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isolation and characterization of chinese hamster ovary cell lines sensitive to mitomycin c and bleomycin.seven chinese hamster ovary k1 cell lines exhibiting sensitivity to anticancer drugs have been isolated by a replica-plating technique. five of the mutants are hypersensitive (approximately 7-fold, as judged by d37 values) to the dna cross-linking agent mitomycin c. of these, one is also appreciably sensitive to uv light. significant variations in their cross-sensitivity to cis-platinum(ii) diammine dichloride, chlorambucil, and adriamycin have also been observed. two additional mutants have bee ...19852413991
characterization of chromosome replication during s-phase with bromodeoxyuridine labelling in chinese hamster ovary and hela cells.chinese hamster ovary (cho) and hela cells were successively pulse labelled at 1-h intervals after the cultures were synchronized at the end of g1 (monitored by flow cytometry). the metaphases analysed afterwards showed r-type replication patterns after 1-h pulses during the early s-phase (se; from h 1-5 after release) and replication of g- and c-bands in late s-phase (sl: from h 6-8 after release). the transition from se to sl is abrupt, constituting a sudden switch of replication between diffe ...19852414077
[experimental induction and cytogenetic study of yolk sac tumors in chinese hamsters].the development of teratomas and yolk sac tumors from displaced yolk sac is well documented in rats and mice. however, precise mechanisms of induction and pathogeneses of these tumors still remain unknown. the present study was undertaken in order to elucidate whether this phenomenon is common to all rodents, to identify the origin of these tumors and to explore specific chromosomal changes in yolk sac tumors. ten benign teratomas and two malignant tumors with teratomatous elements were obtained ...19852414200
the interaction between bleomycin and radiation on cell survival and dna damage in mammalian cell cultures.chinese hamster ovary (cho) cells were exposed to various concentrations of bleomycin (blm) for 0.5, 1, 2, 4, 6, or 24 hours, followed with graded doses (0 to 800 rad) or radiation. the response to this chemotherapy-radiation combination treatment was measured by cell survival studies and dna damage as determined by alkaline elution assay. isobolograms for 1 and 2 log cell kills showed that cytotoxicity from blm and from radiation were additive at 4, 15 and 75 milliunit/ml (mu/ml) for 1 or 4 hou ...19852415499
specific staining of human chromosomes in chinese hamster x man hybrid cell lines demonstrates interphase chromosome territories.in spite of carl rabl's (1885) and theodor boveri's (1909) early hypothesis that chromosomes occupy discrete territories or domains within the interphase nucleus, evidence in favor pf this hypothesis has been limited and indirect so far in higher plants and animals. the alternative possibility that the chromatin fiber of single chromosomes might be extended throughout the major part of even the whole interphase nucleus has been considered for many years. in the latter case, chromosomes would onl ...19852416668
sce induction and harlequin staining in mycoplasma-contaminated chinese hamster cells.chinese hamster v79 and cho cells infected with mycoplasma hyorhinis show elevated sister-chromatid exchange (sce) levels but normal cell proliferation and levels of chromosomal aberrations when compared with uninfected cells. harlequin staining patterns differ from those seen with uninfected cells at similar levels of bromodeoxyuridine (brdurd), indicating that brdurd is rapidly depleted from the medium by the mycoplasmal uridine phosphorylase and therefore becomes unavailable over the two cell ...19852417788
the soluble form of rat liver alpha-mannosidase is immunologically related to the endoplasmic reticulum membrane alpha-mannosidase.the soluble alpha-mannosidase of rat liver, originally described as a cytoplasmic alpha-mannosidase, has been purified to homogeneity by conventional techniques. the purified enzyme has an apparent molecular weight of 350,000 and is composed of 107-kda subunits. the soluble alpha-mannosidase has the same enzymatic properties as the endoplasmic reticulum (er) membrane alpha-mannosidase of rat liver (bischoff, j., and kornfeld, r. (1983) j. biol. chem. 258, 7909-7910) which is believed to play a r ...19862420791
amplification and expression of genes associated with multidrug resistance in mammalian cells.in multidrug resistance, which is observed clinically and in tissue culture, cells that are challenged with certain cytotoxic drugs develop resistance not only to the selective agent but also to other, seemingly unrelated, agents. the multidrug-resistant phenotype is associated with dna sequence amplification and with the overproduction of a number of cytosolic and membrane glycoproteins. the differential amplification and altered expression of at least two related genes, termed multidrug-resist ...19862421411
retinoic acid inhibits junctional communication between animal cells.retinoic acid inhibits junctional communication between a variety of vertebrate cell types in culture. it reduces the intercellular transfer of 3h-nucleotides between syrian hamster kidney fibroblasts (bhk 21/13), chinese hamster lung fibroblasts (v79), rat liver epithelial cells (brl), swiss mouse embryo fibroblasts (3t3), rainbow trout gonadal fibroblasts (rtg2) and xenopus embryo fibroblasts (xen). it also reduces metabolic cooperation between hypoxanthine-guanine phosphoribosyl transferase d ...19862423263
the purification and characterization of rat gamma interferon by use of two monoclonal antibodies.two mouse monoclonal antibodies, designated db-1 and db-2, were isolated and used for the purification and characterization of recombinant rat interferon gamma (rrif-gamma) derived from chinese hamster ovary (cho) cells. the two antibodies belong to different classes (db-1 is an igg1 and db-2 an iga) and display similar epitope specificities as shown in competition binding experiments. both antibodies, raised against rrif-gamma, exhibited high affinity for rat and mouse gamma interferon and effi ...19862423637
relationships between metabolic deactivation of icr compounds and their differential mutagenicity in bacteria and cultured mammalian cells.preparations of chinese hamster ovary (cho) cells decreased the genotoxicity of 3 icr compounds (icr 191, icr 191-oh and icr 170-oh), while they did not affect the genotoxicity of icr 170 in the salmonella reversion test nor in a dna-repair test in e. coli. these data may contribute towards the explanation of the lack of activity of the two hydroxylated compounds in the cho/hgprt forward mutation system, as well as the different rank of mutagenicity of the two chloroethyl compounds in bacteria ( ...19862425256
induction of b2 rna polymerase iii transcription by heat shock: enrichment for heat shock induced sequences in rodent cells by hybridization subtraction.when hybridization subtraction was used to enrich for sequences induced by heat shock in chinese hamster cells, b2 sequences were found to be one of the major sequences enriched. with cloned b2 probes, we found that the level of the short, 0.1 to 0.6 kb, polyadenylated rna polymerase iii transcripts of this repetitive genetic element increased approximately 10 to 20 fold after heat shock. transcription of b2 rna by rna polymerase iii was rapidly induced after heat shock based on time course stud ...19862426659
mutagenicity and carcinogenicity of nitroarenes and their sources in the environment.nitroarenes are postulated to play a principal part among mutagens/carcinogens which are induced in the combustion process and, in addition, are widely distributed in the environment. this review deals with the following points concerning nitroarene toxicity. data on the mutagenicity of nitroarenes obtained by short-term bioassays are expected to provide us with sufficient information for us to determine their genotoxicity and carcinogenicity. therefore, mutagenicity detected with salmonella, es ...19862427276
delayed dna methylation is an integral feature of dna replication in mammalian cells.in the majority of sites of methylation in the dna of mammalian cells, the symmetry of methylation is restored within a few minutes of the passage of a replication fork. however, it has been shown that daughter strand methylation in immortalised cell lines is delayed in a substantial minority of sites for up to several hours after replication. we report here the results of two new approaches to the determination of the functional significance of delayed dna methylation in mammalian cells. firstl ...19862427346
two chinese hamster ovary glycosylation mutants affected in the conversion of gdp-mannose to gdp-fucose.a biochemical basis for the pea and lentil lectin resistance of two chinese hamster ovary (cho) cell mutants, lec13 and lec13a, was investigated. studies of the g glycopeptides of vesicular stomatitis virus grown in the mutants indicated that lec13 cells essentially lack the ability to add fucose to complex carbohydrates while lec13a cells synthesize significant proportions of fucosylated, complex moieties. however, both mutants were known to be reverted to lectin sensitivity by growth in l-fuco ...19862428310
enhancement of o6-methylguanine-dna-methyltransferase activity induced by various treatments in mammalian cells.the o6-methylguanine-dna-methyltransferase (methyltransferase) activity was determined in a rat hepatoma cell line after treatment with ultraviolet or gamma-irradiation, heat treatment, or incubation with cis-diamminedichloroplatinum(ii),2-methyl-9-hydroxyellipticinium, or bleomycin. the assay measured the removal of o6-methylguanine from 3h-alkylated dna by cellular extracts. the results show that 48 h after the various treatments, the methyltransferase activity is increased by 2- to 5-fold. th ...19862428481
simultaneous activation by 5-azacytidine of intracisternal r particles and murine intracisternal-a particle related sequences in syrian hamster cells.5-azacytidine treatment of syrian hamster (mesocricetus auratus) cells, bhk21-cl.13, and primary embryo fibroblasts, activates the production of intracisternal type r particles (irp) as ascertained by electron microscopy scanning and counting. efficiency of the activation is dose- and time of treatment-dependent. the transcription of mouse intracisternal a particle (iap) related sequences, which have been described in syrian hamster genomic dna, is increased at the same time. this correlates wit ...19862430359
species differences in the distribution of substance p and tyrosine hydroxylase immunoreactivity in the olfactory bulb.these studies document species differences in the distribution of the peptide substance p and the catecholamine-synthesizing enzyme tyrosine hydroxylase (th) within a central nervous system region of a number of mammalian species including the mouse, rat, guinea pig, rabbit, cat, and two species of hamster (chinese and syrian). substance p-containing neuronal perikarya were observed in the main olfactory bulb (mob) of both species of the hamster, but not in the mob of the other species examined. ...19862431012
overexpression and amplification of five genes in a multidrug-resistant chinese hamster ovary cell line.multidrug-resistant cells are cross-resistant to a wide range of unrelated drugs, many of which are used in cancer chemotherapy. we constructed a cdna library from rna of the multidrug-resistant chinese hamster ovary cell line chrc5. by differential screening we isolated cdnas derived from mrnas that are overexpressed in this cell line. the cdnas could be grouped in five classes on the basis of transcript lengths detected in rna blots. we infer that each class codes for a separate protein. the c ...19862431283
acid intracellular vesicles and the cytolysis of mammalian target cells by entamoeba histolytica trophozoites.entamoeba histolytica kills mammalian target cells in a multi-step sequential process with separate adherence, cytolytic, and phagocytic events. in the studies reported here, we used fluorescein isothiocyanate linked to dextran to label the endocytic vesicles of the hm1 strain of e. histolytica and measure vesicle ph (5.1 +/- 0.2 by spectrofluorimetry). concentrations of nh4cl (1.0-10.0 mm) sufficient to increase vesicle ph to greater than or equal to 5.7 inhibited amebic killing of target chine ...19862432267
the expression of integrated plasmid dna depends on copy number.the effect of copy number, integration site, and enhancers on the expression of stably integrated exogenous dna was examined in chinese hamster cells. three similar plasmids were constructed with the mouse beta maj-globin promoter fused to the galk gene either with no enhancer or with the sv40 or harvey sarcoma virus (hasv) enhancer. eighteen stable cell lines were obtained and characterized with respect to plasmid copy number and galactokinase activity. at copy numbers of four or less, the enha ...19872433136
scrape-loading and dye transfer. a rapid and simple technique to study gap junctional intercellular communication.gap junction-mediated intercellular communication has been recognized in cells from different tissues of various organisms and has been implicated in a variety of cellular functions and dysfunctions. here we describe a new, direct and rapid technique with which to study this cellular phenomenon. it employs scrape-loading to introduce a low molecular weight (mw) fluorescent dye, lucifer yellow ch (mw 457.2) into cells in culture and allows the monitoring of its transfer into contiguous cells. in ...19872433137
effect of monoclonal antibody to pertussis toxin on toxin activity.two distinct monoclonal antibodies, one to pertussis toxin subunit s2, called 9g8, and another to subunits s2 and s3, called 11e6, were generated from the hybridomas of myeloma sp2/0 and spleen cells of balb/c mice immunized mainly with the subunit s234 complex. binding ability of 9g8 and 11e6 to the subunits was confirmed by the enzyme-linked immunosorbent assay and immunoblotting analysis. generation of 11e6 bound to both s2 and s3 might mean that there is common antigenicity between s2 and s3 ...19872435660
human ribosomal dna fragments amplified in hamster cells are transcribed only by rna polymerase ii and are not silver stained.cloned human rrna gene fragments that included the promoter region were introduced into chinese hamster dihydrofolate reductase-deficient (dhfr-) cells by cotransformation with a dhfr minigene and amplified by selection for methotrexate resistance. the human ribosomal dna was transcribed by rna polymerase ii, not rna polymerase i or iii. the metaphase chromosome regions containing the transcriptionally active human ribosomal dna failed to show silver staining.19872436041
[phenthiuram induction of micronuclei in mammalian somatic cells].genotoxic effects of the preparative-form phenthiuram on the induction of micronuclei in cultured chinese hamster v-79 cells and polychromatic erythrocytes of mouse bone marrow have been studied. the phenthiuram induced cytogenetic effects in cultured somatic cells without metabolic activation being only in toxic concentrations. similar effects were observed in polychromatic erythrocytes after treatment with subtoxic concentrations.19872436363
induced sister chromatid exchange frequency is not increased in homogeneously staining regions that contain amplified genes.gene amplification is a process by which cells become resistant to selective agents by increasing gene copy number and overproducing specific enzymes. the molecular mechanism by which gene amplification occurs is unknown, but unequal sister chromatid exchange (sce) has been suggested as one possibility. unequal sce results in one chromatid containing an extra copy of a selected gene that is deleted in the sister chromatid. two predictions of the unequal sce model are that agents that increase sc ...19872436746
characterization of an endogenous substrate of the insulin receptor in cultured cells.using antiphosphotyrosine antibodies, we have characterized the tyrosine phosphorylation of an endogenous substrate of the insulin receptor in fao hepatoma cells and in chinese hamster ovary cells transfected with a eukaryotic expression vector containing the human insulin receptor cdna. in fao cells, besides the beta-subunit of the insulin receptor, a protein with a molecular mass between 170 and 210 kda designated pp185, undergoes tyrosine phosphorylation immediately after insulin stimulation ...19872439512
high-frequency reactivation of x-linked genes in chinese hamster x human hybrid cells.three genes on the human inactive x chromosome retained in the chinese hamster x human hybrid cell line x8/6t2 have been reactivated using the demethylating agent, 5-azacytidine (5-aza-cr). pulse-labeling and histochemical methods permitted detection and measurement of reactivation rates of the hypoxanthine phosphoribosyltransferase (hpt) and glucose-6-phosphate dehydrogenase (g6pd) genes within 48 h of treatment. about 50% of the cells became active for these genes, which represents a reactivat ...19872440116
quinoneimines as substrates for quinone reductase (nad(p)h: (quinone-acceptor)oxidoreductase) and the effect of dicumarol on their cytotoxicity.several quinoneimines have been shown to be substrates for partly purified rat liver cytosolic quinone reductase with either nadh or nadph as cofactor. km and vmax values with nadh as cofactor for n-acetyl-p-benzoquinoneimine were 54.9 microm and 278 mumol/min/mg; for 2-amino-1,4-naphthoquinoneimine, 2.8 microm and 38 mumol/min/mg; for n,n-dimethylindoaniline, 1.7 microm and 22 mumol/min/mg; and 2-acetamido-n,n-dimethylindoaniline, 0.4 microm and 9 mumol/min/mg. all the quinoneimines showed subs ...19872440444
pyrazoloacridines, a new class of anticancer agents with selectivity against solid tumors in vitro.a series of 2-aminoalkyl-5-nitropyrazolo[3,4,5-kl]acridines (pyrazoloacridines) was evaluated in vitro for activity against a panel of human tumor cell lines of breast, colon, or lung origin. several pyrazoloacridines were found to possess solid tumor selectivity relative to their activity against murine leukemia l1210 cells as well as human lymphoblastoid cells. the superior compounds in this regard were also found to exhibit excellent activity against primary human tumors in stem cell clonogen ...19872440564
sister chromatid exchanges induced by dna demethylating agents persist through several cell cycles in mammalian cells.eukaryotic dna methylation has been extensively studied in recent years. the ability of many carcinogens to interfere with dna methylation has not yet been directly related to their tumorigenic activity. recent data obtained using l-ethionine and 5-azacytidine--both demethylating agents--showed a small but significant increase in the sister chromatid exchange (sce) rate induced in mammalian cells (human lymphocytes and cho cells). in this paper we show that the sce increase induced by both these ...19872440615
a monoclonal antibody against a family of nuclear pore proteins (nucleoporins): o-linked n-acetylglucosamine is part of the immunodeterminant.using nuclear envelopes from chinese hamster ovary (cho) cells as an antigen, a mouse monoclonal antibody (igm; designated mab chon211) that specifically binds to components of the nuclear pore complex (nucleoporins) was isolated. immunofluorescence localization of the antigen recognized by mab chon211 revealed a punctate pattern restricted to the nuclear envelope; this pattern changed dramatically during the cell cycle. when examined by electron microscopy, the antigen was largely restricted to ...19872442757
recovery of dna synthesis from inhibition by ultraviolet light in mammalian cells.in general mammalian cells recover from dna synthesis inhibition by ultraviolet light (u.v.) before most of the pyrimidine dimers have been removed from the genome. this is a complex phenomenon whose biological significance has not been fully assessed. in chinese hamster v79 cells this recovery seems to be directly coupled to an enhanced rate of double-stranded dna elongation. the presence of the dna polymerase alpha inhibitor, aphidicolin, after u.v. irradiation produces two different responses ...19872443519
three distinct epitopes of the human msk20 cell surface antigen are expressed on human and nonhuman primate cells.five monoclonal antibodies which indirectly agglutinate human rbc were isolated and designated 2d8, 4e12, 4h10, 5a8, and 5h5. expression of the antigen defined by 2d8 was 100% concordant with the presence of human chromosome 19 in a panel of human-chinese hamster somatic cell hybrids. secondary clones isolated from antigen positive hybrids exposed to antibody 2d8 and complement were shown to have lost both the cell surface antigen and chromosome 19. small terminal deletions of 19p were observed ...19872443846
characterization of recombinant human alpha 2-antiplasmin and of mutants obtained by site-directed mutagenesis of the reactive site.human alpha 2-antiplasmin (alpha 2ap) has been expressed in chinese hamster ovary cells and purified from conditioned media. the recombinant protein (r alpha 2ap) is immunologically identical with natural alpha 2ap and indistinguishable with respect to plasmin(ogen) binding properties. second-order rate constants (k1) for the interaction of alpha 2ap and r alpha 2ap with plasmin are both (1-2) x 10(7) m-1 s-1. in order to examine the effects of alterations within the reactive site of alpha 2ap, ...19872444252
chinese hamster ovary cell lysosomes rapidly exchange contents.we have used cell fusion to address the question of whether macromolecules are rapidly exchanged between lysosomes. donor cell lysosomes were labeled by the long-term internalization of the fluid-phase pinocytic markers, invertase (sucrase), lucifer yellow, fitc-conjugated dextran, or texas red-conjugated dextran. recipient cells contained lysosomes swollen by long-term internalization of dilute sucrose or marked by an overnight fitc-dextran uptake. cells were incubated for 1 or 2 h in marker-fr ...19872447096
the interaction between pertussis toxin and 10 monoclonal antibodies.data on the epitope specificity of 10 monoclonal hybridoma antibodies (mabs) that showed positive reaction in enzyme-linked immunosorbent assay (elisa) towards pertussins toxin (ptx) are presented. the relative functional affinity of the mabs was determined in a catching elisa system. the mabs were tested for their ability to inhibit the biological activities of this toxin in two in vitro systems, viz. haemagglutination (ha) and chinese hamster ovary cell (cho) test, and in three in vivo assays: ...19872447745
use of dio-c5-3 to improve hoechst 33342 uptake, resolution of dna content, and survival of cho cells.chinese hamster cells (line cho) stained with either 9 microm hoechst 33342 (ho) alone or in combination with the membrane potential fluorochrome dio-c5-3 (dio) were analyzed using uv laser powers between 25 and 500 mw and sorted for determination of survival by a colony formation assay. the combination of ho-dio increased fluorescence twofold and provided coefficients of variation (cvs) as low as 3.0% under conditions where viability of cells, even at 500 mw excitation, was unaffected. ho-stain ...19882448153
neuromodulator-mediated phosphorylation of specific proteins in a neurotumor hybrid cell line (ncb-20).mouse neuroblastoma x embryonic chinese hamster brain explant hybrid cell line (ncb-20) forms functional synapses when intracellular cyclic amp levels are elevated for a prolonged period of time. ncb-20 cells were labeled with [32p]orthophosphate under conditions where 2-chloroadenosine gave maximum increases of 32p incorporation into tyrosine hydroxylase in nerve growth factor dibutyryl cyclic amp-differentiated pc12 (pheochromocytoma) cells. when ncb-20 cells were exposed to activators [5-hydr ...19882450174
methotrexate-induced amplification of the bovine lutropin genes in chinese hamster ovary cells. relative concentration of the alpha and beta subunits determines the extent of heterodimer assembly.methotrexate-induced gene amplification increased the expression of biologically active bovine luteinizing hormone (blh) approximately 11-fold after stable transfection of a line of chinese hamster ovary cells with genes encoding dihydrofolate reductase and the alpha and beta subunits of blh. subsequent analysis of the bovine genes revealed that while the alpha gene was amplified in response to methotrexate selection, the lh beta subunit gene remained unaffected. this effect was probably due to ...19882452162
the role of the asparagine-linked oligosaccharides of the alpha subunit in the secretion and assembly of human chorionic gonadotrophin.human chorionic gonadotropin (hcg) is a member of a family of heterodimeric glycoprotein hormones that have a common alpha subunit but differ in their hormone-specific beta subunit. site-directed mutagenesis of the two asparagine-linked glycosylation sites of hcg alpha was used to study the function of the individual oligosaccharide chains in secretion and subunit assembly. expression vectors for the alpha genes (wild-type and mutant) and the hcg beta gene were constructed and transfected into c ...19882452167
the cellular regulation of vesicle exocytosis by entamoeba histolytica.we studied the cellular regulation of vesicle exocytosis by entamoeba histolytica utilizing release of endocytosed 125iodine (125i) labeled tyrosine conjugated dextran; 125i-dextran entered the acid ph vesicles of the amebae and was not degraded during these studies. exocytosis was temperature dependent with 74%, 36%, 4%, and 0% of 125i-dextran released after 120 min at 37 degrees c, 31 degrees c, 25 degrees c, and 4 degrees c, respectively (p less than 0.01 for each). exocytosis at 37 degrees c ...19882452880
glycolipid reanchoring of t-lymphocyte surface antigen cd8 using the 3' end sequence of decay-accelerating factor's mrna.decay-accelerating factor (daf) is one of a family of cell-associated proteins that undergo posttranslational modifications in which glycolipid anchoring structures are substituted for membrane-spanning sequences. the signals that direct the covalent substitution reaction in these proteins are unknown. human daf was expressed in chinese hamster ovary (cho) cells and murine bw lymphocytes. in both cases, the xenogeneic daf in transfectants incorporated a glycolipid anchor. a chimeric cd8-daf cdna ...19882453063
structural and functional analysis of poly(adp ribose) polymerase: an immunological study.poly(adp ribose) polymerase (ec 2.4.2.30) was studied using monoclonal antibodies for three different epitopes on the enzyme. the epitopes were mapped in relation to the functional domains of the protein and the inhibitory properties of the antibodies. the intranuclear and interspecies immunoreactivity of the enzyme was also investigated. the epitope of antibody 2 was mapped to the 17 kda fragment generated by chymotryptic digestion of the c-terminal 54 kda nad-binding domain. antibody 9 binds t ...19882454668
an acid protease secreted by transformed cells interferes with antigen processing.the major excreted protein of malignantly transformed mouse fibroblasts (mep), which is the precursor to lysosomal cathepsin l, was used to study the effect of exogenous acid proteases on antigen processing. when mep and native pigeon cytochrome c were added to chinese hamster ovary (cho) cells expressing transfected major histocompatability complex class ii gene products, the antigen-specific t-cell hybridoma 2b4 did not respond to the antigen. mep appears to destroy the antigen in an acid comp ...19882454929
monoclonal antibodies to distinctive epitopes on the alpha and beta subunits of the fibronectin receptor.monoclonal antibodies (mabs) have been developed that can recognize epitopes that are unique to either the alpha or beta subunit of the fibronectin receptor (fnr). mabs 11b4 and 7a8 immunoblot the alpha subunit of fnr either in purified form from chinese hamster ovary (cho) cells or in nonionic detergent extracts of cells of human and rodent origin electrophoresed under reducing or nonreducing conditions. the mabs seem to be more reactive to the subunit when it has been electrophoresed under red ...19882455654
structures of the sugar chains of recombinant human granulocyte-colony-stimulating factor produced by chinese hamster ovary cells.recombinant human granulocyte-colony-stimulating factor (g-csf) was purified from chinese hamster ovary cells transfected with human g-csf cdna. the recombinant human g-csf was treated with alkaline borohydride and the oligosaccharide-alditols liberated were fractioned by gel filtration on a bio-gel p-4 column, followed by high-performance liquid chromatography by use of a strong anion exchanger. two oligosaccharide-alditols were obtained and their structures were identified by component analysi ...19882455710
sensitivities of bleomycin-resistant variant cells enhanced by 40 degrees c hyperthermia in vitro.cultured mammalian cells of higher (mouse l) and lower (chinese hamster v-79) sensitivities to bleomycin (blm) were repeatedly treated with blm (0.1 mg/ml), through which blm-resistant variant strains were induced. sensitivities of these variant strains to blm were enhanced by simultaneous treatment with 40 degrees c hyperthermia.19882455760
human immunodeficiency virus type 1 challenge of chimpanzees immunized with recombinant envelope glycoprotein gp120.the major envelope glycoprotein, gp120, of human immunodeficiency virus type 1 (hiv-1) was purified from a chinese hamster ovary cell line transfected with a truncated form of the hiv-1 env gene. the recombinant glycoprotein (rgp120) was formulated with aluminum hydroxide adjuvant and was used to immunize chimpanzees. the recombinant preparation was effective in eliciting cellular and humoral immunity as well as immunologic memory. anti-rgp 120 antibodies reacted with authentic viral gp120 in im ...19882455898
monoclonal antibody jsb-1 detects a highly conserved epitope on the p-glycoprotein associated with multi-drug-resistance.resistance to multiple chemotherapeutic agents is a common clinical problem in the treatment of cancer. this resistance may occur before primary therapy or be acquired during treatment. we have generated a monoclonal antibody (mab) (jsb-i), specific for a conserved epitope on the plasma membrane 170- to 180-kda glycoprotein, the expression of which is strongly correlated with the degree of multi-drug resistance (mdr). jsb-i strongly binds to both chinese-hamster-derived mdr cell lines and human ...19882458323
regulation of glycosphingolipid glycosyltransferase by low density lipoprotein receptors in cultured human proximal tubular cells.we have shown previously that low density lipoproteins (ldl) suppressed the synthesis of lactosylceramide in normal human proximal tubular cells, but stimulated such synthesis in proximal tubular cells from ldl receptor negative subjects (chatterjee, s., clarke, k., and kwiterovich, p.o., jr. (1986) j. biol. chem. 261, 13474-13479). to understand the mechanism(s) of this effect of ldl, we have studied here the effects of ldl on the activity of udp-galcer:beta-galactosyltransferase (galt-2). maxi ...19882458339
determination of monosaccharides in glycoproteins by reversed-phase high-performance liquid chromatography on 2.1-mm narrowbore columns.the monosaccharide composition of glycoproteins was determined by using a supelcosil lc-18 150 x 2.1 mm narrowbore column under reversed-phase conditions. methodology was developed for sample hydrolysis, n-acetylation, benzoylation, and cleanup prior to high-performance liquid chromatography. monosaccharides were identified and quantified at picomole levels. various glycoproteins were then analyzed, including ribonucleases a and b, mucin, and fetuin. in addition, the monosaccharide content of ch ...19882459148
structure and regulation of mammalian s-adenosylmethionine decarboxylase.in order to understand the structure and regulation of s-adenosylmethionine decarboxylase, cdna clones encoding this enzyme have been isolated from rat prostate and human fibroblast cdna libraries. the authenticity of the cdnas was verified by: (a) transfecting the chinese hamster ovary cells with the human cdna in the pcd vector which resulted in a transient 10-20-fold increase in s-adenosylmethionine decarboxylase activity in recipient cells; and (b) translating the mrna formed by transcriptio ...19882460457
site-specific mutagenesis defines the intracellular role of the asparagine-linked oligosaccharides of chorionic gonadotropin beta subunit.the beta subunit of human chorionic gonadotropin contains two asparagine (n)-linked oligosaccharides. to examine the structural and functional roles of these oligosaccharide units in vivo, we constructed mutant genes containing alterations in either the asparagine or threonine codons of the two glycosylation consensus sequences and inserted them into a eukaryotic expression vector. wild-type and mutant cg beta proteins were expressed in chinese hamster ovary cells alone or in the presence of nat ...19882460458
modes of genotoxicity of a macromolecular antibiotic, sn-07, a novel type of interstrand dna cross-linker.the modes of genotoxicity of a novel macromolecular antitumor antibiotic (sn-07) were examined using both prokaryotic and eukaryotic cells in vitro. the antibiotic induced a frameshift-type reverse mutation in ames salmonella typhimurium ta98 at 1.6-400 ng/plate with and without s9 mix. sn-07 also induced chromosomal aberrations and a forward mutation (6-tgr) in chinese hamster v79 cells after 1 h treatment at 12.5-100 ng/ml without metabolic activation. the alkaline elution technique revealed t ...19892462669
rapid fluorescence-based assay for radiosensitivity and chemosensitivity testing in mammalian cells in vitro.an efficient and rapid cytotoxicity assay has been developed, particularly for radiobiological studies, utilizing 96-well microtiter plates. several days after treatment, cell numbers per well were measured by fluorescent intensity using an automatic reader after staining with the dna specific dye hoechst 33258. for radiobiological applications, a microtiter plate irradiation box was designed and built which allowed a variable number of wells (minimum 4, maximum 16) to be irradiated at one time. ...19892463078
coordinate induction of metallothioneins i and ii in rodent cells by uv irradiation.sequence analysis of chinese hamster v79 lung fibroblast cdna clones, which code for uv radiation-inducible transcripts, revealed that many of the clones corresponded to metallothioneins (mts) i and ii. a third cdna clone, ddiu4, was found also to code for a similar-size uv-inducible transcript which was unrelated to mt by both sequence analysis and kinetics of induction. mti and mtii rnas rapidly increased in v79 cells within 1 h after uv irradiation, and maximum induction was seen by 4 h. this ...19882463464
the role of proteinases, interferons and hormones in proliferative activities of nonmalignant and malignant cells.proliferative responses, in form of colony forming ability, of nonmalignant chinese hamster fibroblasts (v79) and malignant human laryngeal carcinoma cells (hep) were examined after treatment with proteinases, proteinase inhibitors, interferons or hormones, in g0-phase, early g1-phase and g1/s point of the cell cycle. these phases of the cell cycle are of particular importance from the point of control and regulation of the proliferative activity of cells, and the effects of proteinases and othe ...19882464142
effects of heat shock proteins (mr 70,000) on protein and dna synthesis at elevated temperatures in vitro.the protective effect of mr 70,000 heat shock protein (hsp-70) during thermotolerance has been previously observed. however, it is not known what cellular processes or components may be protected by this protein during the tolerance state. in the studies reported here, the protective effects of purified hsp-70, the nonspecific heat-stable proteins fetuin and trypsin inhibitor (ovomucoid), and other proteins and agents such as bovine serum albumin, d2o, or glycerol on protein and dna synthesis du ...19892466556
monoclonal antibodies to pertussis toxin: utilization as probes of toxin function.six monoclonal antibodies (mabs) to pertussis toxin (pt) have been generated and characterized. five of these mabs (3cx4, 3c4d, 6d11c, 6fx1, and x2x5) interact with determinants on the catalytic subunit (s1) of pt, and one (6dx3) is specific for subunit s4. the mabs are divided into three groups based upon their ability to neutralize the effects of pt in a chinese hamster ovary (cho) cell assay. three of the mabs (3cx4, 3c4d and 6d11c) had high neutralization titers, one mab (6fx1) displayed wea ...19892466764
expression of a g alpha s/g alpha i chimera that constitutively activates cyclic amp synthesis.a chimeric g alpha subunit cdna, referred to as g alpha s/i(38), was constructed containing the complete 5'-untranslated region of g alpha s, the first 356 codons of the rat g alpha s and the last 36 codons and 428 base pairs of the 3'-untranslated region of the rat g alpha i cdna. transient expression of the g alpha s/i(38) protein in cos cells allowed detection of a chimeric protein which was recognized by antibodies generated against an internal g alpha s sequence as well as antibodies recogn ...19892466829
phase i clinical study for recombinant human granulocyte colony-stimulating factor.the pharmacokinetics, the safety and the efficacy of purified recombinant human granulocyte colony-stimulating factor (rh g-csf) expressed in chinese hamster ovary (cho) cells were studied in normal healthy adults. following the single subcutaneous dose, g-csf levels in sera elevated in a dose-dependent way, with c-max of approximately 140 pg per ml, t-max of 4 hours and t-half of 5 hours at a dose of 10 micrograms per ml. in accord with the elevation, blood neutrophil counts rose promptly and t ...19882467650
differential activation of the hprt gene on the inactive x chromosome in primary and transformed chinese hamster cells.we have investigated the genetic activation of the hprt (hypoxanthine-guanine phosphoribosyltransferase) gene located on the inactive x chromosome in primary and transformed female diploid chinese hamster cells after treatment with the dna methylation inhibitor 5-azacytidine (5azacr). mutants deficient in hprt were first selected by growth in 6-thioguanine from two primary fibroblast cell lines and from transformed lines derived from them. these hprt- mutants were then treated with 5azacr and pl ...19892471066
single channel currents of homo- and heterologous gap junctions between cardiac fibroblasts and myocytes.recently, the use of the double whole-cell patch-clamp technique enabled conductance measurements of single gap junctional channels. different values have been measured in pairs of rat lacrimal cells (6), murine acinar cells and chinese hamster ovary cells (9), embryonic chick heart- (10) and neonatal rat heart myocytes (7). we here present evidence that the conductance of gap junction channels between two different cell types originating from the same tissue, neonatal rat heart, is different. i ...19892471143
monoclonal antibodies that define neutralizing epitopes of pertussis toxin: conformational dependence and epitope mapping.the epitope specificities of 13 hybridomas secreting monoclonal antibodies (mabs) specific for pertussis toxin (pt) is described. hybridoma lines were derived by the fusion of spleen cells from mice immunized with native pt, formalin-detoxified pt, or isolated pt subunits (s1 to s5) with the myeloma line x63-ag8.653. five mabs showed a toxin-neutralizing ability, which was demonstrated by use of a chinese hamster ovary cell assay system and by a nad glycohydrolase assay. all five toxin-neutraliz ...19892474500
site-directed anti-peptide antibodies define the topography of the beta-adrenergic receptor.molecular cloning has revealed the primary structure of a number of g-protein-linked receptors. the organization and topography of these proteins predicted to have seven hydrophobic membrane-spanning domains, in contrast, have not been established. antibodies were prepared against 11 peptides corresponding to each of the hydrophilic sequences of the hamster beta 2-adrenergic receptor. each of the anti-peptide antibodies displayed immunoreactivity for its synthetic peptide antigen and beta 2-adre ...19892474546
generation of recombinant, enzymatically active human thyroid peroxidase and its recognition by antibodies in the sera of patients with hashimoto's thyroiditis.a full-length cdna clone for human thyroid peroxidase (tpo) inserted into the mammalian cell expression vector pece was stably transfected into chinese hamster ovary (cho) cells. clones were assayed for human tpo mrna, tpo protein, and tpo enzymatic activity. one subclone, expressing the highest tpo enzymatic activity, was used in further studies. facs analysis of these cells preincubated in hashimoto's serum revealed approximately 100-fold greater fluorescence compared with controls, indicating ...19892474568
gene for human cd59 (likely ly-6 homologue) is located on the short arm of chromosome 11.the cd59 (mem-43) antigen, which probably is a human homologue of mouse ly-6 antigens, is a broadly expressed mr 18,000-25,000 human leucocyte surface glycoprotein recognized by monoclonal antibody mem-43. ten mouse-human t-lymphocyte hybrids, carrying all mouse chromosomes and a limited number of human chromosomes, were analyzed for expression of cd59 by indirect immunofluorescence and immunoblotting with mem-43 antibody. karyotypic analysis of the tested clones showed that the presence of huma ...19892476389
two distinct pathways for camp-mediated down-regulation of the beta 2-adrenergic receptor. phosphorylation of the receptor and regulation of its mrna level.we have studied cyclic amp-mediated regulation of the beta 2-adrenergic receptor (beta 2ar). the effects of camp were assessed in chinese hamster fibroblast (chw) cells expressing either the wild type human beta 2ar receptor (ch-beta 2) or mutated forms of the receptor lacking the consensus sequences for phosphorylation by the camp-dependent protein kinase. treatment of the ch-beta 2 cells with the camp analogue dibutyryl camp (bt2camp) induces a time-dependent "down-regulation" of the number of ...19892476447
temporal order of gene replication in chinese hamster ovary cells.to investigate the molecular basis of the regulatory mechanisms responsible for the orderly replication of the mammalian genome, we have developed an experimental system by which the replication order of various genes can be defined with relative ease and precision. exponentially growing cho-k1 cells were separated into populations representing various stages of the cell cycle by centrifugal elutriation and analyzed for cell cycle status flow cytometry. the replication of specific genes in each ...19892476659
functional topology of human tissue-type plasminogen activator: characterization of two deletion derivatives and of a duplication derivative.three types of permanent chinese hamster ovary (cho) cell lines with different amplified expression constructs that abundantly secrete derivatives of human tissue-type plasminogen activator (t-pa) were established. the first one expresses a deletion derivative in which the kringle 2 domain (k2) has been removed (fgk1l). in the second derivative, the growth-factor-homologous domain (g) has also been deleted (fk1l); a third line expresses a duplication derivative of k2 (fk2k2l) lacking the (g) and ...19892477307
fine structure analysis of the chinese hamster as gene encoding asparagine synthetase.overlapping cdnas for chinese hamster ovary (cho) asparagine synthetase (as) were isolated from a library prepared from an as-overproducing cell line. the sequence was determined and shown to contain an open reading frame encoding a protein of mr 64,300. the predicted amino acid sequence for the cho as enzyme was compared to that of the human as enzyme and found to be 95% homologous. a potential glutamine amide transfer domain, with sequence similarity to amidotransferases from bacteria and yeas ...19892477309
effects of o-linked glycosylation on the cell surface expression and stability of decay-accelerating factor, a glycophospholipid-anchored membrane protein.decay accelerating factor (daf) is a glycophospholipid-anchored membrane glycoprotein that protects mammalian host cells from inadvertant complement lysis. the effects of inhibiting mucin-type o-glycosylation on the cell surface expression of daf were studied by introducing an expression vector for human daf into wild-type chinese hamster ovary and ldld cells. the ldld cells express reversible defects in the addition of galactose and n-acetylgalactosamine (galnac) to oligosaccharide chains on gl ...19892477368
dna base changes and rna levels in n-acetoxy-2-acetylaminofluorene-induced dihydrofolate reductase mutants of chinese hamster ovary cells.formerly, we isolated a series of dihydrofolate reductase-deficient chinese hamster ovary cell mutants that were induced by n-acetoxy-2-acetylaminofluorene. deletions and complex gene rearrangements were detected in 28% of these mutants; 72% contained putative point mutations. in the present study, we have localized the putative point mutations in the 25,000 base dhfr gene by rnase heteroduplex mapping. assignment of a position for each mutation was successful in 16 of 19 mutants studied. we clo ...19892477551
binding of pertussis toxin to eucaryotic cells and glycoproteins.the binding of pertussis toxin and its subunits to cell surface receptors and purified glycoproteins was examined. the interaction of pertussis toxin with components of two variant chinese hamster ovary (cho) cell lines was studied. these cell lines are deficient in either sialic acid residues (lec 2) or sialic acid and galactose residues (lec 8) on cell surface macromolecules. the binding of pertussis toxin to components of these cells differed from the binding of the toxin to wild-type compone ...19892478471
elimination of disulfide bonds affects assembly and secretion of the human chorionic gonadotropin beta subunit.human chorionic gonadotropin (hcg) consists of two noncovalently joined alpha and beta subunits similar to the other glycoprotein hormones. to study the function of the individual disulfide bonds in subunit assembly and secretion, site-directed mutagenesis was used to convert the 12 cysteine (cys) residues in the beta subunit of hcg to either alanine or serine. both cysteines of proposed disulfide pairs were also mutated. these mutant hcg beta genes were transfected alone or together with the wi ...19892478557
abortive infection with sindbis virus of a chinese hamster ovary cell mutant defective in phosphatidylserine and phosphatidylethanolamine biosynthesis.the effects of phosphatidylserine starvation on the infection with sindbis virus (an enveloped rna virus) have been investigated in a chinese hamster ovary (cho) cell mutant (strain psa-3) which requires exogenously added phosphatidylserine for cell growth because it lacks the ability to synthesize this phospholipid. when psa-3 cells were grown in the absence of phosphatidylserine, the cellular contents of phosphatidylserine and also phosphatidylethanolamine produced through decarboxylation of p ...19892479418
increased expression of glutathione s-transferase gene in cis-diamminedichloroplatinum(ii)-resistant variants of a chinese hamster ovary cell line.we have isolated cis-diamminedichloroplatinum(ii) (cddp)-resistant variants, c/cdp-1 and c/cdp-2, from a chinese hamster ovary (cho) cell line after a stepwise exposure to increasing concentrations of cddp, and a cddp-sensitive revertant, r-1, from c/cdp-2 after continuous incubation for 5 months in the absence of cddp, c/cdp-1 and c/cdp-2 showed 7- and 10-fold higher resistance to cddp, respectively, compared to cho cells. c/cdp-2 was cross-resistant to carboplatin, l-phenylalanine mustard (mel ...19892479474
cloning, sequence analysis and expression of a cdna encoding a novel insulin-like growth factor binding protein (igfbp-2).insulin-like growth factors bind with high affinity to specific binding proteins in extracellular fluids. to identify structural characteristics of igf-binding proteins that might define their physiological roles, we determined the complete primary structure of a novel human igf-binding protein (igfbp-2) from a cloned cdna. the cdna encodes a 328 amino acid igf-binding protein precursor which contains a 39-residue signal peptide. the mature 289 amino acid igfbp-2 has a predicted mr of 31,325. ch ...19892479552
multiple t and b cell epitopes in the s1 subunit ("a"-monomer) of the pertussis toxin molecule.the immunogenicity and reactogenicity of bordetella pertussis vaccine are mediated in part by the s1 subunit of pertussis toxin (pt). to identify the immune epitopes in the s1 subunit of pt, synthetic peptides were prepared and tested for their capacity to induce antibodies in mice with different mhc genotypes. in balb/c mice, peptides corresponding to sequences 1-17, 70-82 and 189-199 generate t cell proliferative responses, induce the production of antibodies capable of neutralization of the t ...19892480389
site-directed mutagenesis of the human chorionic gonadotropin beta-subunit: bioactivity of a heterologous hormone, bovine alpha-human des-(122-145)beta.human cg contains an alpha-subunit, common to the pituitary glycoprotein hormones, and a hormone-specific beta-subunit, but unlike the pituitary beta-subunits, hcg beta is characterized by an o-glycosylated carboxy-terminal extension. a mutant beta-subunit, des-(122-145)hcg beta, was prepared using site-directed mutagenesis, and the prsv expression plasmids were transfected into chinese hamster ovary cells that produce the bovine alpha-subunit (b alpha). the mutant beta-subunit binds to b alpha, ...19892481812
expression of mitogenically active human recombinant platelet-derived growth factor a-chain.expression vectors encoding cdnas for the human platelet-derived growth factor (pdgf) a-chain (pjkl) and murine dihydrofolate reductase (ptkdhfr) were cotransfected into dihydrofolate reductase-deficient chinese hamster ovary cells. methotrexate-induced coamplification of clones, expressing pdgf a-chain resulted in enhanced levels of a-chain-specific dna, rna and protein. a 30,500 mr protein was immunoprecipitated with pdgf antisera from the conditioned media of metabolically labeled cells. redu ...19892482405
mechanism of comutagenesis of sodium arsenite with n-methyl-n-nitrosourea.arsenic compounds are known carcinogens. although many carcinogens are also mutagens, we have previously shown that sodium arsenite is not mutagenic at either the na+/k+ atpase or hprt locus in chinese hamster v79 cells. it can, however, enhance uv-mutagenesis. we now confirm the nonmutagenicity of sodium arsenite in line g12, a psv2gpt-transformed v79 (hprt-) cell line, which is able to detect multilocus deletions in addition to point mutations and small deletions. the lack of arsenic mutagenic ...19892484616
on the mechanism of the comutagenic effect of cu(ii) with ultraviolet light.although cucl2 alone is not mutagenic in e. coli or in chinese hamster cells, exposure of e. coli to cucl2 during uv-irradiation causes enhancement of uv-mutagenesis. the mechanism for this comutagenic effect appears to be owing to increased dna damage by the combined treatment of uv and cu(ii) compared with uv or cu(ii) alone. using a sequencing gel approach, uv alone is found to cause a particular pattern of alkali-labile sites, whereas cucl2 alone caused few such sites. the combined action of ...19892484617
soluble vs insoluble hexavalent chromate. relationship of mutation to in vitro transformation and particle uptake.soluble cacro4 and insoluble pbcro4 were tested for induction of mutation to 6-thioguanine (base-substitution, deletion, addition, and frameshift mutations) or ouabain (base-substitution mutations) resistance in chinese hamster ovary cells and morphological transformation in c3h/10(1/2) mouse embryo cells. cacro4 induced dose-dependent cytotoxicity and mutation to 6-thioguanine resistance, but did not induce mutation to ouabain resistance or morphological transformation. highly cytotoxic amounts ...19892484629
biological and binding studies of acidic fibroblast growth factor in the presence of substituted dextran.heparin has been shown to interact with acidic fibroblast growth factor (afgf) and to potentiate the biological activity of afgf on fibroblastic cells. water-soluble dextran substituted with methyl carboxylic benzylamine and sulfonate groups has been shown to mimic the effect of heparin in its anticoagulant and anticomplement activity. we have studied the effect of a dextran derivative named e (dde), which had an anticoagulant activity equivalent to 0.5 iu heparin/mg, on the mitogenic activity o ...19892484952
deletion formation in mammalian cells: molecular analysis of breakpoints and junctions in the hamster aprt locus.to examine the mechanisms governing deletion formation in mammalian cells, we have analyzed the breakpoints and junction fragments produced by seven such mutations at the aprt locus of chinese hamster ovary cells at the base sequence level. the deletions were heterogeneous both in size, varying from 38 bp to 170 kb, and in sequence in that no recurring sequence or structural motifs were evident. most were simple exchanges at overlapping di- or trinucleotides, but one was the result of a complex ...19892488705
gene inactivation as a mechanism for the expression of recessive phenotypes.a series of chinese hamster ovary cell hybrids were constructed which were heterozygous at the emtb and chr loci. these loci encode two recessive drug-resistance genes (emetine resistance and chromate resistance, respectively) located on a structurally hemizygous region on the long arm of chromosome 2. these heterozygous hybrids therefore exhibit wild-type sensitivity to both emetine and chromate. drug-resistant variants were then selected in medium containing either emetine or chromate, and the ...19892491018
use of the cytokinesis-block method for the analysis of micronuclei in v79 chinese hamster lung cells: results with mitomycin c and cyclophosphamide.the cytochalasin b (cyb)-blocked binucleated cell assay has been explored to analyze micronuclei and cell cycle kinetics using 2 known mutagenic carcinogens in v79 chinese hamster lung cells. to determine the optimum time to obtain the maximum number of binucleated cells for micronucleus analysis, duplicate cultures of exponentially growing cells were treated with 3 micrograms/ml cyb for varying durations (8-48 h). a peak appearance of binucleated cells at 16 h in the presence of cyb suggested t ...19892492079
expression of biologically active human follitropin in chinese hamster ovary cells.to study the structure-function relationships of follitropin (fsh), we expressed the hormone in a heterologous cell system. a genomic clone bearing a 3.7-kilobase fsh beta insert containing the entire coding sequence was transfected alone or together with the alpha subunit gene into chinese hamster ovary cells and stable lines expressing either fsh beta or fsh dimer were selected. pulse-chase experiments revealed that, when transfected alone fsh beta was very slowly secreted similar to lutropin ...19892494176
molecular analysis of spontaneous mutations at the gpt locus in chinese hamster ovary (as52) cells.as52 cells are chinese hamster ovary (cho) cells that carry a single functional copy of the bacterial gpt gene and allow the isolation of 6-thioguanine-resistant (6tgr)mutants arising from mutation at the chromosally integrated gpt locus. the gpt locus in as52 cells is extremely stable, giving rise to 6tgr mutants at frequencies comparable to the endogenous cho hprt locus. in this study, we describe the spectrum of spontaneous mutations observed in as52 cells by southern blot and dna sequence an ...19892494446
construction of a tissue plasminogen activator gene having unique restriction sites to facilitate domain manipulation: a model for the analysis of multi-domain proteins.we have designed and constructed a dna sequence encoding human tissue plasminogen activator (tpa) with convenient restriction sites that flank each of the domains of the heavy chain. to accomplish this, the first 1095 bases of the gene coding for the mature protein were synthesized with unique restriction sites engineered into the interdomainal regions. this synthetic construction was then ligated to a cdna fragment of the tpa gene that encoded the active site, thus generating a full-length tpa ...19892494653
glycosylation of human apolipoprotein e. the carbohydrate attachment site is threonine 194.the glycosylation of human apolipoprotein (apo) e was examined with purified plasma apoe and apoe produced by transfected cell lines. the carbohydrate attachment site of plasma apoe was localized to a single tryptic peptide (residues 192-206). sequence analysis and amino sugar analysis of this peptide derived from asialo-, monosialo-, or disialo-apoe indicated that the carbohydrate moiety is attached only to thr194 in monosialo- and disialo-apoe and that asialo-apoe is not glycosylated. mammalia ...19892498325
expression of apoe gene in chinese hamster cells with a reversible defect in o-glycosylation. glycosylation is not required for apoe secretion.the effects of o-glycosylation on the synthesis and secretion of apolipoprotein e (apoe, a glycoprotein with o- but not n-linked sugars) were studied with a udp-galactose/udp-n-acetylgalactosamine 4-epimerase-deficient cell mutant (ldld cells) which expresses a reversible defect in protein o-glycosylation. under normal culture conditions the mutant ldld cells cannot add n-acetylgalactosamine (galnac) to proteins. galnac is the first sugar of mucin-type o-linked oligosaccharides attached to the p ...19892498327
effect of von willebrand factor coexpression on the synthesis and secretion of factor viii in chinese hamster ovary cells.in plasma, antihemophilic factor (factor viii) exists as a 200-kilodalton heavy-chain polypeptide in a metal ion association with an 80-kilodalton light-chain polypeptide. this complex is bound by hydrophobic and hydrophilic interactions to a large multimeric glycoprotein, von willebrand factor (vwf). accumulation of secreted human factor viii activity expressed in chinese hamster ovary cells requires the addition of serum in the growth medium, which provides vwf. here we report that coexpressio ...19892498645
inhibition of transient gene expression in chinese hamster ovary cells by cyclobutane dimers and (6-4) photoproducts in transfected ultraviolet-irradiated plasmid dna.using a transient gene expression assay to measure host cell reactivation, the effects of cyclobutane dimer and noncyclobutane dimer uv photoproducts on expression of a reporter gene were examined in normal and repair-deficient chinese hamster ovary (cho) cell lines. ultraviolet damage in plasmid prsv beta gal dna, containing the escherichia coli beta-galactosidase gene, resulted in reduced reporter gene expression in both uv-hypersensitive mutant cho cell lines uv5 and uv61 relative to wild-typ ...19892498916
cytogenetical characterization of uv-sensitive repair-deficient cho cell line 43-3b. ii. induction of cell killing, chromosomal aberrations and sister-chromatid exchanges by 4nqo, mono- and bi-functional alkylating agents.an established cell line of chinese hamster ovary (cho-9) cells and its uv-sensitive mutant 43-3b have been studied for the induction of cell killing, chromosomal aberrations and sister-chromatid exchanges (sces) after exposure to different types of dna-damaging agents such as 4-nitroquinoline-1-oxide (4nqo), mitomycin c (mmc), diepoxybutane (deb), methyl methanesulfonate (mms), ethyl methanesulfonate (ems) and ethyl nitrosourea (enu). in comparison with the wild-type cho cells, 43-3b cells show ...19892499773
cytogenetical characterisation of chinese hamster 43-3b transferants with the amplified or non-amplified human dna repair gene ercc-1.a comparative study on the biological responses to different mutagens (uv, 4nqo, mmc, mms and ems) was made on cho wild-type cells (cho-9), its uv-hypersensitive mutant 43-3b, and 2 types of its transferants, i.e., one containing a few copies of the human repair gene ercc-1 and the other having more than 100 copies of ercc-1 (due to gene amplification). cell survival, chromosomal aberrations and sces were used as biological end-points. the spontaneous frequency of chromosomal aberrations in the ...19892499774
enhancing effects of cinoxate and methyl sinapate on the frequencies of sister-chromatid exchanges and chromosome aberrations in cultured mammalian cells.sister-chromatid exchanges (sces) induced by mitomycin c (mmc), 4-nitroquinoline-1-oxide (4nqo) or uv-light in cultured chinese hamster ovary cells (cho k-1 cells) were enhanced by cinoxate (2-ethoxyethyl p-methoxycinnamate) or methyl sinapate (methyl 3,5-dimethoxy 4-hydroxycinnamate). both substances are cinnamate derivatives and cinoxate is commonly used as a cosmetic uv absorber. methyl sinapate also increased the frequency of cells with chromosome aberrations in the cho k-1 cells treated wit ...19892499777
sodium fluoride-induced chromosome aberrations in different stages of the cell cycle: a proposed mechanism.in an attempt to clarify the controversy about sodium fluoride (naf) clastogenicity, the induction of chromosome aberrations in chinese hamster ovary cells (cho) by naf was investigated. following a protocol used for screening chemicals for clastogenic activity, significant increases of aberrant cells were observed when cells were exposed to naf for 4 h and harvested 8 h later. cell-cycle kinetic studies demonstrated most cells were exposed in g2 of the cell cycle. smaller increases in aberrant ...19892500593
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