bluuebunny/crossref_metadata_2025 huggingface dataset:
Enhancer-dependent expression of human kappa immunoglobulin genes introduced into mouse pre-B lymphocytes by electroporation.
Authors: Potter H , Weir L , Leder P
Published: May 2006

We have developed a general method for introducing cloned genes into mammalian cells that affords substantial benefits over current technology. It is simple, rapid, and applicable to many (perhaps all) cell types, including those that are refractory to traditional transfection procedures. The method involves exposure of a suspension of cells and cloned DNA to a high-voltage electric discharge. In a model application of this transfection procedure, we have studied the expression of cloned human and mouse Ig kappa genes stably introduced into mouse pre-B cells and fibroblasts. We find that there is a B-cell-specific enhancer-activator region in the J-C intron of the human kappa gene that is necessary for efficient transcription of the cloned gene in mouse pre-B lymphocytes. This suggests that both the DNA element and the proteins required for its regulatory activity have been highly conserved in evolution and that these elements operate at the pre-B-cell stage of immunocyte development, a stage that precedes productive kappa gene rearrangement.
Monoclonal antibodies and peptide mapping reveal structural similarities between the subunits of the glycine receptor of rat spinal cord.
Authors: Pfeiffer F , Simler R , Grenningloh G , Betz H
Published: May 2006

The glycine receptor of rat spinal cord is an oligomeric membrane glycoprotein of molecular mass 250,000 daltons that contains three polypeptides of 48,000, 58,000, and 93,000 daltons. Monoclonal antibodies (mAbs) were prepared against the affinity-purified glycine receptor protein by using 125I-labeled receptor preparations for the detection of positive hybrids. From nine monoclonal antibodies obtained, six recognized denatured receptor polypeptides blotted to nitrocellulose paper. Two of these antibodies bound to more than one glycine receptor polypeptide: mAb GlyR 4a stained the 48,000- and 58,000-dalton polypeptides, and mAb GlyR 7a stained the 48,000- and 93,000-dalton polypeptides. Common antigenic determinants thus are shared by the different subunits of the glycine receptor. Complementary results were obtained by peptide mapping of 125I-labeled glycine receptor polypeptides with various proteases. A set of peptide fragments of the same apparent molecular mass was produced from the different glycine receptor polypeptides by using V8 protease, chymotrypsin, and elastase. These data suggest that the subunits of the glycine receptor have significant homology within their primary structure and may have evolved from a common ancestor receptor polypeptide.
Regional distribution of putative vasopressin receptors in rat brain and pituitary by quantitative autoradiography.
Authors: Brinton R E , Gee K W , Wamsley J K , Davis T P , Yamamura H I
Published: May 2006

Quantitative light microscopic autoradiography was used to map and characterize the distribution of [3H]arginine vasopressin [( 3H]AVP) binding sites in the rat brain. HPLC analysis for possible degradation of AVP during binding indicated that addition of specific peptidase inhibitors prevented metabolism of AVP. Binding sites for [3H]AVP were observed in the hypothalamus and pituitary as well as in brain regions where AVP may act as a neuroregulator. Within the hypothalamus, dense AVP binding sites were seen in the suprachiasmatic, supraoptic, and paraventricular nuclei. High specific binding was also apparent in the median eminence tubero-infundibular region and in the posterior lobe of the pituitary. [3H]AVP labeling at possible neuroregulatory sites was observed in the hippocampus, lateral septum, superficial cortex, cerebellum, nucleus tractus solitarious, adenohypophysis, and spinal cord.
Opioid receptor reserve in normal and morphine-tolerant guinea pig ileum myenteric plexus.
Authors: Chavkin C , Goldstein A
Published: May 2006

We have measured the opioid receptor reserve in the guinea pig ileum myenteric plexus by means of the site-directed alkylating agent, beta-chlornaltrexamine. Treatment of the tissue with low (less than 10 nM) concentrations of beta-chlornaltrexamine caused a parallel shift of the log concentration-response curves for both normorphine and dynorphin A-(1-13). Analysis of the resulting curves indicated that the Kd values were 1.5 +/- 0.5 X 10(-6) and 10 +/- 4 X 10(-9), respectively. Using the naloxone Ke to distinguish between the mu and kappa receptors in this tissue, we found that the receptor selectivities of normorphine and dynorphin A-(1-13) were unchanged after a maximum parallel shift, thus demonstrating that there are both spare mu and spare kappa receptors present. The spare-receptor fraction for both receptor types was about 90%. In morphine-tolerant preparations (chronic pellet implantation), there was an apparent reduction in the fraction of spare mu receptors without any change in the apparent affinity of normorphine. Reduction in the spare receptor fraction does not necessarily imply reduction in the number of binding sites. We suggest that this reduction in receptor reserve is the basis of opioid tolerance, since the agonist concentration needed to produce a given effect is expected to increase as the receptor reserve decreases.
Characterization of a human liver cytochrome P-450 involved in the oxidation of debrisoquine and other drugs by using antibodies raised to the analogous rat enzyme.
Authors: Distlerath L M , Guengerich F P
Published: May 2006

Debrisoquine 4-hydroxylase activity is a prototype for genetic polymorphism in oxidative drug metabolism in humans; approximately 10% of Caucasian populations exhibit the poor metabolizer phenotype, and the clearance of at least 14 other drugs has been shown to be deficient in patients exhibiting this phenotype. Antibodies prepared to a cytochrome P-450 shown to be responsible for debrisoquine 4-hydroxylation in rats were found to inhibit the oxidation of debrisoquine and sparteine, encainide, and propranolol, three other drugs suggested to be associated with this phenotype, in human liver microsomes. The antibodies did not inhibit the oxidation of seven other cytochrome P-450 substrates. The antibodies recognized a single polypeptide of Mr51,000 after combined sodium dodecyl sulfate/polyacrylamide electrophoresis and immunochemical staining of human liver microsomes. The intensity of this band was significantly correlated with debrisoquine 4-hydroxylase activity when liver microsomes from 44 organ donors were examined. Immunoprecipitation of in vitro translation products of total liver RNA revealed major electrophoretic bands corresponding to the cytochrome P-450 in rats and humans. The level of translatable mRNA coding for the debrisoquine-hydroxylating cytochrome P-450 was an order of magnitude less in human liver than in rat liver. The availability of these antibodies provides a biochemical basis for further basic and clinical studies on the role of a particular cytochrome P-450 polymorphism in humans.
Molecular cloning and nucleotide sequence of cDNA for rat ornithine carbamoyltransferase precursor.
Authors: Takiguchi M , Miura S , Mori M , Tatibana M , Nagata S , Kaziro Y
Published: May 2006

Messenger RNA of rat ornithine carbamoyltransferase (EC 2.1.3.3), a mitochondrial matrix enzyme, was enriched by immunoprecipitation of rat liver free polysomes, and recombinant plasmids were prepared from the enriched mRNA by a vector-primer method. The cDNA clones for ornithine carbamoyltransferase were identified by hybrid-arrested translation and hybrid-selected translation. One of the clones, designated pOTC-1, contained a 1.6-kilobase insert and hybridized to a mRNA of approximately equal to 1.8 kilobases in rat liver. The cDNA clone was subjected to nucleotide sequence analysis. The deduced amino acid sequence indicates that the ornithine carbamoyltransferase precursor consists of the mature enzyme of 322 amino acid residues and an NH2-terminal peptide extension (presequence) of 32 amino acid residues. The presequence contains 8 basic amino acid residues, no acidic residues, and no hydrophobic amino acid stretch. The amino acid sequence of the rat ornithine carbamoyltransferase was compared with the recently reported sequence of the human enzyme [Horwich, A. L., Fenton, W. A., Williams, K. R., Kalousek, F., Kraus, J. P., Doolittle, R. F., Konigsberg, W. & Rosenberg, L. E. (1984) Science 224, 1068-1074]. The sequences of the mature enzyme portion are 93% identical, whereas those of the presequences are 69% identical. There are two highly conserved segments in the presequences of the rat and human enzymes. One of the two conserved segments is significantly similar to a segment of the presequence of yeast mitochondrial elongation factor EF-Tu. These results suggest that the homologous segments are important for the proteins that are synthesized in the cytosol to be transported into the mitochondrial matrix.
Identification of a retrovirus-like repetitive element in human DNA.
Authors: Mager D L , Henthorn P S
Published: May 2006

We describe a 5- to 6-kilobase-pair repetitive family in human DNA. One member of this family is linked to the beta-globin gene cluster and is close to the 3' breakpoints of three different naturally occurring deletions involving this gene cluster. Sequence analysis indicates that this element includes terminal direct repeats of 415 base pairs that exhibit the features of long terminal repeats (LTRs) of retroviruses. A potential histidine tRNA primer binding site occurs just 3' to the 5' direct repeat. This retrovirus-like element interrupts a member of the Kpn I family of repeated DNA and is bracketed by a 5-base-pair directly repeated sequence. When attempts are made to clone the element in bacteriophage, homologous recombination between the LTR-like sequences is very frequently observed. Copy number estimates by two methods indicate that the element is repeated 800-1000 times in the human genome. We term this Homo sapiens family of retrovirus-like elements having a histidine tRNA primer binding site the hsRTVL-H family.
Circumsporozoite gene of plasmodium cynomolgi (Gombak):cDNA cloning and expression of the repetitive circumsporozoite epitope.
Authors: Enea V , Arnot D , Schmidt E C , Cochrane A , Gwadz R , Nussenzweig R S
Published: May 2006

We report the identification, sequence, and expression in Escherichia coli of the immunodominant epitope of the circumsporozoite (CS) gene of Plasmodium cynomolgi (Gombak), a simian malaria parasite. This epitope is encoded by a DNA sequence that is tandemly repeated 10 times in the cDNA clone. Subclones that contain and express only repeats and in variable number have been constructed. We show that the binding of a specific anti-CS protein monoclonal antibody correlates positively with the number of repeats in each subclone. The CS gene of another strain of P. cynomolgi (NIH) encodes an immunodominant epitope that is immunologically distinct from that of the Gombak strain. We present evidence that these two CS genes share extensive overall homology, although the nucleotide sequences that encode the epitopes appear to be unrelated.
Reduction in the incidence of radiation-induced tumors in rats after restriction of food intake.
Authors: Gross L , Dreyfuss Y
Published: May 2006

In our inbred colony of Sprague-Dawley rats, fractionated total-body x-ray irradiation (150 rads five times at weekly intervals; 1 rad = 0.01 gray) increased the incidence of tumors from 22% to 93% in females and from 5% to 59% in males [Gross, L. & Dreyfuss, Y. (1979) Proc. Natl. Acad. Sci. USA 76, 5910-5913]. In experiments reported here, we investigated the influence of reduced food intake on the incidence of radiation-induced tumors in Sprague-Dawley rats. All rats in both groups received fractionated total-body x-ray irradiation similar to that specified above. Among the irradiated rats on full (ad lib) diet (five to six pellets of Purina Rodent Lab Chow per day, each) were 14 females and 9 males, and all (100%) developed tumors (or leukemia) at an average age of 13.7 months in females and 13.4 months in males. Among the litter mates on restricted diet (two pellets of Purina Rodent Lab Chow per day, each), only 9 out of 29 females (31%) developed tumors at an average age of 18.2 months and 1 out of 15 males (6.7%) developed a tumor at 9 months of age. In the full diet group, the majority of tumors developing in females were benign. Among the 9 irradiated males, 1 developed leukemia and the remaining 8 developed tumors; 7 of them were examined, 6 were found malignant, and 1 was benign. In the restricted diet group, of the 9 tumors that developed in females, 4 were malignant and 5 were benign. The tumor that developed in 1 male was a sarcoma.
Defective Ca2+ metabolism in Duchenne muscular dystrophy: effects on cellular and viral growth.
Authors: Fingerman E , Campisi J , Pardee A B
Published: May 2006

Normal fibroblasts in medium containing 0.02 mM CaCl2 arrested growth within 24 hr, whereas Duchenne muscular dystrophy fibroblasts continued to grow for 5 days, albeit at 40% of their rate in standard medium (1.8 mM CaCl2). Moreover, Duchenne cells in calcium-deficient medium showed an enhanced rate of protein synthesis (60% over the rate in standard medium), whereas normal cells were unaffected. Previously we described a general assay for detection of mutant cells by using herpes simplex virus I replication as a probe of cellular function. By altering the growth medium, one can elicit changes in viral DNA replication that depend upon cellular differences. Duchenne fibroblasts in calcium-deficient low-serum (0.5%) medium supported viral replication at a rate 7- to 10-fold greater than did normal cells infected under the same conditions. Using this viral assay, we have successfully identified all 10 samples of a blind coded set of Duchenne muscular dystrophy, normal, and heterozygote cells. In addition, differences of a lower magnitude were found between these cell strains as measured by cellular growth or protein synthesis. Therefore, a cell's ability to grow and support viral replication in calcium-deficient medium can be used to readily distinguish Duchenne muscular dystrophy fibroblasts from normal ones. These results suggest that the viral assay could be used as a prenatal diagnostic test. A defect related to calcium metabolism may be fundamental to this disease.
Serotonin increases intracellular Ca2+ transients in voltage-clamped sensory neurons of Aplysia californica.
Authors: Boyle M B , Klein M , Smith S J , Kandel E R
Published: May 2006

Noxious stimulation of the tail of Aplysia californica produces behavioral sensitization; it enhances several related defensive reflexes. This reflex enhancement involves heterosynaptic facilitation of transmitter release from sensory neurons of the reflex. The facilitation is stimulated by serotonin (5-HT) and involves suppression of a 5-HT-sensitive K+ current (the S current). Suppression of the S current broadens the action potential of the sensory neurons and is thought to enhance transmitter release by prolonging entry of Ca2+ in the presynaptic terminals. We now report a component of enhanced Ca2+ accumulation that is independent of changes in spike shape. We have measured intracellular free Ca2+ transients during long depolarizing steps in voltage-clamped sensory neuron cell bodies injected with the Ca2+-sensitive dye arsenazo III. The free Ca2+ transients elicited by a range of depolarizing voltage-clamp steps increase in amplitude by 75% following application of 5-HT. Since it is observed under voltage-clamp conditions, this increase in the free Ca2+ transients is not merely secondary to the changes in K+ current but must reflect an additional mechanism, an intrinsic change in the handling of Ca2+ by the cell. We have not yet determined whether this change in Ca2+ handling reflects an increase in Ca2+ influx, a reduction in intracellular Ca2+ uptake, or a release of Ca2+ from intracellular stores. Regardless of the underlying mechanism, however, it seems possible that the enhancement of Ca2+ accumulation and the reduction in K+ current act synergistically in producing short-term presynaptic facilitation. Alternatively, this additional modulation of Ca2+ by 5-HT might contribute to processes such as classical conditioning or long-term sensitization that may depend on Ca2+.
The intracellular pathway and assembly of newly formed variable surface glycoprotein of Trypanosoma brucei.
Authors: Grab D J , Webster P , Verjee Y
Published: May 2006

Pulse-chase experiments using L-[35S]methionine suggest that Trypanosoma brucei MITat 1.2 variable surface glycoprotein (VSG) synthesized in the rough endoplasmic reticulum, a process that takes 6-8 min, is shuttled to the Golgi complex 8 min later. Labeling of ultrathin frozen sections with affinity-purified anti-cross-reacting determinant (CRD) IgG followed by protein A-colloidal gold shows that the CRD is localized in the trans-Golgi region. cis-Golgi is not labeled. VSG, when solubilized by treatment with the detergent Nonidet P-40, behaves on sucrose density gradients as a non-membrane protein with a sedimentation value of 5 S. In contrast, VSG solubilized in the presence of Zwittergent TM 3-14 yielded several VSG-containing fractions greater than 5 S, and only the 5S fraction contained the CRD. Lack of the CRD in VSG complexes with sedimentation values greater than 5 S suggests that this determinant is either masked from antibody, perhaps by involvement in polymer formation, or represents the membrane form of VSG recently described by Cardoso de Almeida and Turner [Cardoso de Almeida, M. L. & Turner, M. J. (1983) Nature (London) 302, 349-352].
Calcium/phospholipid-dependent protein kinase (protein kinase C) phosphorylates and activates tyrosine hydroxylase.
Authors: Albert K A , Helmer-Matyjek E , Nairn A C , Müller T H , Haycock J W , Greene L A , Goldstein M , Greengard P
Published: May 2006

Protein kinase C, purified to homogeneity, was found to phosphorylate and activate tyrosine hydroxylase that had been partially purified from pheochromocytoma PC 12 cells. These actions of protein kinase C required the presence of calcium and phospholipid. This phosphorylation of tyrosine hydroxylase reduced the Km for the cofactor 6-methyltetrahydropterine from 0.45 mM to 0.11 mM, increased the Ki for dopamine from 4.2 microM to 47.5 microM, and produced no change in the Km for tyrosine. Little or no change in apparent Vmax was observed. These kinetic changes are similar to those seen upon activation of tyrosine hydroxylase by cAMP-dependent protein kinase. Two-dimensional phosphopeptide maps of tyrosine hydroxylase were identical whether the phosphorylation was catalyzed by protein kinase C or by the catalytic subunit of cAMP-dependent protein kinase. Both protein kinases phosphorylated serine residues. The results suggest that protein kinase C and cAMP-dependent protein kinase phosphorylate the same site(s) on tyrosine hydroxylase and activate tyrosine hydroxylase by the same mechanism.
Toxin-antitoxin selection for isolating somatic cell fusion products between any cell types.
Authors: Wright W E
Published: May 2006

A selective system was developed that permits the isolation of heterokaryons and cell hybrids between any cell types without the use of selectable genetic markers. Antibodies were introduced into two populations of non-transformed normal diploid human fibroblasts by the osmotic lysis of pinosomes containing hypertonic antibody solutions. Heterokaryons formed by fusing antiricin-injected cells to antidiphtheria toxin-injected cells survived an overnight treatment in both ricin and diphtheria toxin. This approach should have general applicability to a variety of investigations such as the complementation analysis of human genetic syndromes, heterokaryon studies of cell differentiation, tumorigenesis and growth control, cell reconstruction experiments, and the production of hybridoma cells.
Polymorphisms of a human variable heavy chain gene show linkage with constant heavy chain genes.
Authors: Johnson M J , Natali A M , Cann H M , Honjo T , Cavalli-Sforza L L
Published: May 2006

In this study, restriction fragment length polymorphisms (RFLPs) were identified with an immunoglobulin variable heavy chain region (Ig VH) probe and the inheritance of the polymorphisms was analyzed in families. Linkage within the VHII gene cluster and between the VHII and Ig CH genes was investigated by lod (logarithm of odds) score analysis. In addition, the position of the VHII genes was determined in relation to another polymorphic locus--D14S1, which is tightly linked and centromeric to the CH genes. Genetic associations between genes in the CH and VH clusters were analyzed. These RFLPs represent genetically characterized VH region polymorphisms and it is hoped that they will facilitate the study of disease correlations as well as further the understanding of the genetics of the immunoglobulin heavy chain genes in humans.
Evidence for linkage between the loci coding for the binding protein for the fourth component of human complement (C4BP) and for the C3b/C4b receptor.
Authors: Rodriguez de Cordoba S , Dykman T R , Ginsberg-Fellner F , Ercilla G , Aqua M , Atkinson J P , Rubinstein P
Published: May 2006

Three pedigrees informative for the segregation of genetic variants of the binding protein for the fourth component of complement (C4BP) and C3b/C4b receptor (C3bR) have been identified. There were 10 informative meioses with no recombinants, indicating a close linkage between the loci encoding C4BP and C3bR, C4BP and C3bR [maximum lod (logarithm of odds of linkage) score: 2.4 at recombinant fraction = 0.0]. In addition, in the four unrelated individuals who were doubly heterozygous (C4BP*1, C4BP*2, C3bR*A, C3bR*B), the infrequent allele C4BP*2 segregated together with the uncommon allele C3bR*B, supporting the hypothesis of linkage between C4BP and C3bR and suggesting that linkage disequilibrium exists between these particular alleles. We conclude that the loci encoding C3bR and C4BP, two functionally related molecules, are linked.
Two endogenous neuropeptides modulate the gill and siphon withdrawal reflex in Aplysia by presynaptic facilitation involving cAMP-dependent closure of a serotonin-sensitive potassium channel.
Authors: Abrams T W , Castellucci V F , Camardo J S , Kandel E R , Lloyd P E
Published: May 2006

We have found that two endogenous neuropeptides in Aplysia, the small cardioactive peptides SCPA and SCPB, facilitate synaptic transmission from siphon mechano-sensory neurons and enhance the defensive withdrawal reflex that these sensory neurons mediate. Single-channel recording revealed that these peptides close a specific K+ channel, the S channel, which is sensitive to cAMP. Moreover, the peptides increase cAMP levels in these sensory neurons. This reduction in K+ current slows the repolarization of the action potential in these cells, which increases transmitter release. In these actions, the SCPs resemble both noxious sensitizing stimuli, which enhance the reflex, and serotonin. Bioassay of HPLC fractions of abdominal ganglion extracts and immunocytochemistry indicate that both the SCPs and serotonin are present in the ganglion and are found in processes close to the siphon sensory neurons, suggesting that these transmitters may be involved in behavioral sensitization. Recent evidence suggests that one group of identified facilitatory interneurons, the L29 cells, does not appear to contain either the SCPs or serotonin but may use yet another facilitatory transmitter. Thus, it appears that several transmitters can converge to produce presynaptic facilitation in the sensory neurons of the defensive withdrawal reflex. All of the transmitters studied here, the SCPs and serotonin, act via an identical molecular cascade: cAMP-dependent closure of the S-K+ channel, broadening of the presynaptic action potential, and facilitation of transmitter release.
Isolation and characterization of a cDNA clone for the complete protein coding region of the delta subunit of the mouse acetylcholine receptor.
Authors: LaPolla R J , Mayne K M , Davidson N
Published: May 2006

A mouse cDNA clone has been isolated that contains the complete coding region of a protein highly homologous to the delta subunit of the Torpedo acetylcholine receptor (AcChoR). The cDNA library was constructed in the vector lambda 10 from membrane-associated poly(A)+ RNA from BC3H-1 mouse cells. Surprisingly, the delta clone was selected by hybridization with cDNA encoding the gamma subunit of the Torpedo AcChoR. The nucleotide sequence of the mouse cDNA clone contains an open reading frame of 520 amino acids. This amino acid sequence exhibits 59% and 50% sequence homology to the Torpedo AcChoR delta and gamma subunits, respectively. However, the mouse nucleotide sequence has several stretches of high homology with the Torpedo gamma subunit cDNA, but not with delta. The mouse protein has the same general structural features as do the Torpedo subunits. It is encoded by a 3.3-kilobase mRNA. There is probably only one, but at most two, chromosomal genes coding for this or closely related sequences.
Compartmental distribution of striatal cell bodies expressing [Met]enkephalin-like immunoreactivity.
Authors: Graybiel A M , Chesselet M F
Published: May 2006

Striatal cell bodies and fibers expressing [Met]enkephalin [( Met]Enk)-like immunoreactivity were studied with two variants of the peroxidase-antiperoxidase method in normal primates and cats and in cats pretreated with colchicine. Strikingly different patterns of [Met]Enk-like immunoreactivity were observed, both in fiber and cell body immunostaining, depending on the technical protocols followed; no single histochemical protocol fully revealed the compartmentalization present. In the dorsal striatum, patches of [Met]Enk-positive neuropil, known to line up with the acetylcholinesterase-poor striatal zones called striosomes, appeared in sections treated by protocols favoring fiber immunostaining. In sections stained by procedures favoring perikaryal staining, the striosomes appeared as Enk-poor patches in a field of immunoreactive cells and neuropil. When cell-body staining was enhanced by pretreatment with colchicine, cells expressing [Met]Enk-like immunoreactivity appeared both in and out of striosomes, and the striosomal neuropil appeared Enk-rich. These results suggest that there are subtypes of Enk-positive neurons in the striatum, including a "colchicine-dependent subtype" in dorsal striosomes, and suggest that the Enk-positive striatal neuropil is also made up of different components. Immunospecificity of this dorsal striosomal system was further demonstrated by the finding that neurons expressing intense immunoreactivity to substance P and to dynorphin B were largely confined to striosomes.
g
-Factor of electron centered in symmetric cavity
Authors: Dehmelt Hans
Published: May 2006

The cyclotron frequency of an electron localized in the center of a cylindrical microwave cavity differs slightly from that in free space. To see this, the system is modeled by a series resonant
lc
circuit for the cyclotron motion connected across the center of a section of two-wire transmission line (the cavity). When the transmission line is infinitely long, its input impedance at terminals in its center is strictly ohmic and equal to half its characteristic impedance. This ohmic impedance completes the
lc
circuit, essentially without changing its resonant frequency but damping it. This is the analog of the cyclotron motion in free space that is damped by spontaneous emission. Free space may be mimicked by a short section of transmission line that is terminated at both ends by resistors equal in value to its characteristic impedance. A slightly lossy cavity is modeled by a section of line terminated by resistors much smaller than the characteristic impedance. When the frequency of the
lc
circuit/cyclotron motion is chosen thus that the line/cavity contains exactly an even number of half-waves, its input impedance will be ohmic and much smaller than its characteristic impedance. Therefore, the frequency of the
lc
circuit/cyclotron motion will be the same as in vacuum but spontaneous emission and associated line broadening will be much reduced. The
g
-factor of the electron measured under these conditions practically equals the free space value. This follows because
g
is determined by the ratio of spin and cyclotron frequencies and the spin practically does not couple to the radiation field.
Synthetic somatomedin C: comparison with natural hormone isolated from human plasma.
Authors: Van Wyk J J , Russell W E , Li C H
Published: May 2006

The biological and immunological properties of a chemically synthesized preparation of somatomedin C (Sm-C) were compared with those of the natural product isolated from human plasma. The two preparations produced identical curves in the radioimmunoassay and radioreceptor assay for Sm-C and in the radioreceptor assay for insulin. They were identical in their ability to stimulate DNA synthesis in confluent BALB/c 3T3 cells previously exposed to platelet-derived growth factor, and the biological activities of both preparations were completely neutralized by a monoclonal antibody raised against native Sm-C. These studies demonstrate that the chemically synthesized product is equivalent to the native molecule in all important respects and that it can be used interchangeably with the natural product for any studies that are contemplated.
Immunofluorescence analysis of the primordial myosin detectable in embryonic striated muscle.
Authors: Sweeney L J , Clark W A , Umeda P K , Zak R , Manasek F J
Published: May 2006

Immunofluorescence analysis showed that the earliest myosin detectable in both the embryonic chicken heart and somitic myotome, the precursor to skeletal muscle, was strongly reactive with two different monoclonal antibodies specific for the heavy chain of cardiac ventricular myosin, but it showed no reactivity with affinity-purified polyclonal antibodies specific for the heavy chains of either fast-twitch or slow-tonic skeletal myosins. The heart remained reactive exclusively with the antibodies to cardiac myosin throughout development, while late embryonic (day 20) skeletal muscles were strongly reactive only with their homologous skeletal myosin antibodies. Our findings suggest that the primordial myosin heavy chain detectable in both forms of embryonic chicken striated muscle, the myotome and the heart, is immunologically distinct from myosins expressed in later embryonic as well as adult skeletal muscles, but it contains antigenic determinants similar to those present in cardiac ventricular myosin.
12S,20-dihydroxyicosatetraenoic acid: a new icosanoid synthesized by neutrophils from 12S-hydroxyicosatetraenoic acid produced by thrombin- or collagen-stimulated platelets.
Authors: Marcus A J , Safier L B , Ullman H L , Broekman M J , Islam N , Oglesby T D , Gorman R R
Published: May 2006

A new metabolite of arachidonic acid, formed during interaction between thrombin- or collagen-stimulated platelets and unstimulated neutrophils, has been demonstrated by both thin-layer radiochromatography and high-performance liquid chromatography. Production of the 3H-labeled metabolite in combined suspensions containing [3H]arachidonate-labeled platelets and unlabeled neutrophils from aspirin-treated donors suggested that platelet 3H-labeled 12S-hydroxy-5,8-cis,10-trans,14-cis-icosatetraenoic acid (12-HETE) was the precursor. This was confirmed by identification of the same product when purified 12-[3H]HETE was added directly to unstimulated neutrophils. Hydrogenation and oxidation of the isolated product, followed by gas chromatography-mass spectrometry showed the structure to be 12S,20-dihydroxyicosatetraenoic acid. These experiments further show that platelet stimuli known to occur in vivo may initiate metabolic interactions between different cell types via the arachidonic acid pathway.
Subcortical projections from ectopic neocortical neurons.
Authors: Jensen K F , Killackey H P
Published: May 2006

There is a high degree of specificity in the efferent connections of the cerebral cortex. In the rodent neocortex, the characteristic band of corticospinal neurons within layer V is present at birth even though changes still occur in the areal distribution of these neurons. Disruption of neocortical development with ionizing radiation before, during, or after the production of neurons destined for layer V results in abnormally located corticospinal neurons. One abnormal location in which corticospinal neurons are found is in ectopic cell clusters beneath the cortical white matter bordering the dorso-medial aspect of the lateral ventricle. Corticospinal neurons only occur in these periventricular ectopias in adult rats irradiated on or before embryonic day 17. A second abnormal location of corticospinal neurons is between layer V and the pial surface. These scattered supragranular corticospinal neurons occur in all adult animals irradiated on embryonic days 16, 17, 18, or 19. The fact that neurons having an unusual position project to a subcortical target appropriate for one neocortical sublayer indicates that neither migratory path nor final position is essential to specifying a subcortical target. In addition, the fact that labeled corticospinal neurons are located in periventricular ectopias only when irradiation occurs on or before embryonic day 17 suggests that the initial projections of corticospinal neurons are determined early in their individual ontogeny prior to migration.
Intermediate filaments in monkey kidney TC7 cells: focal centers and interrelationship with other cytoskeletal systems.
Authors: Celis J E , Small J V , Larsen P M , Fey S J , De Mey J , Celis A
Published: May 2006

Two-dimensional gel electrophoresis of intermediate-sized filament-enriched cytoskeletons of epithelial monkey kidney TC7 cells has shown that they are composed of at least two keratins (isoelectric focusing 36, Mr = 48,500; IEF 46, Mr = 43,500; HeLa protein catalogue number) and vimentin. Indirect immunofluorescence as well as immunoelectron microscopy using antibodies directed against specific polypeptides sometimes revealed a discontinuous staining of keratin-containing filaments. Indirect immunofluorescence analysis of cells stained with keratin or vimentin antibodies also revealed a bright perinuclear staining in 58% of the cells in interphase. Of particular interest were focal centers from which filaments radiated. Double-label immunofluorescence using tubulin and keratin antibodies showed that these centers codistributed with focal arrays of microtubules (most likely centrosomes) in interphase cells but were not colocalized with centrioles in mitosis or, in many cases, with the microtubule organizing centers seen after release from nocodazole treatment. Treatment of TC7 cells with demecolcine (10 micrograms/ml, 20 hr) resulted in a drastic rearrangement of the keratin and vimentin filaments. Likewise, treatment with cytochalasin B (10 micrograms/ml, 1 hr) produced a star-like arrangement of the keratin and vimentin filaments and, in most cases, these codistributed with patches of actin. The results provide evidence for the interaction of intermediate filaments (keratins and vimentin) with both microtubules and microfilaments.
Further clues concerning the vectors essential to regulation of hexose transport, as studied in fibroblast cultures from a metabolic mutant.
Authors: Kalckar H M , Ullrey D B
Published: May 2006

A close study of the metabolic regulation of hexose transport in a hamster fibroblast mutant, highly defective in the enzyme phosphoglucose isomerase (PGI mutant), reveals the requirement for at least three vectors for transport regulation. The downward regulation of the hexose transport system, called the "transport curb," requires (i) a ligand for the transport system, (ii) oxidative energy metabolism, and (iii) some specific enzymes of the glucose-6-phosphate metabolism. Deprivation of glucose was shown to deprive the PGI mutant of UDP hexose, whereas the glucose-fed mutant contained high levels. The parental strain preserved the UDP hexose with or without glucose feeding. Cycloheximide added to the mutant showed two different types of effects. If added at the onset of glucose starvation, the up-regulation of the transport system was scarcely affected. If cycloheximide was added to the mutant at the onset of glucose refeeding, it prevented the development of the glucose-mediated transport curb. In the mutant, the glucose-mediated curb is not derived from energy metabolism but is solely dependent on certain enzymes of glucose-6-phosphate metabolism. The interference of this curb by cycloheximide requires evidently a reassessment, including that of the role of the UDP hexose pathway in regulation of the hexose transport system.
Tumor cell proteinase visualization and quantification using a fluorescent transition-state analog probe.
Authors: Kozlowski K A , Wezeman F H , Schultz R M
Published: May 2006

The fluorescent proteinase transition-state analog inhibitor, dansyl-L-argininal (DnsArgH), may be a selective probe of cysteine and serine-type proteinases in a fibrosarcoma tumor cell line (HSDM1C1). DnsArgH binds with high affinity to proteinases because of its transition-state analog properties, and on association it gives a dramatically increased fluorescent yield. The DnsArgH binding is inhibited by the serine proteinase inhibitor diisopropyl fluorophosphate and by the cysteine proteinase inhibitor p-chloromercuribenzoate. The fluorescence emission appears at its maximum steady-state yield immediately on addition of DnsArgH to the HSDM1C1 fibrosarcoma cells. The immediacy of the DnsArgH reaction supports the contention that DnsArgH binding may be to cell surface-associated proteinases. Quantification of the cell proteinase concentration, by comparison of the fluorescence yield obtained from DnsArgH interactions with bovine trypsin and papain, indicates 10(-15) to 10(-16) mol of proteinase per HSDM1C1 cell. In fluorescence microscopy, DnsArgH fluorescence appears distributed throughout the fibrosarcoma cell without association to organelles. DnsArgH fluorescence from normal fibroblast controls (IMR-90) was found to be substantially lower than in the transformed fibrosarcoma cells, supporting a hypothesis that proteinases have a role in malignancy.
Chloroplast DNA recombination in interspecific hybrids of
Chlamydomonas
: Linkage between a nonmendelian locus for streptomycin resistance and restriction fragments coding for 16S rRNA
Authors: Lemieux Claude , Turmel Monique , Seligy Verner L. , Lee Robert W.
Published: May 2006

Differences in the distribution of
Ava
I and
Bst
EII restriction sites in the chloroplast DNA (cpDNA) of
Chlamydomonas eugametos
and
C. moewusii
have been used to detect extensive cpDNA recombination in the hybrid progeny of these interfertile algae. In the present study, the inheritance of these restriction-site differences was tested for recombination with nonmendelian genetic markers for resistance to streptomycin and erythromycin in interspecific crosses and in hybrid backcrosses to
C. moewusii
. Most of the restriction-pattern markers appear linked to the antibiotic-resistance markers, thus supporting the chloroplast localization of the resistance markers. The streptomycin marker, in particular, shows perfect coordinate inheritance with an
Ava
I band containing one cpDNA fragment and a
Bst
EII band containing two comigrating cpDNA fragments. Molecular hybridization experiments using DNA from the
Ava
I band as a probe show sequence homology between this DNA, the two comigrating
Bst
EII fragments, and cpDNA fragments from
C. reinhardtii
containing the genes for 16S rRNA. The results show the feasibility of using
C. eugametos-C. moewusii
hybrids to identify cpDNA sequences that either contain or are closely linked to nonmendelian genetic markers.
Expression of an immunoglobulin heavy chain gene transfected into lymphocytes.
Authors: Deans R J , Denis K A , Taylor A , Wall R
Published: May 2006

We determined that mouse lymphoid cell lines can be transfected at high efficiencies (10-70%) by a polyoma virus shuttle vector. With this vector, we obtained expression of a cloned mouse alpha heavy chain gene transfected into cell lines representative of all stages in B-lymphocyte development, a T-cell lymphoma line, and 3T3 fibroblasts. Heavy chain gene expression in transfected light chain-producing myeloma cells occurred at levels comparable to those in IgA-secreting myeloma cells. Heavy chains produced in transfected myeloma cells were associated with light chains in membrane-bound IgA. While T-lymphoma cells and fibroblasts were transfected at similar efficiencies to B cells, significantly lower levels of alpha heavy chains were produced. This immunoglobulin gene transfection system provides a powerful approach for defining important regulatory regions in immunoglobulin genes and for identifying lymphoid cell factors involved in immunoglobulin gene expression in B-lymphocyte development.
Cocrystals of the DNA-binding domain of phage 434 repressor and a synthetic phage 434 operator.
Authors: Anderson J , Ptashne M , Harrison S C
Published: May 2006

The amino-terminal domain of the phage 434 repressor forms cocrystals with a synthetic phage 434 operator. The cocrystals diffract to at least 4 A, and x-ray crystallographic analysis of them is in progress. An analysis of the packing in the cocrystals shows that complexes consisting of dimers of amino-terminal domain bound specifically to operators are stacked end to end in longer protein-DNA rods parallel to the unit cell body diagonals. The DNA in the complexes has 10.5 base pairs per turn and a rise per base of 3.26 A--values consistent with B-form DNA--indicating that DNA is neither unwound nor overwound by bound repressor. The packing analysis suggests an approach that might facilitate the cocrystallization of other DNA-binding proteins with the DNA they recognize.
Messenger RNA for an antigen-specific binding molecule from an antigen-specific T-cell hybrid.
Authors: Beaman K D , Ruddle N H , Bothwell A L , Cone R E
Published: May 2006

The T-cell hybridoma 51H7D specifically binds the hapten azobenzenearsonate (ABA). An antiserum (anti-PCIF) specific for antigenic determinants shared by antigen-binding molecules of T cells (TABM) was used to precipitate polyribosomes containing mRNA for TABM from this T-cell hybridoma. The resultant mRNA was translated in vitro. The translated product (TrP51H7D) bound specifically ABA and was bound by both anti-PCIF and anti-T-cell suppressor factor. The parent lymphoma BW5147 yielded a similar translated product with the same antiserum used to isolate specific mRNA containing polysomes. This product (TrPBW) was bound by the antiserum used but did not bind ABA. The specific translated protein from both cells had a pI of approximately equal to 5.0, and apparent molecular weight of 71,000 (reduced) or 145,000 (nonreduced). Both protein products, when treated with guanidine to break down all noncovalent bonds, revealed an elemental peptide of Mr 23,500. The cDNA made from the isolated mRNA had 600-900 bases. mRNA of this size is expected for a protein of Mr 25,000. Our data indicate that a TABM specific for ABA is composed of peptides of Mr 23,500.
Camphor hydroxylase of Pseudomonas putida: vestiges of sequence homology in cytochrome P-450CAM, putidaredoxin, and related proteins.
Authors: Dus K M
Published: May 2006

The amino acid sequences of cytochrome P-450CAM and putidaredoxin of the camphor hydroxylase [camphor, reduced-putida-ferredoxin:oxygen oxidoreductase (5-hydroxylating), EC 1.14.15.1] of Pseudomonas putida are compared to each other and then to the sequences of bovine adrenodoxin and cytochrome b5. The comparisons reveal areas of homology indicating that these four proteins may share a common evolutionary origin. Moreover, homologous segments can be recognized by proper alignment of the sequence of cytochrome P-450CAM to recently determined sequences of other P-450 hemeproteins. Further scrutiny indicates vestiges of internal sequence duplications that have been conserved in limited segments by the constraints of selection over a long time period, while other segments of the sequence diverged. It is predicted that the corresponding reductases will show a similar sequence pattern. The conserved regions of internal sequence repetition may serve a special purpose in protein-protein interactions within the multienzyme systems.
Isolation of an angiotensin II-binding protein from liver.
Authors: Sen I , Bull H G , Soffer R L
Published: May 2006

A protein that specifically binds angiotensin II has been isolated in nearly homogeneous form by two independent approaches after solubilization from rabbit liver particles by treatment with digitonin. The protein purified by either of these methods resembles in size the single radioactive macromolecular component made by using disuccinimidyl suberate to crosslink radioiodinated angiotensin II with its receptor in the solubilized extract. In the first technique, angiotensin II as an affinity ligand specifically extracted the protein from a preparation that had been freed of angiotensin-degrading activity. In the second approach, the angiotensin II-protein complex was specifically precipitated by anti-angiotensin II antibodies and staphylococcal protein A-Sepharose. The protein could be eluted from the affinity column with angiotensin II or 4 M MgCl2. The angiotensin II-protein complex dissociated in the presence of sulfhydryl-containing reagents, and these could therefore be used to elute it from either the chemical or the immunoaffinity-based matrix. This effect of sulfhydryl-containing reagents and the paradoxical observation that the isolated protein after denaturation exhibited a slower electrophoretic mobility in its reduced form that in its unreduced form suggest that the binding configuration of this protein may be sensitive to reduction.
Lipid peroxidation forms ethylene from 1-aminocyclopropane-1-carboxylic acid and may operate in leaf senescence
Authors: Bousquet Jean-Francois , Thimann Kenneth V.
Published: May 2006

An enzyme system is described which oxidizes 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene under physiological conditions. It comprises linoleic acid, pyridoxal phosphate, manganese, and lipoxygenase (linoleate:oxygen oxidoreductase, EC 1.13.11.12). It requires oxygen and is specific for manganese; it can operate but only with greatly reduced yield in the absence of pyridoxal phosphate. An enzyme with the same properties was prepared from microsomal membranes of the seedling shoots of peas. Both have similar reactions to a variety of inhibitors and other reagents. The properties also resemble those of at least two of the
in vivo
systems recorded in the literature. Intact green oat leaves also contain a similar system. Because there is a growing body of evidence that ethylene formation is associated with cell membranes and because the yields of ethylene from the complete system are much higher than those recorded for other enzymes, it may be identical with the
in vivo
system acting in senescent leaves.
Protease inhibitors reduce the frequency of spontaneous chromosome abnormalities in cells from patients with Bloom syndrome.
Authors: Kennedy A R , Radner B S , Nagasawa H
Published: May 2006

Bloom syndrome is an autosomal recessive genetic disease. Cells from patients with this disease are characterized by high levels of chromosome aberrations and sister chromatid exchanges. We show here that the frequency of these chromosomal changes is markedly reduced when the cells are grown in the presence of certain protease inhibitors. In relation to other published data, our results suggest that the primary defect of Bloom syndrome cells may be related to the production of abnormally large amounts of agents, presumably active oxygen species, which are capable of acting like tumor promoters.