bluuebunny/crossref_metadata_2025_split huggingface dataset:
Cellular signal transduction and the reversal of malignancy
Authors: Lockwood Arthur H. , Murphy Suzanne K. , Borislow Steven , Lazarus Adam , Pendergast Maryanne 
Published: December 2004

AbstractAnimal cells contain only a few defined molecular systems that transduce hormonal and growth signals from the external environment to the intracellular milieu to regulate cellular growth and differentiation. Among the most ubiquitous of these “second messenger” pathways are those utilizing cyclic AMP and phosphatidylinositide turnover. The former activates protein kinase A, while the latter leads to the activation of protein kinase C and mobilization of intracellular calcium. Lesions induced by oncogenes in signal transduction systems may be responsible for the cancerous transformation of cells. In many tumor cell lines, including some transformed by the ras and sis oncogenes, activation of protein kinase A by elevation of cyclic AMP or activation of protein kinase C by addition of phorbol esters can restore many normal aspects of growth and morphology. Such “reverse transformation” is accompanied by the phosphorylation of unique cellular proteins and alterations in the phosphoinositide cycle. Molecular mechanisms by which activation of signal transduction systems can attenuate the malignant phenotype are considered in the context of cellular growth and differentiation.
Estradiol‐dependent trans‐acting factor binds preferentially to a dyad‐symmetry structure within the third intron of the avian vitellogenin gene
Authors: Jost Jean‐Pierre , Saluz Hanspeter , Jiricny Josef , Moncharmont Bruno 
Published: December 2004

AbstractThe secondary activation of the avian vitellogenin II gene in isolated liver nuclei by cytoplasmatic liver extracts of estradiol‐treated chicks is accompanied by the binding of a protein from the extract to the structural part of the cloned gene. Both the DNA‐binding and gene‐stimulatory activities, which cochromatograph on heparin‐Sepharose, are apparently present only in the cytoplasmatic liver extracts of estradiol‐treated roosters and in the oviduct extracts of egg‐laying hens. DNA‐binding competition assays combined with exonuclease III footprinting showed that the factor binds to the imperfect dyad‐symmetry structure 5‘GTCTTGTTCCAAAC3’ within the third intron of the gene. The factor is sequence specific and binds equally well to both single‐ and double‐stranded DNA with an estimated dissociation constant of 3.5 × 10−10M.
Isolation of a calcium‐sensitive, 35,000‐dalton microfilament‐ and liposome‐binding protein from ascites tumor cell microvilli: Identification as monomeric calpactin
Authors: Liu Yuecheung , Brew Keith , Carraway Kermit L. , Carraway Coralie A. Carothers 
Published: December 2004

AbstractMicrovilli isolated from the MAT‐C1 ascites subline of the 13762 rat mammary adenocarcinoma contain a major calcium‐sensitive microfilament‐binding protein, AMV‐p35 (ascites microvillar p35). Association of AMV‐p35 with microfilament cores during Triton X‐100 extraction of the microvilli is half‐maximal at 0.1‐0.2 mM calcium. The protein, which comprises 6% of the total microvillar protein, can be isolated from microfilament cores prepared in the presence of calcium by extraction with EGTA and purification by ion‐exchange chromatography. Alternatively, the protein can be isolated from Triton extracts of microvilli prepared in the absence of calcium by precipitation with calcium, solubilization of the precipitate with EGTA, and chromatography on an ion‐exchange column. AMV‐p35 binds to phosphatidylserine liposomes and F‐actin with half‐maximal calcium concentrations of about 10 μM and 0.2 mM, respectively. Treatment of AMV‐p35 with chymotrypsin yields a 33,000‐dalton fragment, behavior similar to the tyrosine kinase substrates calpactins I and II and lipocortins I and II. Immunoblot analyses using antibodies directed against calpactin I, lipocortin I, and lipocortin II showed strong reactivity of AMV‐p35 with anti‐calpactin I and anti‐lipocortin II, but little reactivity toward anti‐lipocortin I. The close relationship between AMV‐p35 and calpactin I was verified by amino acid sequence analyses of peptides isolated from cyanogen bromide digests of AMV‐p35. By gel filtration and velocity sedimentation analyses purified AMV‐p35 is a 35,000‐dalton monomer. Moreover, AMV‐p35 extracted directly from microvilli in Triton/EGTA also behaves as a 35,000‐dalton menomer. These findings indicate that AMV‐p35 is closely related to the pp60src kinase substrate calpactin I (p36). However, AMV‐p35 occurs in the microvilli as a monomer rather than as the heterotetrameric calpactin found in several other cell types.
Hormone‐dependent processing of the avian progesterone receptor
Authors: Sullivan William P. , Smith David F. , Beito Thomas G. , Krco Christopher J. , Toft David O. 
Published: December 2004

AbstractAvian progesterone receptor exists as two forms, A and B, with molecular weights of 79,000 and 110,000 daltons, respectively. The origin and significance of these two forms is an area of active investigation and debate. Monoclonal antibodies produced against these two forms were used to examine receptor stability in cytosol and changes in the receptor forms induced by hormone binding.The lability of hormone binding at elevated temperatures is well documented. Analysis by Western blotting showed the receptor was stable in freshly prepared oviduct cytosol for 2 hr at 37°C, while hormone binding was lost within 30 min. However, loss of receptor through degradation was seen when cytosol was prepared from frozen tissue or when homogenization was excessive.Progesterone was injected into diethylstilbestrol‐stimulated chicks to examine, in vivo, effects of hormone treatment on receptor forms in the cytosol and nuclear fractions. Progesterone treatment caused a time‐ and dose‐dependent conversion of the A receptor to a form (A′) with a slower electrophoretic mobility. The cytosolic progesterone receptor was divided equally between the B and A forms, while the nuclear receptor was predominantly A′. The amount of nuclear receptor was consistently less than cytosolic receptor. Receptor phosphorylation was analyzed by incubating tissue minces with [32P]orthophosphate with or without progesterone followed by immune isolation of receptor forms. Progesterone treatment caused a time‐dependent increase in cytosol receptor phosphorylation which was evident after 5 min of treatment. This phosphorylation was observed with both the A and B receptor forms. The results indicate that receptor phosphorylation is a very early event during progesterone action.
Swiss 3T3 mouse embryo fibroblasts transfected with a human prepro‐GRP gene synthesize and secrete pro‐GRP rather than GRP
Authors: Lebacq‐Verheyden Anne‐Marie , Segal Shoshana , Cuttitta Frank , Battey James F. 
Published: December 2004

AbstractA prepro‐gastrin‐releasing peptide (GRP) gene was introduced into Swiss 3T3 mouse embryo fibroblasts by DNA transfection in an attempt to establish autocrine growth stimulation. Clonal transfectants expressed varying amounts of GRP encoding mRNA. They synthesized and secreted a ∼ 15‐kd pro‐GRP hormone but not fully processed 2.8‐kd GRP. Accordingly, no changes in growth properties were associated with GRP gene expression. We postulate that Swiss 3T3 fibroblasts lack the enzymes necessary to process significantly pro‐GRP into biologically active peptides and that this deficiency may be responsible for the failure to establish autocrine growth stimulation in the transfected cells.
Effect of mutations affecting Na+:H+ antiport activity on tumorigenic potential of hamster lung fibroblasts
Authors: Lagarde Alain E. , Franchi Arlette J. , Paris Sonia , Pouysségur Jacques M. 
Published: December 2004

AbstractMutants unable to regulate intracellular pH through the Na+:H+ antiport system were found to evolve tumors less frequently than wild‐type CCL39 hamster lung fibroblasts, after transplantation in athymic nude mice. When rare tumors arose, they comprised cells which were transformed in vitro, but which upon retransplantation grew at a lower rate than tumor cells originating from CCL39 cells. Both parental and mutant cells became transformed after transfection of the activated Harvey ras oncogene, but transfectants derived from the mutants had a weaker tumorigenic potential. These results suggest that transformed characteristics can be acquired independently from the Na+:H+ antiporter. However, the presence of this system provides a selective growth advantage when cells are confronted with natural environments, as it occurs during the expansion of tumors in a host.
ADP‐ATP Carrier of Saccharomyces cerevisiae contains a mitochondrial import signal between amino acids 72 and 111
Authors: Smagula Cynthia S. , Douglas Michael G. 
Published: December 2004

AbstractThe ADP‐ATP carrier (also referred to as the adenine nucleotide translocator) of Saccharomyces cerevisiae is encoded by a nuclear gene, translated in the cytosol, and imported into the mitochondrial inner membrane. In order to study the determinants of mitochondrial import, a series of fusion proteins, consisting of the first 21, 72, and 111 amino acids of the ADP‐ATP carrier, joined to mouse dihydrofolate reductase were generated. Dihydrofolate reductase is a cytosolic protein that does not bind mitochondria. The reticulocyte lysate reaction containing the 35S‐methionine‐labeled protein was incubated with mitochondria in a buffer containing 3% BSA. Following incubation for import, the reactions were treated with 1 mM PMSF or 25 μg/ml proteinase K; mitochondria were reisolated and analyzed by gel electrophoresis. The 21 and 72 amino acid hybrid proteins showed a low level of binding to mitochondria: the bound form was entirely protease accessible. The 111 amino acid hybrid protein was imported to a protease‐protected location within mitochondria. It is concluded that the first 72 amino acids of the ADP‐ATP carrier do not suffice to import the protein into mitochondria and that the region between amino acids 72 and 111, a region that contains a transmembrane‐spanning domain, constitutes at least part of the mitochondrial import signal.
Monoclonal antibodies to carbohydrate antigens in autologous bone marrow transplantation
Authors: Ball Edward D. , Howell Alexandra L. 
Published: December 2004

AbstractNormal and malignant myeloid cells express a highly immunogenic oligosaccharide, lacto‐n‐fucopentaose‐III (LNF‐III), that has been identified by numerous monoclonal antibodies (MoAb). We have been interested in the use of a particular monoclonal antibody to LNF‐III, PM‐81, in the treatment of patients with acute myelogenous leukemia using the antibody to treat bone marrow in vitro. Following in vitro treatment of bone marrow with PM‐81 and another MoAb, AML‐2–23, the remaining cells are used as an autograft in a patient treated with high‐dose chemotherapy and radiotherapy. In order to enhance the ability of the MoAb to lyse leukemic cells in the remission bone marrow, we have explored the effect of neuraminidase treatment on leukemia cells. In this paper we describe that myeloid leukemia cells expressing low levels of LNF‐III by immunofluorescence can be shown to have high levels of LNF‐III after neuraminidase treatment. In addition, we show that normal bone marrow progenitor cells do not have cryptic LNF‐III antigen, thus allowing the application of this finding to the clinical setting. Moreover, we have shown that leukemia colony‐forming cells from one patient with acute myelogenous leukemia express cryptic LNF‐III and that after exposure to neuraminidase there was an increased ability of PM‐81 in the presence of complement to eliminate these colony forming cells. These data indicate that the LNF‐III moiety is almost universally expressed on myeloid leukemia cells and their progenitors but not expressed on normal progenitors. Thus, it may be possible to enhance leukemia cell kill in vitro by neuraminidase treatment of bone marrow.
Cell surface expression of 4β‐galactosyltransferase accompanies rat parotid gland acinar cell transition to growth
Authors: Marchase Richard B. , Kidd Vincent J. , Rivera Angel A. , Humphreys‐Beher Michael G. 
Published: December 2004

AbstractRat parotid gland acinar cells stimulated to divide by a chronic regimen of isoproterenol demonstrate a dramatic increase in the synthesis of the glycosyltransferase 4β‐galactosyltransferase. A plasma membrane localization for much of the increase in 4β‐galactosyltransferase was determined by density gradient membrane fractionation. Golgi‐enriched fractions showed no increase in specific activity, while plasma membrane activity increased 40‐fold. This selective increase at the cell surface was confirmed by immunofluorescence of intact, nonpermeabilized cells from treated glands, using a monospecific antibody prepared against the purified bovine milk transferase. In detergent‐permeabilized cells staining of nontreated cells was seen only as groups of perinuclear vesicles, presumed to be Golgi apparatus. In isoproterenol‐treated and permcabilized cells both presumptive Golgi and cell surface staining was apparent. Enzyme assays performed on intact cells established that the enzyme's active site was oriented to the exterior of the cells. The transferase could be detected as early as 3 hr after the primary challenge with isoproterenol. Pretrcatment of rats with cycloheximide prevented its appearance.
Inhibition of the proliferation of Nb2 cells by femtomolar concentrations of cholera toxin and partial reversal of the effect by 12‐O‐tetradecanoyl‐phorbol‐13‐acetate
Authors: Pines M. , Ashkenazi A. , Cohen‐Chapnik N. , Binder L. , Gertler A. 
Published: December 2004

AbstractOne hour of exposure to cholera toxin is sufficient to elicit a significant delay in the initiation of DNA synthesis and cell division in lactogenic hormone‐dependent Nb2‐11C lymphoma cells. The inhibitory effect occurs already at very low concentrations of cholera toxin (5–50 fM), at which it is not accompanied by a detectable increase in intracellular cAMP, or ADP‐ribosylation of the alpha subunit of Gs, the stimulatory guanine nucleotide binding protein of adenylate cyclase; IBMX, the phosphodies‐terase inhibitor, acts synergistically to cholera toxin, indicating that a minute increase in cAMP may be sufficient for the inhibition. This indication is substantiated by the finding that dibutyryl cAMP also inhibits cell proliferation. Phorbol diester reverses partially the inhibitory activity of cholera toxin. It is most likely that this effect does not result from blocking the increase in cAMP, but rather from some subsequent, yet unidentified, events. The inhibitory effect of cholera toxin is not dependent on the concentration of the proliferation‐stimulating lactogenic hormone and cannot be abolished or reduced by excess of the hormone. Cholera toxin also inhibits the autonomous proliferation of a lactogenic hormone‐independent cell line (Nb2‐SP); however, in this case the inhibition is not affected by TPA.
Major 56,000‐dalton, soluble phosphoprotein present in bovine sperm is the regulatory subunit of a type II cAMP‐dependent protein kinase
Authors: Paupard Marie‐Christine , MacLeod Janet , Wasco Wilma , Orr George A. 
Published: December 2004

AbstractIt has been shown that cAMP‐dependent phosphorylation of a soluble sperm protein is important for the initiation of flagellar motion. The suggestion has been made that this motility initiation protein, named axokinin, is the major 56,000‐dalton phos‐phoprotein present in both dog sperm and in other cells containing axokinin‐like activity. Since the regulatory subunit of a type II cAMP‐dependent protein kinase is a ubiquitous cAMP‐dependent phosphoprotein of similar subunit molecular weight as reported for axokinin, we have addressed the question of how many soluble 56,000‐dalton cAMP‐dependent phosphoproteins are present in mammalian sperm. We report that in bovine sperm cytosol, the ratio of the type I to type II cAMP‐dependent protein kinase is approximately 1:1. The type II regulatory subunit is related to the non‐neural form of the enzyme and undergoes a phosphorylation‐dependent electrophoretic mobility shift. The apparent subunit molecular weights of the phospho and dephospho forms are 56,000 and 54,000 daltons, respectively. When bovine sperm cytosol or detergent extracts are phosphorylated in the presence of catalytic subunits, two major proteins are phosphorylated and have subunit molecular weights of 56,000 and 40,000 daltons. If, however, the type II regulator subunit (RII) is quantitatively removed from these extracts using cither immobilized cAMP or an anti‐RII monoclonal affinity column, the ability to phosphorylate the 56,000‐ but not 40,000‐dalton polypeptide is lost. These data suggest that the major 56,000 dalton cAMP‐dependent phosphoprotein present in bovine sperm is the regulatory subunit of a type II cAMP‐dependent protein kinase and not the motility initiator protein, axokinin.
Oligosaccharide heterogeneity of glycoproteins sulfated during the vegetative growth of Dictyostelium discoideum
Authors: Davis Simon J. 
Published: December 2004

AbstractMacromolecules are sulfated during the vegetative growth of Dictyostelium discoideum. A characterisation of the structures of sulfated oligosaccharides associated with these macromolecules indicates that the oligosaccharides are heterogeneous. Endoglycosidase and pronase digestion were used with gel‐filtration chromatography to obtain two different Oligosaccharide fractions and a glycopeptide fraction; these were further characterised by ion‐exchange and lectin‐affinity chromatography and by acid hydrolysis. The data indicate that up to 43% of the sulfate is associated with typical N‐linked oligosaccharides, that up to 5% is associated with N‐linked oligosaccharides that are either very large or extremely highly charged, and that the remaining sulfate is associated with oligosaccharides non‐N‐linked to protein. Each fraction was also shown to be heterogeneous at most other structural levels. Electrophoretic analyses following the endoglycosidasc and pronase treatments indicated that all of the macromolecules are glycoproteins and suggested further that at least two of the Oligosaccharide fractions are located on different groups of glycoproteins.
Comparative study of nuclear and cytoplasmic glycogen isolated from mutant HD33 ascites cells
Authors: Kopun Marijana , Granzow Christof , Krisman Clara R. 
Published: December 2004

AbstractMutant cells of the HD33 subline of the Ehrlich‐Lettré ascites tumor synthesize and store glycogen mainly intranuclearly, when growing in vivo, and exclusively in the cytoplasm, when permanently cultivated as a suspension cell strain. To investigate whether there exist differences between glycogen of nuclear and cytoplasmic origin, the ultrastructure and the biophysical and biochemical properties of glycogen from in vivo and in vitro grown HD33 ascites cells were compared. Pronounced heterogeneity and differences in glycogen particle ultrastructure were evident in situ and after isolation of the native, high‐molecular polysaccharide. Nuclear glycogen contains a fraction of heavier molecules (up to 2 × 109) and larger particles (up to 340 nm) which could not be found in the cytoplasmic preparations, which contained only particles smaller than 140 nm. The subparticles of β‐type are similar in both nuclear and cytoplasmic glycogen. The absorption spectra and glucose analysis after degradation with phosphorylase and debranching enzyme indicate that nuclear glycogen has a higher degree of branching, associated with a decrease in the average chain length between the branching points, and shorter external polyglucosidic chains than cytoplasmic glycogen. This is the first report about the analysis and properties of isolated nuclear glycogen.
Type I protein kinase C isozyme in the visual‐information‐processing pathway of monkey brain
Authors: Huang Freesia L. , Yoshida Yasuyoshi , Nakabayashi Hiroki , Friedman David P. , Ungerleider Leslie G. , Young W. Scott , Huang Kuo‐Ping 
Published: December 2004

AbstractPreviously using PKC isozyme‐specific antibodies for immunoblot analysis, we demonstrated the heterogeneous distribution of PKC isozymes in various regions of monkey and rat brains and that type I PKC was most abundant in cerebellum, hippocampus, amygdala, and cerebral cortex (Huang et al.: J Biol Chem 262:15714–15720, 1987). Using these antibodies, we have also demonstrated that type I, II, and III PKC are products of PKC genes γ, β, and α, respectively (Huang et al.: Biochem Biophys Res Commun 149:946–952, 1987). By immunocytochemical analysis, type I PKC‐specific antibody showed strong reactivity in various types of neuron in hippocampal formation, amygdala, cerebellum, and neocortex. In hippocampal formation, granule cells of dentate gyrus and pyramidal cells of hippocampus were heavily stained. By immunoblot analysis, relative levels of PKC isozymes in several areas of monkey cerebral cortex involved in the visual information processing and storage were determined. Both type II and III PKCs appeared to be evenly distributed and at moderate levels, type I PKC formed a gradient of increasing concentration rostral along the cerebral cortex of occipital to temporal and then to the limbic areas. Neurobehavioral studies have demonstrated that the neocortical and limbic areas of the anterior and medial temporal regions participate more directly than the striate, prestriate, and posterior temporal regions in the storage of visual representations and that both hippocampus and amygdala are important in the memory formation. As type I PKC is present at high levels in hippocampus, amygdala, and anterior temporal lobe, we predict that the type I protein kinase C may participate in the plastic changes important for mnemonic function.
Identification of HTLV‐I gag protease and its sequential processing of the gag gene product
Authors: Hatanaka Masakazu , Nam Seok Hyun 
Published: December 2004

AbstractThe full‐length provirus of human T‐cell leukemia virus type I (HTLV‐I) was isolated from MT‐2, a lymphoid cell line producing HTLV‐I. In transfected cells, structural proteins of HTLV‐I, the gag and env products, were formed and processed in the same manner as observed in MT‐2 cells. The nucleotide sequence was determined for a region between the gag and pol genes of the proviral DNA clone containing an open‐reading frame. The deduced amino acid sequences show that this open‐reading frame encodes a putative HTLV‐I protease.The protease gene (pro) of HTLV‐I was investigated using a vaccinia virus expression vector. Processing of 53k gag precursor polyprotein into mature p19, p24, and p15 gag structural proteins was detectable with a recombinant plasmid harboring the entire gag‐ and protease‐coding sequence. We demonstrated that the protease processed the gag precursor polyprotein in a trans‐action. A change in the sequence Asp(64)‐Thr‐Gly, the catalytic core sequence among aspartyl proteases, to Gly‐Thr‐Gly was shown to abolish correct processing, suggesting that HTLV‐I protease may belong to the aspartyl protease group. The 76k gag‐pro precursor polyprotein was identified, implying that a cis‐acting function of HTLV‐I protease may be necessary to trigger the initial cleavage event for its own release from a precursor protein, followed by the release of p53 gag precursor protein. The p53 gag precursor protein is then processed by the trans‐action of the released protease to form p19, p24, and p15.