta cell destruction leading to insulin-dependent (type 1) diabetes mellitus. Conditioned medium from activated mononuclear cells (from human blood) is cytotoxic to isolated rat and human islets of Langerhans. This cytotoxic activity was eliminated from crude cytokine preparations by adsorption with immobilized, purified antibody to interleukin-1 (IL-1). The islet-inhibitory activity and the IL-1 activity (determined by its comitogenic effect on thymocytes) were recovered by acid wash. Purified natural IL-1 and recombinant IL-1 derived from the predominant
p
I 7 form of human IL-1, consistently inhibited the insulin response. The
p
I 6 and
p
I 5 forms of natural IL-1 were ineffective. Natural and recombinant IL-1 exhibited similar dose responses in their islet-inhibitory effect and their thymocyte-stimulatory activity. Concentrations of IL-1 that inhibited islet activity were in the picomolar range. Hence, monocyte-derived
p
I 7 IL-1 may contribute to islet cell damage and therefore to the development of insulin-dependent diabetes mellitus.