bluuebunny/crossref_metadata_2025 huggingface dataset:
Transient Pioneer Neurons Are Essential for Formation of an Embryonic Peripheral Nerve
Authors: Klose Monika , Bentley David
Published: October 2006

In developing nervous systems, many peripheral and central pathways are established by early arising populations of pioneer neurons. The growth cones of these pioneer neurons can migrate while embryonic distances are short and while intervening tissue is relatively uncomplicated. Are these pioneers necessary? In grasshopper embryos, a pair of pioneer neurons arise at the tips of limb buds and extend axons through the limb to the central nervous system. Growth cones of later arising sensory neurons migrate along the pioneer axons. After ingrowth of sensory axons, the pioneer neurons die. If the pioneer neurons are prevented from differentiating by heat shock, then the sensory growth cones that would have migrated along them are blocked and fail to reach the central nervous system. Thus, the pioneer axons are necessary for successful migration of these sensory growth cones. By crossing a segment boundary early in embryogenesis, the pioneers circumvent an incompatibility between differentiated segment boundary cells and growth cone migration. Pioneer neurons may resolve similar problems in many systems.
Molecular Structure of DNA by Scanning Tunneling Microscopy
Authors: Cricenti A. , Selci S. , Felici A. C. , Generosi R. , Gori E. , Djaczenko W. , Chiarotti G.
Published: October 2006

Uncoated DNA molecules marked with an activated tris(1-aziridinyl) phosphine oxide (TAPO) solution were deposited on gold substrates and imaged in air with the use of a high-resolution scanning tunneling microscope (STM). Constant-current and gap-modulated STM images show clear evidence of the helicity of the DNA structure: pitch periodicity ranges from 25 and 35 angstroms, whereas the average diameter is 20 angstroms. Molecular structure within a single helix turn was also observed.
Interleukin-1 Mitogenic Activity for Fibroblasts and Smooth Muscle Cells Is Due to PDGF-AA
Authors: Raines Elaine W. , Dower Steven K. , Ross Russell
Published: October 2006

Both interleukin-1 (IL-1) and platelet-derived growth factor (PDGF) induce proliferation of cultured fibroblasts and smooth muscle cells. These polypeptide mediators are released by activated macrophages and other cell types in response to injury and are thought to have a role in tissue remodeling and a number of pathologic processes. Analysis of the kinetics of [
3
H]thymidine incorporation by cultured fibroblasts demonstrated that the response to IL-1 is delayed approximately 8 hours relative to their response to PDGF. IL-1 transiently stimulated expression of the PDGF A-chain gene, with maximum induction after approximately 2 hours. Subsequent synthesis and release of PDGF activity into the medium was detected as early as 4 hours after IL-1 stimulation, and downregulation of the binding site for the PDGF-AA isoform of PDGF followed PDGF-AA secretion. Antibodies to PDGF completely block the mitogenic response to IL-1. Therefore, the mitogenic activity of IL-1 for fibroblasts and smooth muscle cells appears to be indirect and mediated by induction of the PDGF A-chain gene.
Human Genotoxicity: Pesticide Applicators and Phosphine
Authors: Garry Vincent F. , Griffith Jack , Danzl Thomas J. , Nelson Richard L. , Whorton Elbert B. , Krueger Lisa A. , Cervenka Jaroslav
Published: October 2006

Fumigant applicators who, 6 weeks to 3 months earlier, were exposed to phosphine, a common grain fumigant, or to phosphine and other pesticides had significantly increased stable chromosome rearrangements, primarily translocations in G-banded lymphocytes. Less stable aberrations including chromatid deletions and gaps were significantly increased only during the application season, but not at this later time point. During fumigant application, measured exposure to phosphine exceeds accepted national standards. Because phosphine is also used as a dopant in the microchip industry and is generated in waste treatment, the possibility of more widespread exposure and long-term health sequelae must be considered.
Myristoylated and Nonmyristoylated Forms of a Protein Are Phosphorylated by Protein Kinase C
Authors: Graff Jonathan M. , Gordon Jeffrey I. , Blackshear Perry J.
Published: October 2006

Activation of protein kinase C is thought to require association of the kinase with the cell membrane. It has been assumed that cellular substrates for the kinase must likewise be associated with membranes, and previous studies with membrane-associated myristoylated proteins have supported this view. It is now shown that a mutation that prevents the normal amino-terminal myristoylation of a prominent cellular substrate of protein kinase C, and appears to prevent its membrane association, does not prevent the normal phosphorylation of this protein in intact cells in response to phorbol esters. Thus, membrane association may not be required in order for protein kinase C substrates to undergo phosphorylation.
Chemical Conversion of a DNA-Binding Protein into a Site-Specific Nuclease
Authors: Chen Chi-Hong , Sigman David S.
Published: October 2006

The tryptophan gene (
trp
) repressor of
Escherichia coli
has been converted into a site-specific nuclease by covalently attaching it to the 1,10-phenanthroline-copper complex. In its cuprous form, the coordination complex with hydrogen peroxide as a coreactant cleaves DNA by oxidatively attacking the deoxyribose moiety. The chemistry for the attachment of 1,10-phenanthroline to the
trp
repressor involves modification of lysyl residues with iminothiolane followed by alkylation of the resulting sulfhydryl groups with 5-iodoacetamido-1,10-phenanthroline. The modified
trp
repressor cleaves the operators of
aroH
and
trpEDCBA
upon the addition of cupric ion and thiol in a reaction dependent on the corepressor L-tryptophan. Scission was restricted to the binding site for the repressor, defined by deoxyribonuclease I footprinting. Since DNA-binding proteins have recognition sequences approximately 20 base pairs long, the nucleolytic activities derived from them could be used to isolate long DNA fragments for sequencing or chromosomal mapping.
Two Mutant Alleles of the Insulin Receptor Gene in a Patient with Extreme Insulin Resistance
Authors: Kadowaki Takashi , Bevins Charles L. , Cama Alessandro , Ojamaa Kaie , Marcus-Samuels Bernice , Kadowaki Hiroko , Beitz Laurie , McKeon Catherine , Taylor Simeon I.
Published: October 2006

Insulin receptor complementary DNA has been cloned from an insulin-resistant patient with leprechaunism whose receptors exhibited multiple abnormalities in insulin binding. The patient is a compound heterozygote, having inherited two different mutant alleles of the insulin receptor gene. One allele contains a missense mutation encoding the substitution of glutamic acid for lysine at position 460 in the α subunit of the receptor. The second allele has a nonsense mutation causing premature chain termination after amino acid 671 in the α subunit, thereby deleting both the transmembrane and tyrosine kinase domains of the receptor. Interestingly, the father is heterozygous for this nonsense mutation and exhibits a moderate degree of insulin resistance. This raises the possibility that mutations in the insulin receptor gene may account for the insulin resistance in some patients with non-insulin-dependent diabetes mellitus.
Postsynaptic Calcium Is Sufficient for Potentiation of Hippocampal Synaptic Transmission
Authors: Malenka Robert C. , Kauer Julie A. , Zucker Robert S. , Nicoll Roger A.
Published: October 2006

Brief repetitive activation of excitatory synapses in the hippocampus leads to an increase in synaptic strength that lasts for many hours. This long-term potentiation (LTP) of synaptic transmission is the most compelling cellular model in the vertebrate brain for learning and memory. The critical role of postsynaptic calcium in triggering LTP has been directly examined using three types of experiment. First, nitr-5, a photolabile nitrobenzhydrol tetracarboxylate calcium chelator, which releases calcium in response to ultraviolet light, was used. Photolysis of nitr-5 injected into hippocampal CA1 pyramidal cells resulted in a large enhancement of synaptic transmission. Second, in agreement with previous results, buffering intracellular calcium at low concentrations blocked LTP. Third, depolarization of the postsynaptic membrane so that calcium entry is suppressed prevented LTP. Taken together, these results demonstrate that an increase in postsynaptic calcium is necessary to induce LTP and sufficient to potentiate synaptic transmission.
The Genetic Defect Causing Familial Alzheimer's Disease Maps on Chromosome 21
Authors: St George-Hyslop Peter H. , Tanzi Rudolph E. , Polinsky Ronald J. , Haines Jonathan L. , Nee Linda , Watkins Paul C. , Myers Richard H. , Feldman Robert G. , Pollen Daniel , Drachman David , Growdon John , Bruni Amala , Foncin Jean-Francois , Salmon Denise , Frommelt Peter , Amaducci Luigi , Sorbi Sandro , Piacentini Silva , Stewart Gordon D. , Hobbs Wendy J. , Conneally P. Michael , Gusella James F.
Published: October 2006

Alzheimer's disease is a leading cause of morbidity and mortality among the elderly. Several families have been described in which Alzheimer's disease is caused by an autosomal dominant gene defect. The chromosomal location of this defective gene has been discovered by using genetic linkage to DNA markers on chromosome 21. The localization on chromosome 21 provides an explanation for the occurrence of Alzheimer's disease-like pathology in Down syndrome. Isolation and characterization of the gene at this locus may yield new insights into the nature of the defect causing familial Alzheimer's disease and possibly, into the etiology of all forms of Alzheimer's disease.
Coexistence of Guanylate Cyclase and Atrial Natriuretic Factor Receptor in a 180-kD Protein
Authors: Paul Aranjaniyil K. , Marala Ravi B. , Jaiswal Rama Kant , Sharma Rameshwar K.
Published: October 2006

Atrial natriuretic factor (ANF) is a peptide hormone that is released from atria and regulates a number of physiological processes, including steroidogenesis in adrenal cortex and testes. The parallel stimulation of membrane guanylate cyclase and corticosterone production in isolated fasciculata cells of rat adrenal cortex has supported the hypothesis of a mediatory role for cyclic guanosine monophosphate (cyclic GMP) in signal transduction. A novel particulate guanylate cyclase tightly coupled with ANF receptor was purified approximately 273,000-fold by two-step affinity chromatography. The enzyme had a molecular size of 180 kilodaltons and was acidic in nature with a
p
I of 4.7. Its specific activity was 1800 nanomoles of cyclic GMP formed per minute per milligram of protein. The purified enzyme bound ANF with a specific binding activity of 4.01 nanomoles per milligram of protein, a value that is close to the theoretical binding activity of 5.55 nanomoles per milligram of protein for 1 mole of the ligand binding 1 mole of the receptor protein. These results indicate that the guanylate cyclase-coupled ANF receptor exists in a 180-kilodalton protein of rat adrenocortical carcinoma and represent a step toward the elucidation of the basic mechanism of cyclic GMP-mediated transmembrane signal transduction in response to a hormone.
Full-Wave Rectification from a Mixed Electrical-Chemical Synapse
Authors: Graubard Katherine , Hartline Daniel K.
Published: October 2006

Electrical and chemical synapses usually reinforce one another, but the pyloric late-to-lateral pyloric (PL-to-LP) neuronal connections in lobster stomatogastric ganglia create an inverted U-shaped transfer function between the two neurons: regardless of whether the PL membrane voltage swings positive or negative, the postsynaptic LP voltage will go negative. When the presynaptic cell voltage goes negative, the effect on the LP voltage is due to electrical coupling. During positive presynaptic voltages, the strong contribution of graded chemical inhibition from the PL to the LP neuron overrides the positive electrical coupling to produce net negativity.
Expression of the β-Nerve Growth Factor Gene in Hippocampal Neurons
Authors: Ayer-LeLievre Christiane , Olson Lars , Ebendal Ted , Seiger Åke , Persson Håkan
Published: October 2006

In situ hybridization with complementary DNA probes for nerve growth factor (NGF) was used to identify cells containing NGF messenger RNA in rat and mouse brain. The most intense labeling occurred in hippocampus, where hybridizing neurons were found in the dentate gyrus and the pyramidal cell layer. The neuronal identity of NGF mRNA-containing cells was further assessed by a loss of NGF-hybridizing mRNA in hippocampal areas where neurons had been destroyed by kainic acid or colchicine. RNA blot analysis also revealed a considerable decrease in the level of NGF mRNA in rat dentate gyrus after a lesion was produced by colchicine. This lesion also caused a decrease in the level of Thy-1 mRNA and an increase in the level of glial fibrillary acidic protein mRNA. Neuronal death was thus associated with the disappearance of NGF mRNA. These results suggest a synthesis of NGF by neurons in the brain and imply that, in hippocampus, NGF influences NGF-sensitive neurons through neuron-to-neuron interactions.
Long-Term Synaptic Potentiation
Authors: Brown Thomas H. , Chapman Paul F. , Kairiss Edward W. , Keenan Claude L.
Published: October 2006

Long-term synaptic potentiation (LTP) is a leading candidate for a synaptic mechanism of rapid learning in mammals. LTP is a persistent increase in synaptic efficacy that can be quickly induced. The biophysical process that controls one type of LTP is formally similar to a synaptic memory mechanism postulated decades ago by the psychologist Donald Hebb. A key aspect of the modification process involves the
N
-methyl-D-aspartate (NMDA) receptor-ionophore complex. This ionophore allows calcium influx only if the endogenous ligand glutamate binds to the NMDA receptor and if the voltage across the associated channel is also sufficiently depolarized to relieve a magnesium block. According to one popular hypothesis, the resulting increase in the intracellular calcium concentration activates protein kinases that enhance the postsynaptic conductance. Further biophysical and molecular understanding of the modification process should facilitate detailed explorations of the mnemonic functions of LTP.
Control of Enzyme Activity by an Engineered Disulfide Bond
Authors: Matsumura Masazumi , Matthews Brian W.
Published: October 2006

A novel approach to the control of enzyme catalysis is presented in which a disulfide bond engineered into the active-site cleft of bacteriophage T4 lysozyme is capable of switching the activity on and off. Two cysteines (Thr
21
→ Cys and Thr
142
→ Cys) were introduced by oligonucleotide-directed mutagenesis into the active-site cleft. These cysteines spontaneously formed a disulfide bond under oxidative conditions in vitro, and the catalytic activity of the oxidized (cross-linked) T4 lysozyme was completely lost. On exposure to reducing agent, however, the disulfide bond was rapidly broken, and the reduced (non-cross-linked) lysozyme was restored to full activity. Thus an enzyme has been engineered such that redox potential can be used to control catalytic activity.
Reduction of Intestinal Carcinogen Absorption by Carcinogen-Specific Secretory Immunity
Authors: Silbart Lawrence K. , Keren David F.
Published: October 2006

A secretory immune response to the carcinogen 2-acetylaminofluorene (AAF) was elicited in rabbits by directly immunizing the small intestine with an AAF-cholera toxin conjugate. High-titer, high-affinity secretory immunoglobulin A (IgA) antibody to AAF was secreted into the intestinal lumen in response to this immunogen. Immune secretions reduced the transepithelial absorption of a
125
I-labeled derivative of AAF by more than half. This reduction of absorption by hapten-specific IgA suggests that oral vaccines against carcinogens and toxicants could be developed for humans.
Rapid and Sensitive Protein Similarity Searches
Authors: Lipman David J. , Pearson William R.
Published: October 2006

An algorithm was developed which facilitates the search for similarities between newly determined amino acid sequences and sequences already available in databases. Because of the algorithm's efficiency on many microcomputers, sensitive protein database searches may now become a routine procedure for molecular biologists. The method efficiently identifies regions of similar sequence and then scores the aligned identical and differing residues in those regions by means of an amino acid replaceability matrix. This matrix increases sensitivity by giving high scores to those amino acid replacements which occur frequently in evolution. The algorithm has been implemented in a computer program designed to search protein databases very rapidly. For example, comparison of a 200-amino-acid sequence to the 500,000 residues in the National Biomedical Research Foundation library would take less than 2 minutes on a minicomputer, and less than 10 minutes on a microcomputer (IBM PC).
Huntington's Disease: Two Families with Differing Clinical Features Show Linkage to the G8 Probe
Authors: Folstein Susan E. , Phillips John A. , Meyers Deborah A. , Chase Gary A. , Abbott Margaret H. , Franz Mary L. , Waber Pamela G. , Kazazian Hag H. , Conneally P. Michael , Hobbs Wendy , Tanzi Rudolph , Faryniarz Ann , Gibbons Kerrin , Gusella James
Published: October 2006

To test the hypothesis that interfamily variability in Huntington's Disease (HD) is due to mutation at different loci, linkage analysis was undertaken in two large HD kindreds that differed in ethnicity, age-at-onset, and neurologic and psychiatric features. Both families showed linkage of the HD locus to the G8 probe. Several recombinants were documented in each family, and the best estimate of the recombination fraction for the two families was 6 percent with a 95 percent confidence interval of 0 to 12 percent. Although the data support the existence of a single HD locus, use of the G8 probe for presymptomatic testing in these kindreds would have resulted in a 12 percent error rate in genotype assignment at the HD locus.
A Strong Influence of Serotonin Axons on β-Adrenergic Receptors in Rat Brain
Authors: Stockmeier Craig A. , Martino Andrea M. , Kellar Kenneth J.
Published: October 2006

The role of serotonin axons in modulating the norepinephrine neurotransmission system in rat brain was investigated. Selective lesions of the forebrain serotonergic system were made by injecting 5,7-dihydroxytryptamine into the midbrain raphe nuclei. Four to six weeks after the lesion, the uptake of
3
H-labeled serotonin in the frontal cortex and the hippocampus was reduced by more than 90 percent, while neither the uptake of
3
H-labeled norepinephrine nor the content of norepinephrine was affected in either tissue. The number of β-adrenergic receptors, as measured by radioligand binding with
3
H-labeled dihydroalprenolol, was increased in the frontal cortex and hippocampus of rats with lesions. Similarly, specific lesions of central serotonin axons produced by systemically administered p-chloramphetamine resulted in an increase in the binding of
3
H-labeled dihydroalprenolol to β-adrenergic receptors and in the production of adenosine 3′,5′-monophosphate in response to isoproterenol. These results indicate that serotonin axons may regulate β-adrenergic receptor number and function in brain.
Structure of the Human Interleukin-2 Receptor Gene
Authors: Leonard Warren J. , Depper Joel M. , Kanehisa Minoru , Krönke Martin , Peffer Nancy J. , Svetlik Penny B. , Sullivan Margery , Greene Warner C.
Published: October 2006

The gene encoding the human interleukin-2 (IL-2) receptor consists of 8 exons spanning more than 25 kilobases on chromosome 10. Exons 2 and 4 were derived from a gene duplication event and unexpectedly also are homologous to the recognition domain of human complement factor B. Alternative messenger RNA (mRNA) splicing may delete exon 4 sequences, resulting in a mRNA that does not encode a functional IL-2 receptor. Leukemic T cells infected with HTLV-I and normal activated T cells express IL-2 receptors with identical deduced protein sequences. Receptor gene transcription is initiated at two principal sites in normal activated T cells. Adult T cell leukemia cells infected with HTLV-I show activity at both of these sites, but also at a third transcription initiation site.
Genetic Variation in HTLV-III/LAV Over Time in Patients with AIDS or at Risk for AIDS
Authors: Hahn Beatrice H. , Shaw George M. , Taylor Maria E. , Redfield Robert R. , Markham Phil D. , Salahuddin S. Z. , Wong-Staal Flossie , Gallo Robert C. , Parks Elizabeth S. , Parks Wade P.
Published: October 2006

In a study of genetic variation in the AIDS virus, HTLV-III/LAV, sequential virus isolates from persistently infected individuals were examined by Southern blot genomic analysis, molecular cloning, and nucleotide sequencing. Four to six virus isolates were obtained from each of three individuals over a 1-year or 2-year period. Changes were detected throughout the viral genomes and consisted of isolated and clustered nucleotide point mutations as well as short deletions or insertions. Results from genomic restriction mapping and nucleotide sequence comparisons indicated that viruses isolated sequentially had evolved in parallel from a common progenitor virus. The rate of evolution of HTLV-III/LAV was estimated to be at least 10
-3
nucleotide substitutions per site per year for the
env
gene and 10
-4
for the
gag
gene, values a millionfold greater than for most DNA genomes. Despite this relatively rapid rate of sequence divergence, virus isolates from any one patient were all much more related to each other than to viruses from other individuals. In view of the substantial heterogeneity among most independent HTLV-III/LAV isolates, the repeated isolation from a given individual of only highly related viruses raises the possibility that some type of interference mechanism may prevent simultaneous infection by more than one major genotypic form of the virus.
Studies of the Human c-
myb
Gene and Its Product in Human Acute Leukemias
Authors: Slamon Dennis J. , Boone Thomas C. , Murdock Douglas C. , Keith Duane E. , Press Michael F. , Larson Richard A. , Souza Lawrence M.
Published: October 2006

The
myb
gene is the transforming oncogene of the avian myeloblastosis virus (AMV); its normal cellular homolog, c-
myb
, is conserved across a broad span of evolution. In humans, c-
myb
is expressed in malignant hematopoietic cell lines and in primary hematopoietic tumors. Partial complementary DNA clones were generated from blast cells of patients with acute myelogenous leukemia. The sequences of the clones were compared to the c-
myb
of other species, as well as the v-
myb
of AMV. In addition, the carboxyl terminal region of human c-
myb
was placed in an expression vector to obtain protein for the generation of antiserum, which was used to identify the human c-
myb
gene product. Like v-
myb
, this protein was found within the nucleus of leukemic cells where it was associated with the nuclear matrix. These studies provide further evidence that c-
myb
might be involved in human leukemia.
Gene Amplification of c-
myc
and N-
myc
in Small Cell Carcinoma of the Lung
Authors: Wong Albert J. , Ruppert John M. , Eggleston Joseph , Hamilton Stanley R. , Baylin Stephen B. , Vogelstein Bert
Published: October 2006

The relationship of the copy numbers of the c-
myc
and N-
myc
oncogenes to tumor formation and progression was studied in small cell carcinoma of the lung. When 96 neoplastic lesions from 45 patients were examined, these lesions could be grouped into three categories: high copy (tumors with greater than 3 copies of the N-
myc
or c-
myc
gene per haploid genome), middle copy (1.5 to 3 copies per genome), and normal copy. Fourteen of the patients had middle copy tumors, but this was almost always a result of chromosome duplication rather than the amplification of a small genetic locus. In contrast, five patients had high copy tumors, with the increased copy number in each case due to gene amplification. The amplification did not occur in a heterogeneous fashion within individual patients, since all metastatic lesions from patients with high copy lung tumors were also high copy, while none of 41 metastatic lesions from the other patients were high copy. These data suggest that gene amplification is an important step in neoplastic growth in a subset of patients with small cell carcinoma of the lung and that this genetic event occurs relatively early (before metastasis) in this subset.
Pain Relief by Electrical Stimulation of the Central Gray Matter in Humans and Its Reversal by Naloxone
Authors: Hosobuchi Yoshio , Adams John E. , Linchitz Rita
Published: October 2006

Relief of intractable pain was produced in six human patients by stimulation of electrodes permanently implanted in the periventricular and periaqueductal gray matter. The level of stimulation sufficient to induce pain relief seems not to alter the acute pain threshold. Indiscriminate repetitive stimulation produced tolerance to both stimulation-produced pain relief and the analgesic action of narcotic medication; this process could be reversed by abstinence from stimulation. Stimulation-produced relief of pain was reversed by naloxone in five out of six patients. These results suggest that satisfactory alleviation of persistent pain in humans may be obtained by electronic stimulation.
In Vivo Half-Life of a Protein Is a Function of Its Amino-Terminal Residue
Authors: Bachmair Andreas , Finley Daniel , Varshavsky Alexander
Published: October 2006

When a chimeric gene encoding a ubiquitin-β-galactosidase fusion protein is expressed in the yeast
Saccharomyces cerevisiae
, ubiquitin is cleaved off the nascent fusion protein, yielding a deubiquitinated β-galactosidase (βgal). With one exception, this cleavage takes place regardless of the nature of the amino acid residue of βgal at the ubiquitin-βgal junction, thereby making it possible to expose different residues at the amino-termini of the otherwise identical βgal proteins. The βgal proteins thus designed have strikingly different half-lives in vivo, from more than 20 hours to less than 3 minutes, depending on the nature of the amino acid at the amino-terminus of βgal. The set of individual amino acids can thus be ordered with respect to the half-lives that they confer on βgal when present at its amino-terminus (the "N-end rule"). The currently known amino-terminal residues in long-lived, noncompartmentalized intracellular proteins from both prokaryotes and eukaryotes belong exclusively to the stabilizing class as predicted by the N-end rule. The function of the previously described posttranslational addition of single amino acids to protein amino-termini may also be accounted for by the N-end rule. Thus the recognition of an amino-terminal residue in a protein may mediate both the metabolic stability of the protein and the potential for regulation of its stability.
Tissue-Specific and Ectopic Expression of Genes Introduced into Transgenic Mice by Retroviruses
Authors: Soriano Philippe , Cone Roger D. , Mulligan Richard C. , Jaenisch Rudolf
Published: October 2006

Recombinant retroviruses containing the complete genomic human β globin gene (under the control of its own promoter) and the bacterial neomycin phosphotransferase gene (under the control of the normal or enhancerless viral promoter) were used to derive transgenic mouse strains by infection of preimplantation embryos. Expression of the β globin gene in hematopoietic tissues was observed in all transgenic strains. In addition, one strain showed ectopic expression of β globin in the same tissues that also expressed high levels of RNA from the viral promoter. It is likely that expression from the long terminal repeat (LTR), in contrast to expression from the internal promoter, is dependent on the site of integration. Thus, retroviral vectors can be used for tissue-specific expression of foreign genes in transgenic mice, as well as for the identification of loci that allow developmental activation of a provirus.
The Biology and Chemistry of Fertilization
Authors: Wassarman Paul M.
Published: October 2006

Fertilization of eggs by sperm, the means by which sexual reproduction takes place in nearly all multicellular organisms, is fundamental to the maintenance of life. In both mammals and nonmammals, the pathway that leads to fusion of an egg with a single sperm consists of many steps that occur in a compulsory order. These steps include species-specific cellular recognition, intracellular and intercellular membrane fusions, and enzyme-catalyzed modifications of cellular investments. In several instances, the molecular mechanisms that underlie these events during mammalian fertilization are beginning to be revealed.
HTLV
x
Gene Mutants Exhibit Novel Transcriptional Regulatory Phenotypes
Authors: Wachsman William , Cann Alan J. , Williams Janice L. , Slamon Dennis J. , Souza Larry , Shah Neil P. , Chen Irvin S. Y.
Published: October 2006

The human T-cell leukemia viruses, HTLV-I and HTLV-II, contain a gene, termed
x
, with transcriptional regulatory function. The properties of the
x
proteins were analyzed by constructing mutant genes containing site-directed deletions and point mutations. The results demonstrate that the amino terminal 17 amino acids of the
x
protein constitute part of a functional domain that is critical for the transcriptional activating properties of the protein. Within this region, substitution of a leucine residue for a proline residue results in major changes in the
trans
-activation phenotype of the protein. The mutant HTLV-II
x
protein, though incapable of activating the HTLV-II long terminal repeat, will block
trans
-activation of the HTLV-II long terminal repeat by the wild-type protein. The altered phenotype of this mutant suggests a potential negative regulatory function of the
x
protein.
Zinc-Dependent Structure of a Single-Finger Domain of Yeast ADR1
Authors: Párraga Grace , Horvath Suzanna J. , Eisen Amur , Taylor Wayne E. , Hood Leroy , Young Elton T. , Klevit Rachel E.
Published: October 2006

In the proposed "zinc finger" DNA-binding motif, each repeat unit binds a zinc metal ion through invariant Cys and His residues and this drives the folding of each 30-residue unit into an independent nucleic acid-binding domain. To obtain structural information, we synthesized single and double zinc finger peptides from the yeast transcription activator ADR1, and assessed the metal-binding and DNA-binding properties of these peptides, as well as the solution structure of the metal-stabilized domains, with the use of a variety of spectroscopic techniques. A single zinc finger can exist as an independent structure sufficient for zinc-dependent DNA binding. An experimentally determined model of the single finger is proposed that is consistent with circular dichroism, one- and two-dimensional nuclear magnetic resonance, and visual spectroscopy of the single-finger peptide reconstituted in the presence of zinc.
Scanning Tunneling Microscopy and Atomic Force Microscopy: Application to Biology and Technology
Authors: Hansma P. K. , Elings V. B. , Marti O. , Bracker C. E.
Published: October 2006

The scanning tunneling microscope (STM) and the atomic force microscope (AFM) are scanning probe microscopes capable of resolving surface detail down to the atomic level. The potential of these microscopes for revealing subtle details of structure is illustrated by atomic resolution images including graphite, an organic conductor, an insulating layered compound, and individual adsorbed oxygen atoms on a semiconductor. Application of the STM for imaging biological materials directly has been hampered by the poor electron conductivity of most biological samples. The use of thin conductive metal coatings and replicas has made it possible to image some biological samples, as indicated by recently obtained images of a recA-DNA complex, a phospholipid bilayer, and an enzyme crystal. The potential of the AFM, which does not require a conductive sample, is shown with molecular resolution images of a nonconducting organic monolayer and an amino acid crystal that reveals individual methyl groups on the ends of the amino acids. Applications of these new microscopes to technology are demonstrated with images of an optical disk stamper, a diffraction grating, a thin-film magnetic recording head, and a diamond cutting tool. The STM has even been used to improve the quality of diffraction gratings and magnetic recording heads.
Rates of DNA Sequence Evolution Differ Between Taxonomic Groups
Authors: Britten Roy J.
Published: October 2006

The mutation rates of DNA sequences during evolution can be estimated from interspecies DNA sequence differences by assaying changes that have little or no effect on the phenotype (neutral mutations). Examination of available measurements shows that rates of DNA change of different phylogenetic groups differ by a factor of 5. The slowest rates are observed for higher primates and some bird lineages, while faster rates are seen in rodents, sea urchins, and drosophila. The rate of DNA sequence change has decreased markedly during primate evolution. The contrast in rates of DNA sequence change is probably due to evolutionary variation and selection of biochemical mechanisms such as DNA replication or repair.
Inhibition of Development of Exoerythrocytic Forms of Malaria Parasites by γ-Interferon
Authors: Ferreira Arturo , Schofield Louis , Enea Vincenzo , Schellekens Huub , van der Meide Peter , Collins William E. , Nussenzweig Ruth S. , Nussenzweig Victor
Published: October 2006

A specific DNA probe was used to study the effect of recombinant rat, mouse, and human γ-interferon (γ-IFN) on the course of sporozoite-induced malaria infections. In mice and rats infected with sporozoites of
Plasmodium berghei
, mouse and rat γ-IFN's strongly inhibited the development of the exoerythrocytic forms in the liver cells of the hosts, but not the development of the erythrocytic stages. The degree of inhibition of the exoerythrocytic forms was proportional to the dose of γ-IFN administered, but was independent of the number of sporozoites used for challenge. A 30 percent reduction in the development of exoerythrocytic forms in rat liver was achieved when 150 units (about 15 nanograms of protein) of rat γ-IFN were injected a few hours before sporozoite challenge; the reduction was 90 percent or more with higher doses of γ-IFN. The effect was less pronounced if the γ-IFN was administered 18 hours before or a few hours after challenge. Human γ-IFN also diminished the parasitemia in chimpanzees infected with sporozoites of the human malaria parasite
Plasmodium vivax
. The target of γ-IFN activity may be the infected hepatocytes themselves, as shown by in vitro experiments in which small doses of the human lymphokine inhibited the development of exoerythrocytic forms of
Plasmodium berghei
in a human hepatoma cell line. These results suggest that immunologically induced interferon may be involved in controlling malaria infection under natural conditions.
Induction of Membrane Ruffling and Fluid-Phase Pinocytosis in Quiescent Fibroblasts by
ras
Proteins
Authors: Bar-Sagi Dafna , Feramisco James R.
Published: October 2006

Expression of the
ras
oncogene is thought to be one of the contributing events in the initiation of certain types of human cancer. To determine the cellular activities that are directly triggered by
ras
proteins, the early consequences of microinjection of the human H-
ras
proteins into quiescent rat embryo fibroblasts were investigated. Within 30 minutes to 1 hour after injection, cells show a marked increase in surface ruffles and fluid-phase pinocytosis. The rapid enhancement of membrane ruffling and pinocytosis is induced by both the proto-oncogenic and the oncogenic forms of the H-
ras
protein. The effects produced by the oncogenic protein persist for more than 15 hours after injection, whereas the effects of the proto-oncogenic protein are short-lived, being restricted to a 3-hour interval after injection. The stimulatory effect of the
ras
oncogene protein on ruffling and pinocytosis is dependent on the amount of injected protein and is accompanied by an apparent stimulation of phospholipase A
2
activity. These rapid changes in cell membrane activities induced by
ras
proteins may represent primary events in the mechanism of action of
ras
proteins.
Cytochrome P-450—Catalyzed Formation of Δ4-VPA, a Toxic Metabolite of Valproic Acid
Authors: Rettie Allan E. , Rettenmeier Albert W. , Howald William N. , Baillie Thomas A.
Published: October 2006

Liver damage induced by the antiepileptic drug valproic acid (VPA) is believed to be mediated by an unsaturated metabolite of the drug, Δ4-VPA. In studies of the biological origin of this hepatotoxic compound, it was found that liver microsomes from phenobarbital-treated rats catalyzed the desaturation of VPA to Δ4-VPA. Indirect evidence suggested that cytochrome P-450 was the responsible enzyme, a conclusion that was verified by studies with a purified and reconstituted form of the hemoprotein, which catalyzed the oxidation of VPA to 4- and 5-hydroxyvalproic acid and to Δ4-VPA. Desaturation of a nonactivated alkyl substituent represents a novel metabolic function of cytochrome P-450 and probably proceeds via the conversion of substrate to a transient free radical intermediate, which partitions between recombination (alcohol formation) and elimination (olefin production) pathways. These findings have broad implications with respect to the metabolic generation of olefins and may explain the increased hepatotoxic potential of VPA when it is administered in combination with potent enzyme-inducing anticonvulsants such as phenobarbital.
Rheumatoid Factor Secretion from Human Leu-1
+
B Cells
Authors: Hardy Richard R. , Hayakawa Kyoko , Shimizu Masatoshi , Yamasaki Katsuhiko , Kishimoto Tadamitsu
Published: October 2006

A human B cell subpopulation identifiable by the expression of the cell surface antigen Leu-1 (CD5) is responsible for most of the immunoglobulin M rheumatoid factor secreted in vitro after the cells are stimulated with
Staphylococcus aureus
. The ability of B cells bearing the Leu-1 marker (Leu-1
+
) to secrete rheumatoid factor is present early in development and extends to adulthood, since Leu-1
+
B cells from cord blood and from peripheral blood lymphocytes of both normal adults and patients with certain autoimmune conditions secrete rheumatoid factor in comparable amounts. The neonatal enrichment of Leu-1
+
B cells, the presence of Leu-1
+
B cells in increased frequencies in patients with autoimmune disease, and the involvement of Leu-1
+
B cells in autoantibody secretion suggest both developmental and functional homologies between this human B cell subpopulation and the murine Ly-1 B cell subpopulation.
Insulin Rapidly Increases Diacylglycerol by Activating De Novo Phosphatidic Acid Synthesis
Authors: Farese Robert V. , Konda Thomas Suman , Davis John S. , Standaert Mary L. , Pollet Robert J. , Cooper Denise R.
Published: October 2006

The mechanisms whereby insulin increases diacylglycerol in BC3H-1 myocytes were examined. When [
3
H]arachidonate labeling of phospholipids was used as an indicator of phospholipase C activation, transient increases in [
3
H]diacylglycerol were observed between 0.5 and 10 minutes after the onset of insulin treatment. With [
3
H]glycerol labeling as an indicator of de novo phospholipid synthesis, [
3
H]diacylglycerol was increased maximally at 1 minute and remained elevated for 20 minutes. [
3
H]Glycerol-labeled diacylglycerol was largely derived directly from phosphatidic acid. Insulin increased de novo phosphatidic acid synthesis within 5 to 10 seconds; within 1 minute, this synthesis was 60 times greater than that of controls. Thus, the initial increase in diacylglycerol is due to both increased hydrolysis of phospholipids and a burst of de novo phosphatidic acid synthesis. After 5 to 10 minutes, de novo phosphatidic acid synthesis continues as a major source of diacylglycerol. Both phospholipid effects of insulin seem important for generating diacylglycerol and other phospholipid-derived intracellular signaling substances.
Corkscrew-Like Shortening in Single Smooth Muscle Cells
Authors: Warshaw David M. , McBride Whitney J. , Work Steven S.
Published: October 2006

The slower and more economical contraction of smooth muscle as compared to that of skeletal muscle may relate to the arrangement of its contractile apparatus. Because the arrangement of the contractile apparatus determines the manner in which a single smooth muscle cell shortens, shortening of a contracting cell was examined by tracking of marker bead movements on the cell surface by means of digital video microscopy. Smooth muscle cells were observed to freely shorten in a unique corkscrew-like fashion with a pitch of 1.4 cell lengths (that is, the length change required for one complete rotation of cell) at a rate of 27 degrees per second. Corkscrew-like shortening was interpreted in terms of a structural model in which the contractile apparatus or cytoskeleton (or both) are helically oriented within the cell. Such an arrangement of these cytoarchitectural elements may help to explain in part the contractile capabilities of smooth muscle.
Thyroid Hormone Regulates TRH Biosynthesis in the Paraventricular Nucleus of the Rat Hypothalamus
Authors: Segerson Thomas P. , Kauer John , Wolfe Hubert C. , Mobtaker Hamid , Wu Ping , Jackson Ivor M. D. , Lechan Ronald M.
Published: October 2006

Thyroid hormone is important in the regulation of synthesis and secretion of thyroid-stimulating hormone (TSH) in the anterior pituitary, but its role in the control of hypothalamic thyrotropin-releasing hormone (TRH) is controversial. To determine whether thyroid hormone regulates the function of TRH in the hypothalamic tuberoinfundibular system, a study was made of the effect of hypothyroidism on thyrotropin-releasing hormone messenger RNA (proTRH mRNA) and TRH prohormone in the rat paraventricular nucleus. Extracts of rat hypothalamic paraventricular nucleus were examined by quantitative Northern blot analysis, and coronal sections of rat brain were examined by in situ hybridization histochemistry and immunocytochemistry. A nearly twofold increase in proTRH mRNA was observed in hypothyroid animals; this increase could be obliterated by levothyroxine treatment, suggesting an inverse relation between circulating thyroid hormone and proTRH mRNA. In situ hybridization showed that this response occurred exclusively in medial parvocellular neurons of the paraventricular nucleus. A simultaneous increase in proTRH mRNA and immunoreactive TRH prohormone in this region suggests that hypothyroidism induces both transcription and translation of the TRH prohormone in the paraventricular nucleus.
Interleukin-2: Inception, Impact, and Implications
Authors: Smith Kendall A.
Published: October 2006

Interleukin-2 (IL-2), the first of a series of lymphocytotrophic hormones to be recognized and completely characterized, is pivotal for the generation and regulation of the immune response. A T lymphocyte product, IL-2 also stimulates T cells to undergo cell cycle progression via a finite number of interactions with its specific membrane receptors. Because T cell clonal proliferation after antigen challenge is obligatory for immune responsiveness and immune memory, the IL-2-T cell system has opened the way to a molecular understanding of phenomena that are fundamental to biology, immunology, and medicine.
Isolation of T-Cell Tropic HTLV-III-Like Retrovirus from Macaques
Authors: Daniel M. D. , Letvin N. L. , King N. W. , Kannagi M. , Sehgal P. K. , Hunt R. D. , Kanki P. J. , Essex M. , Desrosiers R. C.
Published: October 2006

The isolation of a T-cell tropic retrovirus from three immunodeficient macaques and one macaque with lymphoma is described. The morphology, growth characteristics, and antigenic properties of this virus indicate that it is related to the causative agent of acquired immune deficiency syndrome in humans (HTLV-III or LAV). This virus is referred to as simian T-lymphotropic virus type III (STLV-III) of macaques. The existence of a cytopathic, T-cell tropic virus resembling HTLV-III in monkeys may facilitate study of disease induction and vaccine development in an animal model.
A Bitter Substance Induces a Rise in Intracellular Calcium in a Subpopulation of Rat Taste Cells
Authors: Akabas Myles H. , Dodd Jane , Al-Awqati Qais
Published: October 2006

The sense of taste permits animals to discriminate between foods that are safe and those that are toxic. Because most poisonous plant alkaloids are intensely bitter, bitter taste warns animals of potentially hazardous foods. To investigate the mechanism of bitter taste transduction, a preparation of dissociated rat taste cells was developed that can be studied with techniques designed for single-cell measurements. Denatonium, a very bitter substance, caused a rise in the intracellular calcium concentration due to release from internal stores in a small subpopulation of taste cells. Thus, the transduction of bitter taste may occur via a receptor-second messenger mechanism leading to neurotransmitter release and may not involve depolarization-mediated calcium entry.
Molecular Cloning of Two Types of GAP Complementary DNA from Human Placenta
Authors: Trahey Meg , Wong Gail , Halenbeck Robert , Rubinfeld Bonnee , Martin George A. , Ladner Martha , Long Christopher M. , Crosier Walter J. , Watt Ken , Koths Kirston , McCormick Frank
Published: October 2006

The
ras
p21 GTPase-activating protein (GAP) was purified from human placental tissue. Internal amino acid sequence was obtained from this 120,000-dalton protein and, by means of this sequence, two types of complementary DNA clones were isolated and characterized. One type encoded GAP with a predicted molecular mass of 116,000 daltons and 96% identity with bovine GAP. The messenger RNA of this GAP was detected in human lung, brain, liver, leukocytes, and placenta. The second type appeared to be generated by a differential splicing mechanism and encoded a novel form of GAP with a predicted molecular mass of 100,400 daltons. This protein lacks the hydrophobic amino terminus characteristic of the larger species, but retains GAP activity. The messenger RNA of this type was abundantly expressed in placenta and in several human cell lines, but not in adult tissues.
Nonoxidative Glucose Consumption During Focal Physiologic Neural Activity
Authors: Fox Peter T. , Raichle Marcus E. , Mintun Mark A. , Dence Carmen
Published: October 2006

Brain glucose uptake, oxygen metabolism, and blood flow in humans were measured with positron emission tomography, and a resting-state molar ratio of oxygen to glucose consumption of 4.1:1 was obtained. Physiological neural activity, however, increased glucose uptake and blood flow much more (51 and 50 percent, respectively) than oxygen consumption (5 percent) and produced a molar ratio for the increases of 0.4:1. Transient increases in neural activity cause a tissue uptake of glucose in excess of that consumed by oxidative metabolism, acutely consume much less energy than previously believed, and regulate local blood flow for purposes other than oxidative metabolism.
Nef
Protein of HIV-1 Is a Transcriptional Repressor of HIV-1 LTR
Authors: Ahmad Nafees , Venkatesan Sundararajan
Published: October 2006

In studies of the genetics of human immunodeficiency virus type 1 (HIV-1), the product of the
nef
gene, formerly known as F, 3′-
orf
, or B-ORF, was a negative regulator of HIV-1 replication. Proviruses with mutations in the
nef
gene replicated better than their standard counterparts during transient expression, and the mutant virus maintained its enhanced replication even after serial passages in T lymphocytes. The
nef
protein trans-suppressed, in a dose-dependent manner, the replication of wild-type and
nef
mutant proviruses and the expression of reporter genes linked to the HIV-1 long terminal repeat (LTR). The repression induced by the
nef
protein was mediated by inhibition of transcription from the HIV-1 LTR, which contains a far upstream cis element (previously recognized to be a negative regulatory element) between 340 and 156 nucleotides upstream of the RNA initiation site.