bluuebunny/crossref_metadata_2025 huggingface dataset:
Intermolecular crosslinking of fatty acyl chains in phospholipids: use of photoactivable carbene precursors.
Authors: Gupta C M , Radhakrishnan R , Gerber G E , Olsen W L , Quay S C , Khorana H G
Published: May 2006

Phospholipids containing photolysable carbene precursors (beta-trifluoro-alpha-diazopropionoxy and m-diazirinophenoxy groups) in omega-positions of sn-2 fatty acyl chains were prepared. Photolysis of their vesicles produced crosslinked products in 40-60% yields. Crosslinking was mostly intermolecular and occurred by carbene insertion into the C-H bonds of a second fatty acyl chain. Crosslinking products were characterized by (i) their gel permeation behavior, (ii) analysis of products formed by base-catalyzed transesterification, (iii) degradation with phospholipases A2 and C, (iv) gas chromatography/mass spectrometry, and (v) use of mixtures of phospholipids carrying the carbene precursors and a phospholipid containing radioactively labeled fatty acyl groups. Nitrenes generated from the aliphatic or aromatic azido groups in phospholipids were unsatisfactory for forming crosslinks by insertion in C-H bonds.
Chemoattractants stimulate degradation of methylated phospholipids and release of arachidonic acid in rabbit leukocytes.
Authors: Hirata F , Corcoran B A , Venkatasubramanian K , Schiffmann E , Axelrod J
Published: May 2006

When rabbit peritoneal leukocytes were treated with chemoattractants such as fMet-Leu-Phe, an apparent decrease of [3H]methyl incorporation into the lipid fraction from L-[methyl-3H]methionine was observed. This decrease was a result of increased degradation of methylated phospholipids, not of decreased synthesis. Chemotactic peptides did not affect the metabolism of the phospholipids in which [methyl-14C]choline was incorporated. The disappearance of the [3H]methyl group was associated with the release of [1-14C]arachidonic acid from phospholipids prelabeled with these compounds. These findings suggested the activation by chemoattractants of phospholipase A2, an enzyme that removes an unsaturated fatty acid from phospholipids. The order of potency of chemoattractants for the stimulated degradation of phospholipids was in good agreement with that for chemotaxis. Mepacrine (quinacrine) and hydrocortisone inhibited and a phorbol ester enhanced both chemotaxis and phospholipase A2 activity. These results, taken together, suggest close association of the metabolism of methylated phospholipids with chemotaxis in rabbit peritoneal leukocytes.
DNA endonucleases associated with the avian myeloblastosis virus DNA polymerase.
Authors: Samuel K P , Papas T S , Chirikjian J G
Published: May 2006

A DNA endonuclease, Endo-I, which cleaves superhelical DNAs, has been isolated from avian myeloblastosis virions stripped of their coats by mild detergent treatment. The enzyme has a broad pH optimum around 7.5-8.0 and requires Mg2+ for activity. A second endonuclease, Endo-II, with a requirement for Mn2+, also present in viral cores, copurified with avian myeloblastosis virus alpha beta DNA polymerase (reverse transcriptase, RNA-dependent DNA nucleotidyltransferase) and similarly cleaved superhelical DNAs. Heat denaturation and sodium fluoride and N-ethylmaleimide inhibition studies were carried out to demonstrate a possible relationship between the two endonucleases and the viral DNA polymerase and RNase H activities. It appears that Endo-II may be an intrinsic activity of the polymerase.
Initial glycosylation of proteins with acetylgalactosaminylserine linkages.
Authors: Strous G J
Published: May 2006

Epithelial glycoprotein like that produced by the gastric surface consists of a polypeptide chain rich in serine and threonine; to these amino acid residues oligosaccharide chains of variable length are linked. The linking sugar is acetylgalactosamine. To find out whether the initial glycosylation takes place at the ribosomal level. I treated purified peptidyl-tRNA, derived from rat gastric membrane-bound polysomes, with alkali in the presence of boro[3H]hydride. Alkali specifically splits glycosidic bonds between serine or threonine and oligosaccharide side chains (beta-elimination reaction). The linking sugar is converted to an alditol and simultaneously labeled. GalNAc was identified as the linking sugar by paper chromatography. Furthermore, nascent peptides with lengths between 40 and 60 amino acid residues already contained this linking sugar. Gel filtration on Bio-Gel P-2 of 3H-labeled saccharides revealed that nascent chains contained mainly monosaccharides, but some di- or trisaccharides were found with GalNAc as the linkage sugar. These findings demonstrate that initial glycosylation of epithelial glycoprotein occurs at the ribosomal level rather shortly after the nascent peptide chain has reached the cisternal lumen of the endoplasmic reticulum.
Increased levels of adenosine 3',5'-cyclic monophosphate in Tetrahymena stimulated by glucose and mediated by Ca2+ and epinephrine.
Authors: Nandini-Kishore S G , Thompson G A
Published: May 2006

Stationary phase Tetrahymena cells exposed to 0.5-1% glucose experience an 8-fold increase in intracellular cyclic AMP within 1 hr. The increase follows an enhanced uptake of Ca2+ triggered by the glucose addition. Inhibiting the Ca2+ influx with ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid blocks the increase in cyclic AMP, but the block can be reversed by the addition of epinephrine. Both this epinephrine effect and the glucose-induced increase in cyclic AMP are counteracted by the beta-adrenergic inhibitor dichloroisoproterenol.
Regulation of dihydrofolate reductase synthesis in an overproducing 3T6 cell line during transition from resting to growing state
Authors: Wiedemann Leanne M. , Johnson Lee F.
Published: May 2006

We have isolated a methotrexate (MTX)-resistant clone of mouse 3T6 cells, designated M50L3, which grows normally in the presence or absence of 50 μM MTX and produces a level of dihydrofolate reductase (DHFR; 5,6,7,8-tetrahydrofolate:NADP
+
oxidoreductase, EC 1.5.1.3) that is increased about 300-fold compared to the parental 3T6 cells. The cells retain the ability to rest in the G
0
state when maintained in medium containing 0.5% calf serum and can be stimulated to reenter the cell cycle by increasing the serum concentration to 10%. The rate of accumulation of DHFR in resting M50L3 cells is about 1/25th of that in exponentially growing cells. When resting cells are stimulated to reenter the cell cycle, the rate of accumulation of DHFR starts to increase at about 8 hr and reaches a maximum (25-fold increase) at about 16 hr after stimulation. Pulse-labeling experiments show that the increase in DHFR accumulation is due to an increased rate of synthesis. This increase occurs at about the same time the cells enter S phase. However, inhibitors of DNA synthesis have no effect on the increase in DHFR accumulation after serum stimulation, indicating that there is no tight coupling of the two events. Actinomycin D inhibits the subsequent increase in DHFR accumulation if added 8 hr after stimulation but has no effect if added 16 hr after stimulation. This is consistent with the idea that the increase in DHFR gene expression depends on transcription of the gene and that DHFR mRNA synthesis begins at about the time the cell initiates DNA replication. DHFR gene expression appears to be regulated in the same manner in the overproducing cells as we found in the parental 3T6 cells [Johnson, L. F., Fuhrman, C. L. & Wiedemann, L. M. (1978)
J. Cell. Phys.
97, 397-406]. Therefore, the alterations that are responsible for DHFR overproduction (presumably DHFR gene amplification) do not interfere with the ability of the cell to regulate the rate of synthesis of the enzyme after serum stimulation.
Continuous variation caused by genes with graduated effects.
Authors: Matthysse S , Lange K , Wagener D K
Published: May 2006

The classical polygenic theory of inheritance postulates a large number of genes with small, and essentially similar, effects. We propose instead a model with genes of gradually decreasing effects. The resulting phenotypic distribution is not normal; if the gene effects are geometrically decreasing, it can be triangular. The joint distribution of parent and offspring genic value is calculated. The most readily testable difference between the two models is that, in the decreasing-effect model, the variance of the offspring distribution from given parents depends on the parents' genic values. The more the parents deviate from the mean, the smaller the variance of the offspring should be. In the equal-effect model the offspring variance is independent of the parents' genic values.
Enkephalin-containing amacrine cells in the avian retina: immunohistochemical localization.
Authors: Brecha N , Karten H J , Laverack C
Published: May 2006

The distribution of enkephalin-like immunoreactivity within the avian retina was studied by immunofluorescence and immunoperoxidase techniques with antiserum to [Met5]enkephalin and [Leu5]enkephalin. Formaldehyde-fixed retinae were sectioned and incubated in antiserum to either [Met5]- or [Leu5]enkephalin. Specificity to the antiserum was established by absorption of the antiserum with synthetic [Met5]- or [Leu5]enkephalin at 1 mM. Positive immunohistochemical staining for enkephalin was observed in the somata of amacrine cells and their processes within the inner plexiform layer. A large number of enkephalin-containing amacrine cells were distributed throughout the retina, and their density appeared to be greatest within central retinal regions. The majority of labeled amacrine cells were about 7.5 micrometers in diameter although, occasionally, amacrine cells were observed that were 12--15 micrometers in diameter. Amacrine cells had a cell-to-cell spacing of approximately 40 micrometers within central retinal regions. Labeled processes of the amacrine cells were observed to project into the inner plexiform layer where they arborized as a fine plexus, within laminae 1, 3--5 of the inner plexiform layer. These observations demonstrate the existence of opioid peptides in seemingly select populations of amacrine cells within the retina. The localization of enkephalin-like immunoreactivity within the retina suggests that opioid peptides play a specific and unique functional role in retinal processing.
Electron microscopic analysis of transcription: mapping of initiation sites and direction of transcription.
Authors: Brack C
Published: May 2006

An electron microscope technique is described that allows rapid characterization of transcription in vitro. DNA is transcribed with Escherichia coli RNA polymerase in vitro, and the RNA is hybridized to its template. Measurement of the resulting transcription R-loop molecules allows accurate mapping of transcription initiation sites (promoter sites) and analysis of the direction and rate of transcription and the level of transcription from each initiation site. The two major early promoters pR and pL of bacteriophage lambda have been mapped within 0.1-0.3 map units of the known positions and three additional sites have been confirmed. Six transcription initiation sites have been preliminarily mapped on plasmid pSF2124 DNA.
Antineoplastic activity of poly(L-lysine) with some ascites tumor cells.
Authors: Arnold L J , Dagan A , Gutheil J , Kaplan N O
Published: May 2006

We have found that poly(L-lysine) can be a very effective agent in preventing the growth of Ehrlich ascites tumors in mice. When given optimal doses of poly(L-lysine) (Mr 60 x 10(3)) intraperitoneally for 5 consecutive days, beginning on day 1 after inoculation with Ehrlich ascites cells. White Swiss mice show nearly a 100% remission from subsequent tumor growth. Rechallenge of "cured" animals with tumor cells, however shows no long-term immunological protection. In tissue culture, poly(L-lysine) shows a related potent cytotoxicity with HeLa cells; interestingly, the D isomer. In addition, there is a strong molecular weight dependence in that the small polylysine (Mr 3 x 10(3)) possesses less than 1/20th the cytotoxicity of large polymers (Mr 70 x 10(3)) on a weight basis in both cell culture and animal studies. At the same time, none of these lysine polymers gives any significant increase in life span to BDF1 mice infected with L1210 murine leukemia cells. We have also further explored the mechanism by which the polylysines express their cytotoxicity. These data indicate that lysine polymers show cell specificity in their action and in some cases they may be beneficial as potent antineoplastic agents, particularly when molecular weight is taken into consideration.
Evidence from ultraviolet absorbance measurements for a codon-induced conformational change in lysine tRNA from Escherichia coli.
Authors: Möller A , Wild U , Riesner D , Gassen H G
Published: May 2006

From experiments with equilibrium dialysis it was concluded earlier that formation of the codon-anticodon complex triggers a conformational change in the tertiary structure of tRNAPhe from Escherichia coli. A similar conformational transition is demonstrated here in the poly(A)/tRNALys system. C-G-A or C-G-A-A was used as a probe for the conformational transition in tRNA. These probes bound to tRNAPhe and tRNALys more strongly in the presence of the corresponding codons than in the absence. In order to verify these data by an independent method, the decrease in absorbance at 300 nm that occurs on formation of the codon-anticodon complex in tRNALys (which contains 2-thio-5-methylaminomethyluridine, s2mam5U) was used. The binding constants for formation of A3 . tRNALys (Ka = 2.4 . 10(4) M-1) and A4 . tRNALys (Ka = 2.5 . 10(5) M-1) are very close to those obtained by equilibrium dialysis. In the presence of C-G-A the apparent binding constant of A3 to tRNA was raised 10-fold to 2.5 . 10(-5) M-1. It was calculated that the constant for the binding of C-G-A to the binary complex A3 . tRNALys is approximately 2 . 10(4) M-1, whereas binding to the free tRNA is lower than 10(3) M-1. Under appropriate conditions binding of A3 to tRNALys can be induced directly by the addition of C-G-A. These data demonstrate that codon-anticodon complex formation induces a conformational change in the tRNA that as a consequence allows the binding of a trinucleoside diphosphate, presumably to the T-psi-G region.
A factor in serum and amniotic fluid is a substrate for the tRNA-modifying enzyme tRNA-guanine transferase.
Authors: Katze J R , Farkas W R
Published: May 2006

Q factor, a substance found in animal serum that enables cultured mammalian cells (L-M) to produce tRNA containing queuine (the base of "nucleoside Q", queuosine), has been purified to homogeneity from bovine amniotic fluid. Q factor causes the appearance of Q-containing tRNAAsp in the L-M cells cultivated in serum-free medium, and this was used as an assay to monitor the purification of Q factor. Q factor is a competitive inhibitor of guanine for rabbit reticulocyte tRNA-guanine trnsferase, with a K1 of 4.5 x 10(-8) M. Q factor is inactivated in both the L-M cell and tRNA-guanine transferase assays by treatment with periodate or cyanogen bromide, both of which react with queuine. In L-M cells, nearly complete conversion of Q-free to Q-containing tRNAAsp is observed within 24 hr after addition of pure Q factor to the medium; actinomycin D, cycloheximide, and cycloleucine, inhibitors of RNA synthesis, protein synthesis, and nucleic acid methylation, respectively, do not inhibit this conversion. The product of the reaction, catalyzed by pure rabbit reticulocyte tRNA-guanine transferase, between Q factor and rabbit reticulocyte Q-free tRNAHis is chromatographyically indistinguishable from Q-containing tRNAHis.
Receptor-mediated endocytosis: insights from the lipoprotein receptor system.
Authors: Brown M S , Goldstein J L
Published: May 2006

The low density lipoprotein (LDL) receptor system coordinates the metabolism of cholesterol, an essential component of the plasma membrane of all mammalian cells. Study of this system has led to an enhanced understanding of the cellular basis of cholesterol homeostasis. It has also brought into focus an important mechanism of metabolic regulation--the process of receptor-mediated endocytosis. In this article, we first describe the receptor-mediated endocytosis of LDL, a sequence of events in which receptor binding and internalization are coupled in specialized regions of the plasma membrane called coated pits. Second, we trace the cellular functions of the cholesterol derived from internalized LDL. Third, genetic evidence is presented to indicate that both the binding and internalization of LDL are mediated by a single receptor molecule that contains two active sites, one mediating binding and the other internalization. Finally, the characteristics of the LDL receptor system are used to suggest models for receptor systems in general.
Role of carbohydrate in biological function of the adhesive glycoprotein fibronectin.
Authors: Olden K , Pratt R M , Yamada K M
Published: May 2006

We have investigated the role of the carbohydrate moiety in the biological activity of fibronectin in vitro by using tunicamycin to inhibit the glycosylation of this glycoprotein. Tunicamycin is a glucosamine-containing antibiotic that specifically inhibits glycosylation of protein asparaginyl residues mediated by dolichol pyrophosphate. Fibronectin synthesized in the presence of 0.5 microgram of tunicamycin per ml was not glycosylated, as determined by amino sugar analysis, lack of incorporation of [14C]glucosamine and [3H]mannose, and concanavalin A binding studies. Nonglycosylated fibronectin that was isolated from chicken embryo fibroblasts and added to transformed cells in vitro was as effective as the glycosylated protein in promoting a more normal fibroblastic phenotype, including cell flattening, elongation of cell processes, and parallel alignment of cells. The nonglycosylated protein was also as effective as the glycosylated species in mediating cell attachment to collagen and spreading on plastic, as well as in agglutination of formalin-fixed sheep erythrocytes. The nonglycosylated protein was twice as sensitive as the glycosylated protein to proteolytic hydrolysis in vitro as had been suggested by previous studies with intact cells [Olden, K., Pratt, R.M. & Yamada, K.M. (1978) Cell 13, 461-473]. We conclude that the carbohydrate moiety of fibronectin is not required for the mediation of a number of biological activities characteristic of this glycoprotein.
Junctional modulation in mouse embryonal carcinoma cells by Fab fragments of rabbit anti-embryonal carcinoma cell serum.
Authors: Dunia I , Nicolas J F , Jakob H , Benedetti E L , Jacob F
Published: May 2006

Mouse embryonal carcinoma PCC4 cells are connected by extensive gap and tight junctions. When the cells are incubated in a medium containing Fab fragments against embryonal carcinoma F9 cells, they round up and a process of junctional removal is initiated. In particular, gap junctions are internalized and after 30 hr of incubation with the anti-F9 Fab fragments both tight and gap junctions are no longer present at the cell surface; however, the cells are still in contact by means of small attachment sites.
Dynamics of activated processes in globular proteins.
Authors: McCammon J A , Karplus M
Published: May 2006

A procedure for the dynamical simulation of activated processes, such as ligand binding and enzymatic reactions, in a globular protein is outlined. Preliminary calculations of the transition state geometry and barrier crossing trajectories are presented for a model reaction, the rotation of an aromatic ring in the bovine pancreatic trypsin inhibitor. The results show that repulsive nonbonded interactions between the ring atoms and the atoms in the surrounding protein matrix determine the dynamical character of the reorientation process; the nonbonded interactions are the source of the rotational barrier and of the impulses that speed up or slow down the ring motion during the barrier crossings.
Adenine aminohydrolase: occurrence and possible significance in trypanosomid flagellates.
Authors: Kidder G W , Nolan L L
Published: May 2006

Adenine aminohydrolase (EC 3.5.4.2) from four species of Leishmania and from Crithidia fasciculata was examined for specific activities, affinity for substrate (adenine), and stability to heat. All were found to be strongly and non-competitively inhibited by both coformycin and deoxycoformycin, two tight-binding inhibitors of adenosine deaminase (adenosine aminohydrolase, EC 3.5.4.4). Deoxycoformycin is the more potent inhibitor of the two. Neither inhibitor was active against the purine phosphoribosyltransferases. When deoxycoformycin was added to the defined growth medium containing hypoxanthine as the purine source, the growth of C. fasciculata was unaffected, but when adenine was the purine source for the organism, severe inhibition resulted. This implies that hypoxanthine is the obligatory base for nucleotide synthesis and that the adenine phosphoribosyltransferase (AMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.7) is, in some manner,idenied access to exogenous substrate.
Covalently bound non-coenzyme phosphorus residues in flavoproteins:
31
P nuclear magnetic resonance studies of
Azotobacter
flavodoxin
Authors: Edmondson Dale E. , James Thomas L.
Published: May 2006

In addition to the 5′-phosphate ester on its flavin mononucleotide (FMN) moiety, flavodoxin from
Azotobacter vinelandii
contains 2 moles of tightly bound phosphate. One non-coenzyme phosphate group is covalently bound to the protein, as it remains with the protein on acid precipitation, whereas the other phosphate is released. The invariance of the
31
P nuclear magnetic resonance chemical shift of the covalently bound phosphate (-0.8 ppm relative to 85% phosphoric acid) with pH, even in the presence of protein denaturants, implies it is in a diester linkage to the protein. Because no evidence could be found for the presence of covalently bound sugars, nucleotides, or phospholipids, it is suggested that the phosphate residue forms a diester linkage with two hydroxyl amino acids in the protein. The only other suggestion of a phosphodiester linkage in proteins is from previous studies on pepsin and pepsinogen [Perlmann, G. E. (1955)
Adv. Prot. Chem.
10, 1-30]. The observed changes in
31
P chemical shift with pH show that the covalent phosphorus in pepsinogen has ionization properties of a monoester rather than a diester. The
31
P resonance of the FMN phosphate occurs at -5.6 ppm in native
Azotobacter
flavodoxin. No ionization of the protein-bound FMN phosphate is observed since the chemical shift does not change appreciably in the pH range of 5.5-9.5. The chemical shift data suggest, but do not prove, that the coenzyme phosphate in its protein-bound form is dianionic. Chemical analysis of several other flavoenzymes from a variety of sources shows the presence of covalently bound phosphorus in quantities stoichiometric with the flavin content in most of the enzymes tested. Thus, the presence of covalent phosphorus in flavoenzymes may be a general phenomenon with currently unknown catalytic significance.
In vivo distribution and turnover of fluorescently labeled actin microinjected into human fibroblasts.
Authors: Kreis T E , Winterhalter K H , Birchmeier W
Published: May 2006

Graessmann's microinjection technique was chosen to introduce fluorescently labeled muscle actin and other proteins into WI-38 human fibroblasts. The injected cells were examined during culture by fluorescence and reflection contrast microscopy. Within 30 min after injection, rhodamine-labeled actin was incorporated into a distinct network of fluorescent filaments, resembling the stress fibers observed by classical immunofluorescence microscopy. Cytochalasin b prevented the formation of these fibers, but colchicine did not. Neighboring fibers often converged into distinct focal points that appeared to be concentrated near the base of the cell. Examination of these fluorescent fibers and focal points by reflection contrast microscopy confirmed their close location to the substratum. After 6 hr of culture, fluorescent actin and the control proteins were segregated into granules located mainly near the cell nucleus. Thus, the injected actin both enters the intrinsic actin pool and participates in an assembly and disassembly of filamentous structures. Segregation into granules traces the natural turnover of this protein within the cell.
Immunotitration of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in various physiological states.
Authors: Hardgrave J E , Heller R A , Herrera M G , Scallen T J
Published: May 2006

The immunotitration of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase [mevalonate:NADP+ oxidoreductase [CoA-acylating), EC 1.1.1.34], the major regulatory enzyme in cholesterol biosynthesis, by HMG-CoA reductase antiserum is described. This technique provides the advantage that one can rapidly measure relative changes in both enzyme concentration and enzyme activity in different physiological states by using enzyme samples all the way from crude liver microsomes to purified enzyme. Regarding the diurnal rhythm of HMG-CoA reductase, the major difference noted between animals killed near the middle of the dark period (D 4.5) compared to animals killed near the middle of the light period (L 4.5) was in the concentrations of HMG-CoA reductase present, rather than in the activity of the enzyme, with substantially more enzyme found near mid-dark. Cholestyramine treatment resulted in both an increased concentration of HMG-CoA reductase and a catalytically more active enzyme. On the other hand, cholesterol feeding resulted in both a decreased concentration of HMG-CoA reductase and a catalytically less active enzyme.
Unstable transpositions of his4 in yeast.
Authors: Greer H , Fink G R
Published: May 2006

Unstable transpositions in yeast have been selected in which the his4C gene from chromosome III is inserted into chromosome XII. This event is associated with the generation of a recessive lethal mutation, resulting from the integration of his4C into an essential gene. Strains with these transpositions are viable as diploids or aneuploids for chromosome XII. The event that generates the transpositions does not lead reciprocally to a deletion on chromosome III, implying that synthesis of a new copy of his4C and subsequent transposition may have occurred. The his4C transpositions are unstable and give rise to C- segregants at a high frequency, as a result of either precise excision of the his4C gene (restoring function of the gene into which insertion had occurred) or chromosome loss.
Human cell lines expressing intestinal alkaline phosphatase.
Authors: Benham F J , Harris H
Published: May 2006

At least three loci determine human alkaline phosphatases [orthophosphoric-monoester phosphohydrolase (alkaline optimum), EC 3.1.3.1]: one coding for the placental form of the enzyme, at least one coding for the intestinal forms, and at least one for the liver, bone, and kidney forms. The alkaline phosphatase in cell line D98/AH-2 has been characterized by inhibition, thermostability, and electrophoretic studies. It is intestinal in type and resembles the fetal intestinal form somewhat more closely than the adult intestinal form. Intestinal alkaline phosphatase was found in the related cell lines Detroit 98, D98/S, and D98/AH-R. No placental alkaline phosphatase could be detected in any of these cell lines. This series of cell lines are believed, on the basis of earlier investigations, to be HeLa in origin but other HeLa cell lines show placental alkaline phosphatase. Loss of expression of the placental alkaline phosphatase locus probably occurred prior to the separation of Detroit-98 from the lineage leading to other HeLa cell lines and this has persisted in the Detroit-98 derivatives D98/AH-2, D98/S, and D98/AH-R. Another possibility is that placental alkaline phosphatase expression only appeared in the HeLa lineage subsequent to the separation of Detroit-98.
Amyloid protein SAA is an apoprotein of mouse plasma high density lipoprotein.
Authors: Benditt E P , Eriksen N , Hanson R H
Published: May 2006

Mouse plasma contains a protein antigenically related to mouse protein AA, the principal protein derived from tissue deposits of amyloid substance in mice. In this work the plasma antigen, SAA, was found mainly as a high molecular weight form (2 x 10(5)) residing for the most part in the density interval 1.063-1.21 g/cm3 (high density lipoprotein, HDL); the largest amount of SAA, absolute and relative to total protein, was found in the density interval 1.125-12.1 g/cm3 (HDL3). When apoproteins of the mouse HDL obtained by delipidation of the lipoprotein particles were chromatographed in acid/urea, the antigenic activity appeared in the 10,000- to 15,000-dalton portion of the apoprotein complex. In these characteristics mouse SAA closely resembles human SAA and the behavior of the protein related to amyloid protein AA indicates that is one of the apoproteins of the HDL complex in both species. Therefore we suggest that it be named apoSAA.
The Bezold reflex: A special case of the left ventricular mechanoreceptor reflex
Authors: Estrin J. A. , Emery R. W. , Leonard J. J. , Nicoloff D. M. , Swayze C. R. , Buckley J. J. , Fox I. J.
Published: May 2006

Our previous finding that increasing myocardial contractility caused reflex systemic hypotension, the left ventricular (LV) mechanoreceptor reflex, suggested that the classical Bezold reflex (systemic hypotension and bradycardia after intracoronary administration of veratrum alkaloids) may be initiated by these same LV mechanoreceptors. In our working LV preparation with the coronary and systemic circulations isolated and perfused separately, intracoronary injection of veratrum alkaloids, like that of catecholamines or ouabain, had a positive inotropic effect which produced the hypotensive response typical of the LV mechanoreceptor reflex. To test directly if veratridine's positive inotropic effect initiates the Bezold reflex, verapamil, which blocks the slow Ca
2+
channels of myocardial cells but leaves intracardiac nerves unaffected, was injected by the intracoronary route to prevent the increased contractility from intracoronary injection of veratridine which also abolished the reflex hypotension, demonstrating conclusively that increasing myocardial contractility and thereby activating LV mechanoreceptors but not chemoreceptors initiates the Bezold reflex. Contrariwise, decreasing contractility or cardiac asystole by administration of tetrodotoxin, verapamil, or EDTA resulted in an increase in the systemic resistance, indicating that changes in the magnitude of the stimulus initiating the LV mechanoreceptor reflex (i.e., changes in myocardial contractility) lead to directionally opposite changes in peripheral resistance, as in the sino-aortic mechanoreflexes. Thus, it is concluded that the Bezold reflex is a special case of the LV mechanoreceptor reflex. The latter, by means of feedback mechanisms, functions normally by continuously matching the peripheral resistance to the LV contractile state so as to maintain the arterial pressure constant, thereby playing an important role in blood pressure regulation.
Photoaffinity labeling of cyclic-AMP- and AMP-binding proteins differentiating Dictyostelium discoideum cells.
Authors: Wallace L J , Frazier W A
Published: May 2006

Cyclic-AMP-binding proteins play important roles during the differentiation of the cellular slime mold Dictyostelium discoideum. The photoaffinity reagent 8-N3-cyclic [32P]AMP has been used to label developmentally regulated cyclic-AMP-binding proteins of intact cells, membranes, and cytoplasm. 8-N3-Cyclic AMP is a chemoattractant for differentiated D. discoideum cells and is a substrate for the membrane phosphodiesterase (mPDE). When mPDE is inhibited, the only specifically labeled protein on intact cells has a molecular weight of 40,000 on sodium dodecyl sulfate gels. The developmental time course of appearance of this protein and its high specificity for cyclic AMP identify it as the cell surface chemotactic receptor for cyclic AMP. The concentration dependence of labeling of this protein is consistent with the measured chemotactic potency of 8-N3-cyclic AMP, which is about 1/100th that of cyclic AMP. Three developmentally regulated proteins (Mr 26,000, 33,000, and 36,000) of the soluble fraction (cytoplasm) are labeled by the photoaffinity reagent and are specific for cyclic AMP. By analogy with other systems, these may be regulatory subunits of protein kinases. The mPDE of ghosts or plasma membrane fractions converts the reagent to 8-N3-[32P]AMP, which specifically photoaffinity labels a protein of Mr 42,000 associated with the cytoplasmic face of the plasma membrane.
Identification of gamma-glutamyl phosphate in the alpha 2 chains of chicken bone collagen.
Authors: Cohen-Solal L , Cohen-Solal M , Glimcher M J
Published: May 2006

Purified components of chicken bone collagen contain approximately 4 atoms of organic phosphorus per mol of collagen, located principally in the alpha 2 chains. Previous analyses have demonstrated the absence of O-phosphoserine, O-phosphothreonine, and other phosphorylated hydroxy amino acids, phosphoamidated amino acids, and phosphorylated sugars. In the present report we establish that chicken bone collagen contains gamma-glutamyl phosphate. This was accomplished by the isolation of tritiated alpha-amino-delta-hydroxyvaleric acid after reductive cleavage with NaB[3H]H4 of the gamma components, the alpha 2 chains, and peptides enriched in organic phosphorus that were derived from the alpha 2 chains. Tritiated alpha-amino-delta-hydroxyvaleric acid was not detected in any of the following unphosphorylated proteins after cleavage with NaB[3H]H4:albumin and lysozyme, the alpha 2 chains of several unmineralized tissues, and, most importantly, dephosphorylated alpha 2 chains of chicken bone collagen. The alpha 2 chain of chicken bone collagen is the first structural protein found to contain an acyl phosphate.
Hydroperoxides can modulate the redox state of pyridine nucleotides and the calcium balance in rat liver mitochondria
Authors: Lötscher Hans Ruedi , Winterhalter Kaspar H. , Carafoli Ernesto , Richter Christoph
Published: May 2006

When rats are fed a selenium-deficient diet, the glutathione peroxidase activity in liver mitochondria decreases within 5 weeks to 0-6% of that of control animals fed on a diet supplemented with 0.5 ppm of selenium as sodium selenite. Analysis of the temperature dependence of energy-linked Ca
2+
uptake by means of Arrhenius plots reveals two breaks (at around 11°C and 24°C) in mitochondria isolated from selenium-supplemented animals, whereas in selenium-deficient rats the break at 11°C is absent. Ca
2+
-loaded mitochondria of selenium-supplemented rats—i.e., with active glutathione peroxidase in the matrix—lose Ca
2+
rapidly, with a concomitant oxidation of endogenous NAD(P)H, when exposed to
t
-butyl hydroperoxide or H
2
O
2
. In contrast, in selenium deficiency,
t
-butyl hydroperoxide and H
2
O
2
induce neither a release of Ca
2+
nor an oxidation of NAD(P)H. The peroxide-induced oxidation of NAD(P)H is reversible in the presence of succinate when no Ca
2+
has been taken up. When Ca
2+
has previously been accumulated, however, the oxidation of NAD(P)H is irreversible. Enzymatic analysis of mitochondrial pyridine nucleotides reveals that the peroxide-induced oxidation of NAD(P)H in Ca
2+
-loaded mitochondria leads to a loss of NAD
+
and NADP
+
. It is proposed that the redox state of mitochondrial pyridine nucleotides can be or is in part controlled by glutathione peroxidase and glutathione reductase and is a factor in the balance of Ca
2+
between mitochondria and medium.
Biological detection of specific mRNA molecules by microinjection.
Authors: Liu C P , Slate D L , Gravel R , Ruddle F H
Published: May 2006

We have used direct microinjection of messenger RNA into individual mouse and human cells to assay for specific translation products. We have been able to detect the synthesis of human fibroblast interferon, thymidine, kinase, hypoxanthine phosphoribosyltransferase, adenine phosphoribosyltransferase, and propionyl-CoA carboxylase in response to injected mRNA. Using the interferon system as a model, we have quantitated interferon synthesis and followed partial purification of interferon mRNA sequences on sucrose density gradients. The methods we have utilized should be applicable to other systems in which sensitive assays exist for gene products and should provide a screening procedure for isolating specific mRNA sequences.
Transcription promotes recA-independent recombination mediated by DNA-dependent RNA polymerase in Escherichia coli.
Authors: Ikeda H , Matsumoto T
Published: May 2006

The Rpo-mediated recombination of phage lambda takes place independently of the recA function and is promoted by DNA-dependent RNA polymerase of Escherichia coli [Ikeda, H. & Kobayashi, I. (1977) Proc. Natl. Acad Sci. USA 74, 3932--3936]. The crossovers were particularly frequent to the cIII-N and N-cII regions which are transcribed actively. To determine whether the transcription process required for the recombination is the initiation step or the chain elongation step, we have examined the effect of bacterial rho mutation, which affects transcription termination, on the distribution of crossover points in the lambda phage genome. The crossovers in the cII-S interval took place more frequently in rho mutant strains than in wild-type strains. Analysis of lambda mRNA showed that much more O-P-Q mRNA is synthesized in the rho mutant cells than in the wild-type cells and is largely produced by the readthrough from the PR promotor. These results strongly suggest that the chain elongation in transcription plays an essential role in this recombination. Physical analysis of the recombinant phage DNA showed that this recombination is a legitimate type. Models are presented to explain how the transcription complex can promote this recA-independent recombination.
Enhanced mitotic recombination in a ligase-defective mutant of the yeast Saccharomyces cerevisiae.
Authors: Game J C , Johnston L H , von Borstel R C
Published: May 2006

The temperature-sensitive Saccharomyces cerevisiae cell cycle mutant cdc9 is defective in DNA ligase, and the DNA synthesized at the restrictive temperature contains many single-strand breaks. We find that holding a diploid homozygous for cdc9 at the restrictive temperature and then plating cells at the permissive temperature gives rise to increased intragenic and intergenic recombination. In the latter case, recombinants signaled by the ade2 locus rise to about 4% of the survivors after 6 hr of incubation at the restrictive temperature. We propose that the single-strand breaks left in DNA synthesized at the restrictive temperature may lead to recombination.
Localization of spontaneously hyperactive B cells of NZB mice to a specific B cell subset.
Authors: Nakajima P B , Datta S K , Schwartz R S , Huber B T
Published: May 2006

NZB mice produce numerous autoantibodies and have a subpopulation of B cells characterized by marked spontaneous hypersecretion of IgM. The latter trait is determined by autosomal genes, in F1 hybrids of NZB and normal strains. We tested the hypothesis that the hypersecreting B cells of NZB mice are contained within a specific subpopulation by examining (CBA/N X NZB)F1 hybrids. CBA/N mice have an X-linked recessive defect that results in the absence of a functionally distinct B cell subpopulation and impaired antibody responses. The hyperactivity of B cells, characteristic of the NZB parent, was transmitted to the F1 female, but was not expressed by the F1 male, which manifested the CBA/N B cell hyporesponsiveness. By contrast, the NZB xenotropic virus was expressed equally by both male and female F1 mice. We conclude that the NZB B cell abnormality resides within the B cell subpopulation affected by the CBA/N mutation.
Cell cytotoxicity due to specific influenza antibody production in vitro after recent influenza antigen stimulation.
Authors: Greenberg S B , Six H R , Drake S , Couch R B
Published: May 2006

Peripheral blood leukocytes, obtained from volunteers after vaccination or natural illness with influenza, were assayed for cytotoxicity against influenza virus-infected cells. Approximately 7 days after vaccination or the onset of respiratory illness, peak cytotoxicity was demonstrated in a chromium-release assay. Secretion of specific antibody against hemagglutinin from the leukocytes during in vitro incubation was demonstrated in quantities that would mediate the cell cytotoxicity observed. Antibody secretion was inhibited by exposure to cycloheximide but not by exposure to trypsin. The secretion of antibody against hemagglutinin from peripheral blood leukocytes occurred only at the time of maximal cytotoxicity. We thus demonstrate secretion of specific antibody in vitro after recent viral antigen stimulation. Moreover, this antibody is capable of conveying cytotoxic capacity to peripheral blood leukocytes that may be important in the recovery process from acute viral infection.
Reovirus-induced modification of cap-dependent translation in infected L cells.
Authors: Skup D , Millward S
Published: May 2006

The translational apparatus in cell-free extracts prepared from L cells infected with reovirus undergoes a time-dependent transition from cap dependence to cap independence. Extracts from uninfected L cells translate capped reovirus mRNA at high efficiency and synthesize the expected three size classes of reovirus polypeptides, and the translation is sensitive to m7G(5')ppp. This same extract translates uncapped mRNA at a much lower efficiency. In contrast, extracts from infected L cells translate uncapped reovirus mRNA at high efficiency and synthesize the correct three size classes of polypeptides, and the translation is not sensitive to inhibition by m7G(5')ppp. Infected cell extracts translate capped mRNA at reduced efficiency (a,proximately 25%), the translation is not sensitive to inhibition by m7G(5')ppp, and the correct three size classes of viral polypeptides are not synthesized. These observations may explain how reovirus takes over the host translational apparatus.
Auxin-induced changes in the patterns of protein synthesis in soybean hypocotyl
Authors: Zurfluh Linda L. , Guilfoyle Tom J.
Published: May 2006

The patterns of protein synthesis in elongating and mature (basal) sections of soybean hypocotyl were examined after incubation in a medium containing auxin (auxin-treated) or a medium lacking auxin (untreated). The hypocotyl sections (1.2 cm) were labeled with [
35
S]methionine, and polypeptide patterns were analyzed by one- and two-dimensional polyacrylamide gel electrophoresis. Auxin treatment altered the pattern of protein synthesis in both elongating and basal soybean hypocotyl sections. Excision of terminal segments from incubated sections was required to clearly observe auxin-induced changes in the synthesis of polypeptides. Polypeptides synthesized in terminal segments, possibly in response to wounding, can mask subtle changes in the spectrum of polypeptides synthesized in response to auxin. Cytokinin treatment caused a decrease in [
35
S]methionine incorporation into polypeptides and altered the pattern of protein synthesis in untreated and auxin-treated elongating hypocotyl sections.
Localization of calmodulin in rat tissues.
Authors: Harper J F , Cheung W Y , Wallace R W , Huang H L , Levine S N , Steiner A L
Published: May 2006

The localization of calmodulin, a calcium-dependent modulator of many enzymes, was studied in rat liver, skeletal muscle, and adrenal slices. Calmodulin is found in liver cytoplasm, nucleus, and plasma membrane. Much of the cytoplasmic calmodulin is associated with glycogen particles presumably bound to enzymes involved in glycogen metabolism. Skeletal muscle calmodulin is found on the I-band, also associated with glycogen particles. Intermyofibrillar staining that is not glycogen associated is also observed. Calmodulin is localized in the cytoplasm and nucleus of adrenal cortex cells. Injection of corticotropin leads to a greatly increased localization of calmodulin in nuclei of the adrenal cortex. These observations suggest that one role of calmodulin may be the regulation of hormone effects on nuclear processes.
Target cell specificity of defective avian leukemia viruses: hematopoietic target cells for a given virus type can be infected but not transformed by strains of a different type.
Authors: Graf T , Beug H , Hayman M J
Published: May 2006

Defective avian leukemia viruses of the avian erythroblastosis (AEV), avian myelocytomatosis (MC29), and avian myeloblastosis (AMV) type induce the proliferation of leukemic cells with properties of erythroblasts, macrophages, and myeloblasts, respectively. Their target cells can be separated and have properties of cells of the erythroid (AEV) and myeloid lineage (MC29 and AMV), respectively. In the present study we have shown that this target cell specificity is not due to the ability of the different strains to infect only certain types of hematopoietic cells. Instead, AEV was found to replicate in macrophages and to induce the expression of p75 AEV, its presumptive transforming protein. Likewise, MC29 was found to replicate in AEV-infected erythroblasts as well as in AMV-infected myeloblasts and to express the p110 MC29 protein in these cells. Superinfection with MC29 or AMV of ts34 AEV-infected erythroblasts did not impair their capacity to accumulate hemoglobin after shift to nonpermissive temperature. Our results support a model in which the transforming proteins of AEV, MC29, and MAV block the differentiation of their target cells by competitively inhibiting the action of a hypothetical homologous cellular differentiation protein synthesized in the corresponding target cells only.
Growth and differentiation of embryonal carcinoma cell line F9 in defined media.
Authors: Rizzino A , Crowley C
Published: May 2006

This paper reports the growth and differentiation of the mouse embryonal carcinoma cell line F9 in completely defined culture media. The defined growth medium, referred to as EM-3, contains plasma fibronectin, insulin, and transferrin in place of serum. F9 cells cultured in EM-3 for over 15 generations retain their ability to form tumors and to differentiate. Fibronectin is essential for the attachment of F9 cells in defined media and its effect can be blocked with affinity-purified anti-fibronectin. When retinoic acid was added to EM-3, the F9 cells differentiated. The majority of the the newly formed cells differed from patient F9 cell two major respects: (i) they were morphologically different; and (ii) they secreted plasminogen activator, and the secretion was stimulated by dibutyrlyl adenosine cyclic monophosphate.
Lentropin: a factor in vitreous humor which promotes lens fiber cell differentiation.
Authors: Beebe D C , Feagans D E , Jebens H A
Published: May 2006

An activity has been identified in chicken vitreous humor which stimulates embryonic chicken lens epithelial cells to elongate and specialize for lens crystallin synthesis. The activity is heat-labile and is destroyed by treatment with trypsin or agents that reduce disulfides. Gel filtration and ultrafiltration analyses indicate that it has an apparent molecular weight of approximately 60,000. Its properties differ from those of an activity present in serum which also can promote lens fiber cell formation in vitro. We call this material "lentropin" and suggest that it is responsible for stimulating lens fiber cell formation in vivo and, consequently, plays an important role in determining the shape and polarity of the lens.
Phylogenetic analysis of the mycoplasmas.
Authors: Woese C R , Maniloff J , Zablen L B
Published: May 2006

The phylogenetic relationships between the mycoplasmas and bacteria have been established from a comparative analysis of their 16S rRNA oligonucleotide catalogs. The genera Mycoplasma, Spiroplasma, and Acholeplasma arose by degenerative evolution, as a deep branch of the subline of clostridial ancestry that led to Bacillus and Lactobacillus. Thermoplasma has no specific relationship to the other mycoplasmas; it belongs with the archaebacteria.