bluuebunny/crossref_metadata_2025 huggingface dataset:
Fast axonal transport in auditory neurons of the guinea pig: a rapidly turned-over glycoprotein.
Authors: Tytell M , Gulley R L , Wenthold R J , Lasek R J
Published: May 2006

Proteins of the fast component of axonal transport were analyzed by one- and two-dimensional polyacrylamide gel electrophoresis in the guinea pig spiral ganglion, which has its cell bodies in the cochlea and its axons in the eighth cranial nerve projecting to the ipsilateral cochlear nucleus. We found that we could easily identify the proteins of the fast component even though these axons are only about 3 mm long because the cochlea minimized diffusion of labeled precursor into the cochlear nucleus. The composition of the fast component of the spiral ganglion cells was similar, but not identical, to the fast component of guinea pig retinal ganglion cells. One difference was the predominance in the spiral ganglion cell fast component of a rapidly turned-over glycoprotein (RTGP) with a molecular weight of 110,000-140,000 and an isoelectric point of 5.0 RTGP accumulated in the cochlear nucleus for just the first 3 hr after the application of the labeled precursor and then rapidly disappeared, whereas the other major fast component polypeptides continued to accumulate for 12-24 hr. RTGP was also tentatively identified in the fast component of retinal ganglion cells, but was not as prominently labeled relative to the other fast-component proteins in those cells. The rapid disappearance of RTGP from spiral ganglion cell terminals in the cochlear nucleus may be a result of secretion, perhaps as part of a synaptic vesicle, or retrograde transport as a feedback signal. The difference in the relative amounts of RTGP found in spiral ganglion and retinal ganglion cell terminals may reflect differences in the fundamental properties of the two groups of neurons.
Ternary transition metal phosphides: High-temperature superconductors
Authors: Barz H. , Ku H. C. , Meisner G. P. , Fisk Z. , Matthias B. T.
Published: May 2006

Two systems of ternary transition metal phosphides with the ordered Fe
2
P-type hexagonal structure are reported. They have the general formula MRuP and MOsP, in which M can be Ti, Zr, or Hf. An onset of the superconducting transition temperature as high as 13.0 K is reported for the ZrRuP compound.
Synthesis, cloning, and identification of DNA sequences complementary to mRNAs for alpha and beta subunits of thyrotropin.
Authors: Vamvakopoulos N C , Monahan J J , Kourides I A
Published: May 2006

Double-stranded cDNA sequences were synthesized, by using as templates mRNA for alpha and beta subunits of thyrotropin purified from mouse thyrotrophic pituitary tumours and cloned in Escherichia coli RR1 by insertion in the Pst I site of the bacterial plasmid pBR322 by use of poly(dA) x poly(dT) homopolymeric extensions. Plasmids containing inserted cDNA sequences were selected by resistance to tetracycline and sensitivity to ampicillin; those containing thyrotropin cDNA sequences were identified by colony hybridization with an 125I-labeled mixture of alpha and beta thyrotropin mRNAs. Plasmids carrying either alpha or beta thyrotropin cDNA sequences were further identified by hybridization to highly purified 125I-labeled alpha or beta thyrotropin mRNA, respectively. Two plasmids, one containing a 400-nucleotide alpha thyrotropin cDNA insert and the other containing a 710-nucleotide beta thyrotropin cDNA insert, were purified and characterized by restriction endonuclease digestions. Plasmid [32P]DNA containing either alpha or beta thyrotropin cDNA was then used as a hybridization probe for further characterization of alpha and beta thyrotropin mRNA from the mouse thyrotropic tumor. RNA was fractionated by agarose gel electrophoresis under denaturing and nondenaturing conditions and transferred to diazobenzyloxymethyl-paper. alpha thyrotropin mRNA of two sizes, 650 and approximately equal to 1500 nucleotides long, were identified. The larger alpha thyrotropin mRNA appeared to have marked secondary structure in its native form in contrast to the 650-nucleotide alpha thyrotropin mRNA. However, only one form of beta thyrotropin mRNA was detected with an apparent size of 710 nucleotides. We have successfully cloned and identified alpha and beta thyrotropin cDNA sequences in bacterial plasmids and used them to identify a second form of alpha thyrotropin mRNA.
Sequence of 1060 3'-terminal nucleotides of poliovirus RNA as determined by a modification of the dideoxynucleotide method.
Authors: Kitamura N , Wimmer E
Published: May 2006

The dideoxynucleotide method for sequencing DNA developed by Sanger et al. [Sanger, F., Nicklen, S. & Coulson, A. (1977) Proc. Natl. Acad. Sci. USA 74, 5463-5467] was modified to allow sequence analysis of poliovirus RNA without recourse to cloning. Our method involves reverse transcription of poliovirus RNA followed by cDNA-dependent DNA synthesis in the presence of unlabeled dNTPs and 2',3'-dideoxynucleoside triphosphates, with Escherichia coli DNA polymerase I (Klenow) used to catalyze the reaction. DNA synthesis is primed by 5'-32P-labeled RNase T1- or RNase A-resistant oligonucleotides generated from poliovirus RNA. The sequence of 1060 nucleotides preceding the 3'-terminal poly(A) is presented. Based on the position of termination codons we propose that viral translation terminates at nucleotide -562.
Isolation and preliminary characterization of single amino acid substitution mutants of aspartate carbamoyltransferase.
Authors: Kantrowitz E R , Foote J , Reed H W , Vensel L A
Published: May 2006

In order to isolate functional Escherichia coli aspartate carbamoyltransferase (carbamoylphosphate:L-aspartate carbamoyltransferase, EC2.1.3.2) with single amino acid replacements, a series of pyrB nonsense mutants has been isolated. These nonsense mutants were induced by 2-aminopurine mutagenesis and selected by a combination of antibiotic treatments, direct enzyme assays, and suppressibility tests. Suppression of the pyrB nonsense mutation with various suppressors, which insert different amino acids, has resulted in the formation of a series of mutant aspartate carbamoyltransferases, each differing in one amino acid from the wild-type enzyme. After partial purification, kinetic studies revealed that some of the mutant enzymes had altered homotropic and heterotropic interactions. The mutants that had a tyrosine insert showed the most pronounced changes, followed by those with a serine insert. The mutants having a glutamine insert, howevr, were indistinguishable from the wild-type enzyme, supporting the conclusion that, because of the specificity of the mutagen, the glutamine insert had regenerated the wild-type enzyme.
Four sizes of transcript produced by a single sea urchin gene expressed in early embryos.
Authors: Lee A S , Thomas T L , Lev Z , Britten R J , Davidson E H
Published: May 2006

This report concerns a set of sea urchin egg and embryo transcripts complementary to a single-copy region of a cloned DNA fragment (Sp88). Three distinct 16-cell embryo polysomal RNA species were found to hybridize with this fragment. These RNAs are about 1700, 3000, and 4000 nucleotides (nt) in length, and the same species were identified in unfertilized eggs. A significant fraction of all three species of the egg and early embryo transcripts is polyadenylylated. At gastrula stage Sp88 transcripts are almost completely confined to the nucleus [Lev, Z., Thomas, T. L., Lee, A. S., Angerer, R. C., Britten, R. J. & Davidson, E. H. (1980) Dev. Biol, 75, in press]. The Sp88 transcripts of gastrulae are present as a fourth RNA species approximately 5800 nt in length. The four species share a sequence element of cloned DNA fragment that is about 1000 nt long. These RNAs constitute a set of alternative partially overlapping transcripts from the same genomic region.
Replication of duplex DNA of phage phi X174 reconstituted with purified enzymes.
Authors: Arai K , Arai N , Shlomai J , Kornberg A
Published: May 2006

Replication of the covalently closed duplex replicative form (RF) of phage phi X174 DNA has been achieved by coupling two known enzyme systems: (i) synthesis of viral strand circles (SS) from RF, and (ii) conversion of SS to nearly complete RF (RF II). In this coupled system, activated RF (gene A . RF II complex) was a more efficient template and generated as many as 10 RF II molecules per RF input, at a rate commensurate with SS synthesis. The 11 proteins required for the two component systems were all needed in the coupled RF duplication system; no new factors were required. Single-stranded DNA binding protein was needed for RF duplication at only 4% the level needed in its stoichiometric participation in SS synthesis. In addition to RF II, more complex replicative forms appeared late in the reaction, and their possible origin is discussed.
cis
-4-Cyclohexene-1,2-dicarboximide: Inhibitor of phytochrome-promoted seed germination
Authors: Bewley J. Derek , Oaks Ann
Published: May 2006

cis
-4-Cyclohexene-1,2-dicarboximide (CHDC) inhibits the germination of light-requiring seeds in both light and darkness but has no effect upon the germination of non-light-requiring seeds. In lettuce seeds, CHDC inhibits the action of far-red-absorbing form of phytochrome in breaking dormancy. This inhibition can be overcome by benzyladenine and red light together, but not by a combination of red light and gibberellic acid. Gibberellic acid-induced germination of lettuce seeds in darkness is inhibited also by CHDC. Embryos isolated from dark-imbibed lettuce seeds germinate on the inhibitor. CHDC was thought to be an “analogue” of cycloheximide, but it does not inhibit protein synthesis in lettuce seeds. Our results lead us to conclude that CHDC inhibits germination of seeds that require red light to break dormancy and interferes with some aspect of metabolism that is stimulated by far-red-absorbing form of phytochrome.
Culture of hormone-dependent functional epithelial cells from rat thyroids.
Authors: Ambesi-Impiombato F S , Parks L A , Coon H G
Published: May 2006

Primary cultures of rat thyroid cells were made in medium supplemented with 0.1--0.5% calf serum and containing six hormones or growth factors: insulin, thyrotropin, transferrin, hydrocortisone, somatostatin, and glycyl-L-histidyl-L-lysine acetate. The FRTL strain was purified by successive colonial isolations and was found to maintain highly differentiated features (secretion into the culture medium of physiological amounts of thyroglobulin and concentration of iodide by 100-fold). The FRTL strain has been observed for more than 3 years in continuous culture. It has maintained the same biochemical and morphological characteristics that typified the primary cultures of thyroid follicular cells immediately after their enzymatic release from the rat thyroid. Thyroid epithelial cells that were grown under more conventional cell culture conditions failed to retain these specialized characteristics. We show that maintenance in vitro of these specialized functions of rat thyroid follicular cells is dependent on low serum concentrations and supplementation with hormones in the primary cultures. Our observations indicate that this culture strategem may be aplicable to the general problem of maintenance of differentiated characteristics in cultures of other epithelial cells.
Regulation of the stability of poly(I)xpoly(C)-induced human fibroblast interferon mRNA: selective inactivation of interferon mRNA and lack of involvement of 2',5'-oligo(A) synthetase activation during the shutoff of interferon production.
Authors: Sehgal P B , Gupta S L
Published: May 2006

The inactivation of interferon mRNA during the shutoff phase of interferon production in poly(I)xpoly(C)-induced human fibroblast cultures is selective. We have determined that the shutoff of interferon production, which takes place from 3 to 8 hr after the beginning of induction, is not associated with an appreciable declined in the rate of bulk cellular protein synthesis or of cellular protein secretion. While the amount of translatable interferon mRNA declined markedly during the shutoff phase, the level of translatable bulk cellular mRNA and the stability of [3H]uridine-labeled mRNA were unaffected. Superinduction with actinomycin D selectively stabilized interferon mRNA with no apparent effect on the stability of bulk cellular mRNA. Furthermore, an activation of the 2',5'-oligo(A) synthetase/endonuclease system does not appear to be involved in the shutoff phenomenon. Uninduced FS-4 cells contained a low basal level of 2'5'-oligo(A) synthetase activity, which was unchanged in poly(I)xpoly(C)-induced cells during the shutoff phase. Treatment of FS-4 cells with interferon for 16-18 hr prior to induction increased the enzyme activity by approximately 200-fold. However, this did not inhibit interferon production after induction with poly(I)xpoly(C) alone or after superinduction with cycloheximide or actinomycin D or both. Furthermore, the rates of decay of interferon production were comparable in cells with either a basal or an increased level of 2',5'-oligo(A) synthetase. Thus a 200-fold increase in 2',5'-oligo(A) synthetase level did not affect either the stability of interferon mRNA or the efficacy of interferon superinduction by metabolic inhibitors.
Identification of 5-methylcytosine in DNA fragments immobilized on nitrocellulose paper.
Authors: Sano H , Royer H D , Sager R
Published: May 2006

A method to identify 5-methylcytosine (m5Cyt) in DNA immobilized on nitrocellulose paper by using antibody against m5Cyt raised in rabbits is described. Immobilized restriction fragments of DNA are incubated first with purified antibody against m5Cyt and then with goat anti-rabbit IgG labeled with 125I. Restriction fragments containing m5Cyt are visualized by autoradiography. By using this method, a heavily methylated fragment of about 1700 base pairs was identified in nuclear DNA fom Chinese hamster cells, the methylation pattern of calf thymus satellite I DNA was examined, and chloroplast DNAs that were extracted from various stage of the Chlamydomonas life cycle were compared. Little if any methylation was detected in chloroplast DNA from vegetative cells or from male gametes, whereas homologous DNAs from female gametes and from zygotes were heavily methylated. The sensitivity of the method was examined with calf thymus satellite I DNA (which contains approximately 40 m5Cyt residues per repeat unit of 1400 base pairs) and with phi X174 virion DNA (which contains a single m5Cyt per molecule). The presence of m5Cyt was detected with as little as 40 ng of phi X174 DNA containing 0.02 pmol of m5Cyt and with 100 ng of satellite DNA containing about 0.5 pmol of m5Cyt. Thus, the method makes possible the identification of individual methylated sites in purified DNAs in the size range of single genes.
Studies on the mechanism of skin tumor promotion: Evidence for several stages in promotion
Authors: Slaga T. J. , Fischer S. M. , Nelson K. , Gleason G. L.
Published: May 2006

The effects of nonpromoting and weakly promoting diterpenes on skin tumor promotion by 12-
O
-tetradecanoylphorbol 13-acetate (TPA) were investigated. When phorbol and phorbol 12,13-diacetate (both nonpromoting) were given simultaneously with TPA after 7,12-dimethylbenz[
a
]-anthracene (DMBA) initiation in female mice, they had no effect on TPA promotion. However, the nonpromoter 4-
O
-methyl-TPA and the weak promoter mezerein were found to inhibit TPA promotion in a dose-dependent manner when given simultaneously with TPA. Because mezerein was found to be an effective inhibitor of TPA promotion when given simultaneously and because it induces many biological responses similar to those to TPA, the capacity of mezerein to act as an incomplete promoter in a two-stage promotion protocol was also investigated. Twice-weekly applications of 1,2, or 5 μg of TPA for 2 weeks after DMBA initiation produced 0, 0, and 0.5 papilloma per mouse, respectively, at 20 weeks. When the twice-weekly applications of TPA for 2 weeks were followed by twice-weekly treatments with 2 μg of mezerein for 18 weeks, the number of papillomas per mouse was 2.2, 3.5, and 9.0, respectively. Twice-weekly applications of 2 μg of TPA for 2 weeks followed by twice-weekly treatments with 1, 2, or 4 μg of mezerein for 18 weeks produced 2.1, 3.5, and 6.8 papillomas per mouse, respectively, in DMBA-treated mice. Twice-weekly doses as high as 40 μg of 4-
O
-methyl-TPA were not effective in producing tumors when given after a limited treatment with TPA; however, 4-
O
-methyl-TPA had weak activity as a first-stage promoter. The results suggest that although mezerein by itself is a weak promoter and mimics TPA in many biochemical and morphological effects it is a potent second-stage promoter in a two-stage promotion regimen.
Enkephalin and fear-motivated behavior.
Authors: Rigter H , Jensen R A , Martinez J L , Messing R B , Vasquez B J , Liang K C , McGaugh J L
Published: May 2006

Leu[Enkephalin (400.0 micrograms/kg) and the enkephalin analog [DAla,DLeu]enkephalin (0.4, 4.0, and 40.0 micrograms/kg) were intraperitoneally administered to rats 5 min before they were trained on aversively motivated tasks. The peptides impaired acquisition of a one-way active avoidance response, facilitated acquisition of an inhibitory avoidance response, and had no effect on acquisition of a swim-escape response. The data indicate that the enkephalin effects are not mediated through actions on locomotor activity, pain perception or reactivity, or light sensitivity. Rather, we suggest that [Leu]enkephalin and its analog strengthen the tendency of rats to suppress behavior in the presence of cues previously associated with aversive stimulation. This effect may be due to an enkephalin-induced increase in fear or arousal. The enkephalin effect on behavior was obtained at low doses and was observed within a few minutes after administration. These findings are consistent with the interpretation that enkephalin effects may be initiated at a peripheral site.
Characterization of neurotensin production by a line of rat medullary thyroid carcinoma cells.
Authors: Zeytinoğlu F N , Gagel R F , Tashjian A H , Hammer R A , Leeman S E
Published: May 2006

The rMTC 6-23 cell line was derived from a calcitonin-producing rat medullary thyroid carcinoma. During characterization of these cells we discovered that they also synthesize and secrete neurotensin (NT), a tridecapeptide originally isolated from the hypothalamus and not previously associated with C cells. Immunoreactive NT (iNT) was measured with two antisera: HC-8, which recognizes the COOH-terminal eight amino acids, and TG-1, which binds the NH2-terminal region of NT. By use of these antisera, 2 M acetic acid extracts of rMTC 6-23 cells were found to contain 0.7--5.0 pmol of iNT per mg of cell protein. Successive fractionation of iNT from cell extracts by gel filtration, ion-exchange chromatograpy, and reverse-phase high-pressure liquid chromatography showed at each step a peak of iNT that was indistinguishable from synthetic NT in its chromatographic behavior. Biologic activity of the iNT was confirmed by demonstrating the characteristic fall in arterial blood pressure in the rat after intravenous injection of material purified from rMTC 6-23 cells. Calcium (0.5--4.0 mM) stimulated release of iNT in a dose-dependent manner; the effect was maximal at 3--4 mM calcium. K+ (50 mM) stimulated release of iNT that was maximal in the presence of 1.0--1.5 mM calcium. Synthesis and secretion of iNT by these cells were shown during a 16-day growth experiment. These results demonstrate that rMTC 6-23 cells contain NT and suggest the possibility of an association between neurotensin and calcitonin.
Small phospholipid vesicles: internal pressure, surface tension, and surface free energy.
Authors: White S H
Published: May 2006

Tanford [Tanford, C. (1979) Proc. Natl. Acad. Sci. USA 76, 3318-3319] used thermodynamic arguments to show that the pressure difference across the bilayer of small phospholipid vesicles must be zero. This paper analyzes the implications of this conclusion in terms of Laplace's law and the basic thermodynamics of interfaces. In its usual form, Laplace's law is of questionable value for the vesicle. If the vesicle is in a state of metastable equilibrium, the surface free energy must be minimal with respect to several thermodynamic variables; the condition (delta F/ delta A) = 0 is not adequate by itself.
Analysis of the nucleotide sequence of an invertible controlling element.
Authors: Zieg J , Simon M
Published: May 2006

The nucleotide sequence of the inversion region responsible for flagellar phase variation in Salmonella was determined. The inversion region is 995 base pairs (bp) in length and is bounded by a 14-bp inverted repeat sequence. A homologous recombination event between the 14-bp inverted repeat sequences would result in the inversion of the DNA segment between them. Sequence homologies with other systems suggest that the 14-bp inverted repeat sequences may have some general significance as sites for specific recombinational events. The gene which specifies H2 flagellin synthesis begins 16 bp outsie the inversion region. Within the inversion region, an open translational frame exists which could encode a low molecualr weight polypeptide (190 amino acids).
Sickle-cell hemoglobin: fall in osmotic pressure upon deoxygenation.
Authors: Hargens A R , Bowie L J , Lent D , Carreathers S , Peters R M , Hammel H T , Scholander P F
Published: May 2006

Macromolecules such as hemoglobin exert both kinetic and matrix effects on osmotic pressure. The kinetic osmotic pressure of sickle-cell hemoglobin is lost upon deoxygenation at physiological erythrocyte concentrations. The non-kinetic or matrix component of osmotic pressure remains relatively unchanged. Loss of thermal-osmotic activity during deoxygenation occurs throughout a hemoglobin concentration range between 2.5 and 35 g/100 ml. Deoxygenation of sickle-cell hemoglobin causes aggregation such that the matrix effect is unchanged but the kinetic (van't Hoff) effect nearly vanishes. A loss of intracellular osmotic pressure during deoxygenation could dehydrate the erythrocyte sufficiently to promote more rapid sickle-cell hemoglobin aggregation. Subsequently, complete gelation of these aggregates could cause additional water loss and thrust the sickled cell into an irreversible cycle. The osmotic pressure of normal hemoglobin does not change appreciably during deoxygenation and is essentially the same as the osmotic pressure of oxygenated sickle-cell hemoglobin.
Translation of epigenetic rules of individual behavior into ethnographic patterns
Authors: Lumsden Charles J. , Wilson Edward O.
Published: May 2006

The pivotal process in gene-culture coevolution is envisaged to be the evolution of behavioral epigenesis. From premises based on the known properties of enculturation and usage diffusion within societies, a probabilistic model is constructed to estimate the degree to which rules governing individual development canalize ethnographic curves (the probability density distributions of societies engaged in varying patterns of usage). The results indicate that under most conceivable conditions the translation from individual epigenesis to social pattern is amplified, to an extent that differences in bias too faint to be detected in ordinary developmental studies can generate conspicuous variation in the ethnographic curves. Examples are cited of sufficiently biased epigenesis in human behavior.
Bäcklund transformations for multidimensional sine-Gordon equations
Authors: Benguria R. , Case K. M.
Published: May 2006

A class of Bäcklund transformations for multidimensional sine-Gordon equations is shown to relate solutions of only one- and two-dimensional equations. Also, a proposed superposition principle is shown to be useless.
Boundary methods: A criterion for completeness
Authors: Herrera Ismael
Published: May 2006

The application of methods which constitute an alternative to boundary integral equations to specific problems depends on development of complete systems of solutions, convergence of approximating procedures, and formulation of variational principles. This paper establishes a criterion for completeness. In this manner, greater flexibility of the theory is achieved; for example, systems of functions which are complete for different types of boundary conditions are developed.
Direct inhibition of testicular function by gonadotropin-releasing hormone: mediation by specific gonadotropin-releasing hormone receptors in interstitial cells.
Authors: Clayton R N , Katikineni M , Chan V , Dufau M L , Catt K J
Published: May 2006

Agonist analogs of gonadotropin-releasing hormone (GnRH) have been shown to exert antigonadal effects in male and female animals. In hypophysectomized male rats treated with follicle-stimulating hormone, administration of a potent GnRH agonist caused depletion of luteinizing hormone and prolactin receptors and marked suppression of serum testosterone levels. The possibility that such direct effects of GnRH agonists on testicular function could be expressed through specific receptors located in the interstitial cells of the testis was supported by the selective concentration of a 125I-labeled GnRH agonist by the testis in vivo. Specific receptors for the releasing hormone were demonstrated in testis particles and dispersed interstitial cells by direct binding analysis with the 125I-labeled GnRH agonist. The binding affinity (Ka = G X 10(9) M-1) and peptide specificity of the testicular GnRH binding sites were similar to those of anterior pituitary and ovarian GnRH receptors. The presence of GnRH receptors in the testis indicates that these sites mediate the direct inhibitory actions of GnRH agonists upon testicular endocrine function.
Molecular cloning of developmentally regulated, low-abundance mRNA sequences from embryonic muscle.
Authors: Ordahl C P , Kioussis D , Tilghman S M , Ovitt C E , Fornwald J
Published: May 2006

To study the role of low-abundance, embryonic muscle-specific gene transcripts, we have developed a method to screen cDNA clones from embryonic muscle for such sequences. The protocol involves two stages: first, partial enrichment for cDNA clones carrying possible embryo-specific sequences by selecting clones of low-abundance sequences; and second, determination, by hybridization to RNA attached to diazobenzyloxymethyl-paper, which sequences from this category are regulated in an embryonic muscle-specific manner during development. At least three different clones were obtained which hybridized to sequences present in early muscle development but absent, or present at relatively low levels, at late embryonic and adult muscle stages. Two of these clones were not muscle-specific because they hybridized to poly(A)+RNA from liver or brain or both. The third clone, 106A4, did not detectably hybridize to total poly(A)+RNA at any stage of brain or liver development tested. This sequence also was not detectable in poly(A)+RNA from embryonic muscle progenitor cells. Thus, the 106A4 sequence is a likely candidate for an embryonic muscle-specific sequence. We have demonstrated that the 106A4 sequence is a mRNA, although the specific identity and function of the translated product is unknown. The method used to identify embryonic muscle-specific cDNA clones should be generally applicable for obtaining clones for low abundance transcripts regulated in a tissue-specific or developmental stage-specific manner.
Intermediate filaments of baby hamster kidney (BHK-21) cells and bovine epidermal keratinocytes have similar ultrastructures and subunit domain structures.
Authors: Steinert P M , Idler W W , Goldman R D
Published: May 2006

Structural features of the intermediate filaments (IF) of bovine epidermal keratinocytes (keratin filaments) and of baby hamster kidney (BHK-21) cells were studied to define the molecular basis of the similarities and differences in their properties. Purified subunits of the IF were specifically cleaved at their methionine and tryptophan residues. Peptide maps indicated that the IF subunits of BHK-21 cells were similar but not identical to each other and quite different from each of the keratin IF subunits. Intact IF were subjected to limited tryptic digestion, and structurally identical alpha-helix-enriched particles were released. Their properties indicated these IF were composed of a similar three-chain unit which contained regions of coiled-coil alpha-helix interspersed with region of non-alpha-helix. These two types of experiments permitted the construction of subunit domain maps which revealed a common structure: all subunits possessed two alpha-helical domains of the same size that were adjoined by non-alpha-helical domains of variable size. We propose that the reported solubility and immunological differences in these IF and perhaps those of other types of cells are due largely to variations in the size, configuration, and amino acid sequence of the non-alpha-helical regions of the subunits in the IF.
Apurinic/apyrimidinic endonucleases in repair of pyrimidine dimers and other lesions in DNA.
Authors: Warner H R , Demple B F , Deutsch W A , Kane C M , Linn S
Published: May 2006

The characteristics of the nicks (single-strand breaks) introduced into damaged DNA by Escherichia coli endonucleases III, IV, and VI and by phage T4 UV endonuclease have been investigated with E. coli DNA polymerase I (DNA nucleotidyltransferase). Nicks introduced into depurinated DNA by endonuclease IV or VI provide good primer termini for the polymerase, whereas nicks introduced into depurinated DNA by endonuclease III or into irradiated DNA by T4 UV endonuclease do not. This result suggests that endonuclease IV nicks depurinated DNA on the 5' side of the apurinic site, as does endonuclease VI, whereas endonuclease III has a different incision mechanism. T4 UV endonuclease also possesses apurinic endonuclease activity that generates nicks in depurinated DNA with low priming activity for the polymerase. The priming activity of DNA nicked with endonuclease III or T4 UV endonuclease can be enhanced by an additional incubation with endonuclease VI and, to a lesser extent, by incubation with endonuclease IV. These results indicate that endonuclease III and T4 UV endonuclease (acting upon depurinated and irradiated DNA, respectively) generate nicks containing apurinic/apyrimidinic sites at their 3' termini and that such sites are not rapidly excised by the 3' leads to 5' activity of DNA polymerase I. However, endonuclease IV or VI apparently can remove such terminal apurinic/apyrimidinic sites as well as cleave on the 5' side of the unnicked sites. These results suggest roles for endonucleases III, IV, and VI in the repair of apurinic/apyrimidinic sites as well as pyrimidine dimer sites in DNA. Our results with T4 UV endonuclease suggest that the incision of irradiated DNA by T4 UV endonuclease involves both cleavage of the glycosylic bond at the 5' half of the pyrimidine dimer and cleavage of the phosphodiester bond originally linking the two nucleotides of the dimer. They also imply that the glycosylic bond is cleaved before the phosphodiester bond.
Electron microscope study of duck globin mRNA precursor crosslinked in situ.
Authors: Reymond C , Aglianó A , Spohr G
Published: May 2006

Double-stranded RNA segments present in duck globin pre-mRNA were crosslinked in situ with aminomethyltrioxalen and UV light. The secondary structure of the crosslinked pre-mRNA was then studied by electron-microscopic analysis of pre-mRNA . cDNA hybrids. The data suggest that duck globin pre-mRNAs contain intervening sequences that are excised stepwise. Excision and subsequent ligation appears to occur on precursor molecules that are stabilized by base-paired regions.
Nerve growth factor receptors on PC12 cells: ligand-induced conversion from low- to high-affinity states.
Authors: Landreth G E , Shooter E M
Published: May 2006

The pheochromocytoma PC12 cell possesses specific cell surface receptors that bind nerve growth factor (NGF) with two different affinities. The rate of dissociation of NGF from the higher affinity receptor is slower than from the lower affinity receptor; this rate is reduced to essentially zero at low temperature, allowing the extent of high-affinity binding to be determined. When NGF is added to PC12 cells, only low-affinity binding is observed. After a short lag period, high-affinity binding also appears and increases slowly. If NGF is removed from the medium after binding is initiated, high-affinity receptors continue to be formed at the expense of low-affinity receptors. The increase in receptor affinity is accompanied by a transfer of the NGF-receptor complex from a trypsin-sensitive to a trypsin-resistant state. This transfer does not involve internalization of the NGF. The data show that NGF binds first to receptors of low affinity and that the binding induces a conversion of a proportion of the receptors to a higher affinity state. It is also consistent with a model in which the change in affinity is due either to conformational changes in the receptor or to interaction of the occupied receptor with other receptors or with effector proteins in cell plasma membrane.
Talpid2 mutant of the chicken with perturbed cartilage development has an altered precartilage-specific chromatin protein.
Authors: Perle M A , Newman S A
Published: May 2006

An abundant nonhistone protein (Mr, 125,000) is lost from the chromatin embryonic chicken precartilage mesenchyme cells as they differentiate into cartilage [Newman, SA., Birnbaum, J & Yeoh, G.C.T.(1976) Nature (London) 259, 417-418]. We have now examined the chromatin proteins of precartilage and cartilage cells of chicken embryos carrying the talpid2 gene which causes a perturbed patern of cartilage differentiation in the homozygous state. We find that homozygous talpid2 precartilage chromatin differs from that of the normal cell type in having its abundant precartilage chromatin protein decreased to a Mr of approximately 120,000 and in having a "precocious" cartilage-like pattern of proteins in the Mr 35,500-36,500 region. The precartilage chromatin of talpid2 heterozygotes is completely normal within the resolution of our techniques, as is the cartilage chromatin of the homozygote and heterozygote talpid2 embryos. The correlation of an aberration in a developmentally significant chromatin protein with the perturbed development of its tissue of origin is discussed.
Control of gene expression by a mobile recombinational switch.
Authors: Berg D E
Published: May 2006

Transposable recombinational switches may play important roles in the evolution of bacterial populations by increasing flexibility in the control of expression of particular genes and thereby maintaining heterogeneity in clones of cells growing in a uniform environment. Experiments reported here show that Tn5-112, a deletion derivative of kanamycin-resistance transposon Tn5, can function as such a mobile recombinational switch. The internal deletion in Tn5-112 removes transcription termination signals and permits transcription initiated within the element to continue into nearby bacterial genes. Consequently, in one orientation Tn5-112 stimulates distal gene expression, whereas in the other orientation the normal polarity imposed by wild-type Tn5 intervenes and distal gene expression is not stimulated. Because Tn5-112 contains terminal inverted repeats, intramolecular recombination can invert the Tn5-112 element and alter gene expression. Tn5-112 is transposition deficient. Its mobility derives from the recessive nature of the transposition deficiency and, in this study, from the possibility of homologous recombination which permits its placement in either orientation at any site occupied by nother Tn5 element.
A transformation-defective mutant of Abelson murine leukemia virus lacks protein kinase activity.
Authors: Witte O N , Goff S , Rosenberg N , Baltimore D
Published: May 2006

A transformation-defective mutant of Abelson murine leukemia virus (A-MuLV), called A-MuLV-P92td, has been isolated. The mutant encodes a serologically identifiable A-MuLV protein of molecular weight 92,000 (P92) but it lacks the ability to transform either fibroblasts or bone marrow lymphoid cells. In contrast to the protein made by transforming strains of A-MuLV, the protein made by A-MuLV-P92td does not becme phosphorylated during in vitro incubation with [gamma-32P]ATP. If the protein is mixed with proteins from cells transformed by a functional A-MuLV strain, phosphorylation of P92 occurs, showing that its ability to accept phosphate is not altered by the mutation. These parallel changes provide genetic evidence that the A-MuLV protein is a transforming protein and that its associated protein kinase activity (EC 2.7.1.37) is a crucial part of its transforming ability.
Retinal rhythms in chicks: circadian variation in melantonin and serotonin N-acetyltransferase activity.
Authors: Hamm H E , Menaker M
Published: May 2006

There is a large-amplitude circadian rhythm of indoleamine metabolism in the retina-pigment epithelium of the chicken. N-Acetyltransferase activity (arylamine acetyltransferase; acetyl-CoA:arylamine N-acetyltransferase, EC 2.3.1.5) and melatonin content are 15-fold higher at night than during the day in a cycle of a 4-fold increase during the subjective night. Light at midnight inactivates N-acetyltransferase and lowers melatonin. N-Acetyltransferase activity is found predominantly in the retina. The circadian rhythm of this enzyme activity persists in pinealectomized chicks. Thus the pineal is not responsible for retinal indoleamine rhythms. Retinal and pineal levels of N-acetyltransferase activity behave similarly under several conditions. In the chicken, the eye is a major site of rhythmic indoleamine metabolic activity.
A remark on the Conway-Norton conjecture about the “Monster” simple group
Authors: Kac Victor G.
Published: May 2006

By the method of second quantization we construct an infinite-dimensional graded module for the centralizer of an involution of the Fischer-Griess “Monster” simple group
F
1
so that the Thompson series are those conjectured by Conway and Norton. As a consequence we obtain some identities and congruences relating modular functions
j
and η. We are hopeful that this representation can be extended to the whole group
F
1
, proving all the conjectures by Conway and Norton in their recent paper [Conway, J. H. & Norton, S. P. (1979)
Bull. London Math. Soc.
11, 308-339].
Spectral theory for domains in R

n

of finite measure
Authors: Jørgensen Palle E. T.
Published: May 2006

Let Ω be a measurable subset of R

n

of finite positive Lebesgue measures. The following two problems are considered: (
i
) Find commuting self-adjoint extensions of the minimal operators -
i
∂
/∂

x
k
, k

= 1,...,
n
(Ω open). (
ii
) Find a set Λ ⊂ R

n

such that the functions
e
λ
= exp(
i
λ
1
x
1
+... +
i
λ

n

x
n

) for λ ∈ Λ form an orthonormal basis for
L
2
(Ω). The problems are known to be equivalent under mild regularity conditions on Ω, and existence holds in two cases: (
i
) there is a connected open set Ω′ such that the symmetric difference ΩΔΩ′ is a null set and Ω′ is a fundamental domain for a discrete total subgroup; and (
ii
)Ω = [unk]

a∈R

(
a
+ [unk]), disjoint union neglecting null sets, in which [unk] is a fundamental domain and
R
is a “set of representors” for a finite group of translations. Case
i
is equivalent to a function theoretic condition of Forelli, and case
ii
is established when the existence of a discrete covariance group is assumed. Generalizations of the geometric results
i
and
ii
for spectral sets in arbitrary Lie groups are indicated.
Mechanism of translocation: relative arrangement of tRNA and mRNA on the ribosome.
Authors: Matzke A J , Barta A , Kuechler E
Published: May 2006

AcPhe-tRNAPhe from yeast can be photocross-linked to poly(U) on Escherichia coli ribosomes. The photoreaction occurs at the wybutine base situated next to the 3' side of the anticodon. The kinetics and efficiency of crosslinking of AcPhe-Phe-tRNA are the same at both the acceptor site and the peptidyl site. Therefore, the orientation of wybutine with respect to the mRNA is similar in both the pretranslocational and posttranslocational states. AcPhe-Phe-tRNA crosslinked at the acceptor site can still be translocated to the peptidyl site, demonstrating that tRNA and mRNA are transported together. The experiments support a model of translocation in which the conformation of the anticodon loop of tRNA is similar in both the peptidyl site and the acceptor site.
Selective effects of phorbol 12-myristate 13-acetate on myofibrils and 10-nm filaments.
Authors: Croop J , Toyama Y , Dlugosz A A , Holtzer H
Published: May 2006

Phorbol 12-myristate 13-acetate (PMA) has a prompt and selective catabolic effect on striated myofibrils in postmitotic myotubes. Fluorescein-labeled antibodies against light meromyosin were used to follow the effects of PMA on the muscle-specific myosin in myofibrils. The response of actin filaments was monitored by decoration with heavy meromyosin. The response of the two types of 10-nm filaments in myotubes was followed by fluorescein- and rhodamine-labeled antibodies to the fibroblastic and muscle-specific filament proteins, respectively. Within 2-3 days, PMA induced dismantling of virtually every striated myofibril in every myotube in the culture. These myotubes bound little or no anti-light meromyosin, and tests to detect the alpha-actin filaments of the myofibrils with heavy meromyosin were negative. In contrast, the nonmuscle actin in the subsarcolemmal microfilaments persisted in PMA-treated myotubes and was decorated with heavy meromyosin. The sarcoplasmic reticulum, mitochondria, and Golgi bodies appeared normal. Myotubes depleted of myofibrils by PMA displayed large numbers of muscle-specific 10-nm filaments. This preferential degradation of the myosin and actin of the myofibrils were reversible. These myotubes formed a normal complement of myofibrils 24-48 hr after removal of PMA. When, after 3 days in PMA, the cultures were treated for an additional 3-8 days, a transitory subpopulation of PMA-resistant myotubes appeared.
Preferential inclusion of extrachromosomal genetic elements in yeast meiotic spores.
Authors: Brewer B J , Fangman W L
Published: May 2006

During meiosis and sporulation in the yeast Saccharomyces cerevisiae, extrachromosomal traits are efficiently transmitted to haploid spores. Although the pattern of inheritance of chromosomal traits reflects the mechanism of regular chromosomal segregation in meiosis, it is not known what processes are reflected by the efficient inheritance of extrachromosomal traits. Because extrachromosomal genetic elements in yeast are present in multiple copies, perpetuation of an extrachromosomal trait could occur by the passive envelopment of a subset of copies or by an active sequestering of all or a subset of copies within the four spores. We show that only subsets of the four extrachromosomal nucleic acids commonly found in yeast are transmitted through meiosis--55% of mitochondrial DNA copies, 82% of the 2-micron DNA plasmids, and about 70% of the L and M double-stranded RNAs. However, electron micrographs of serial sections through yeast asci indicate that the four spore enclose only 30% of the total ascus material. Thus these extrachromosomal elements are preferentially included within the spores, indicating that their inheritance is not a random process. Transmission of mitochondrial DNA can be accounted for by the observed enclosure of 52% of the mitochondrial volume within the spores. The high transmission frequencies of the double-stranded RNAs (which exist as virus-like particles in the cytoplasm) and 2-micron DNA must indicate that either these nucleic acids are actively recruited from the cytoplasm by some mechanism or they are associated in some way with the nucleus during meiosis.
Selective killing of normal or neoplastic B cells by antibodies coupled to the A chain of ricin.
Authors: Krolick K A , Villemez C , Isakson P , Uhr J W , Vitetta E S
Published: May 2006

Highly specific antibodies (affinity-purified or hybridoma) directed against cell surface immunoglobulins on normal or neoplastic murine B lymphocytes were covalently coupled to the A chain of the plant toxin ricin. Such conjugates containing antibodies specific for IgM, for either of the two allotypes of IgD, or for the idiotype of the B cell tumor BCL1 rapidly bound in vitro to cells expressing the corresponding surface antigen and inhibited protein synthesis in such cells. The results demonstrate that A chain-coupled anti-idiotype antibody may be useful as a tumor-specific cytotoxic agent.
Low density lipoprotein receptor activity in human monocyte-derived macrophages and its relation to atheromatous lesions.
Authors: Traber M G , Kayden H J
Published: May 2006

Human peripheral monocytes, isolated from a previously unused source (a leukocyte concentrate byproduct of the plateletphoresis procedure for platelet transfusion), transformed into macrophages while cultured with 5% human serum or isolated lipoprotein fractions. Used for the study of their cholesterol (Chol) metabolism, these human monocyte-derived macrophages had a high-affinity receptor that is saturable, specific for low density lipoprotein (LDL) and maximally induced by incubation for as little as 24 hr in medium devoid of lipoproteins or Chol. Macrophages were shown to have separate receptor activity for 125I-labeled LDL and 125I-labeled acetylated LDL; macrophages that had been incubated with lipoprotein-depleted serum degraded native and acetylated LDL at similar rates. The receptor for LDL was functional in maintaining Chol homeostasis, as demonstrated by feedback inhibition of Chol synthesis after culture with LDL or very low density lipoprotein (VLDL). The Chol content of macrophages was doubled by incubation with medium containing VLDL compared to incubation with LDL (60 microgram of Chol per ml of medium). Incubation with native lipoproteins did not lead to accumulation of esterified Chol by macrophages. Changes in the Chol metabolism of macrophages, rather than modifications in circulating LDL, may cause these cells to store cholesteryl ester and take on the characteristics of cholesteryl ester-laden macrophages of atheromatous lesions.
Anti-Thy 1.2 monoclonal antibody linked to ricin is a potent cell-type-specific toxin.
Authors: Youle R J , Neville D M
Published: May 2006

The cell-type specificity of the toxin ricin, which ordinarily binds, enters, and kills cells through receptors containing galactose, has been altered by covalently binding it to a monoclonal antibody and by reversibly binding it to lactose. The antibody, a monoclonal rat IgG2b directed against the Thy 1.2 antigen, provides ricin with a new binding site for the murine thymus cell surface. Addition of lactose saturates the galactose-binding site on ricin and inhibits ricin from binding and killing cells via the galactose-containing receptors. The antibody-ricin hybrid protein, anti-Thy 1.2-ricin, formed with a thioether linkage, has been purified by size exclusion and affinity chromatography. When assayed by inhibition of protein synthesis of EL-4 cells, which express the Thy 1.2 antigen anti-Thy 1.2-ricin is equally as toxic as ricin on a molar basis. The hybrid protein toxicity is unchanged in the presence of 100 mM lactose, whereas unmodified ricin toxicity is reduced to 1% of its toxicity in the absence of lactose. This demonstrates the altered receptor specificity of the ricin hybrid. The cell-type specificity of the anti-Thy 1.2-ricin inhibition of protein synthesis correlates with the presence of the Thy 1.2 antigen. Anti-Thy 1.2-ricin at 4 microgram/ml in the presence of lactose inhibits protein synthesis within 3.5 hr by 60-80% in EL-4 cells but does not affect Thy 1.1 alloantigen and HeLa cells that lack the Thy 1 antigen. Anti-Thy 1.2-ricin in the presence of lactose selectively kills EL-4 cells at concentrations that do not kill AKR-K36 cells. This selectivity, expressed as the ratio of anti-Thy 1.2-ricin concentrations required to kill 40% of both cell types, is 700. Ricin-monoclonal antibody hybrids of this type combine a high degree of cell-type selectivity and toxicity and may have pharmacologic utility as antitumor reagents.