Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “KB Cells”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 271 records · Page 15Linked to original sources
PubMed1985

Ultrastructural immunocytochemical localization of the transferrin receptor using a monoclonal antibody in human KB cells.

Using a monoclonal antibody (HB21) against the human transferrin receptor, we have localized this receptor in cultured KB human carcinoma cells by fluorescence and ultrastructural immunocytochemistry. The receptor was found diffusely distributed on the cell surface, concentrated in clathrin-coated pits of the cell surface, in intracellular endocytic vesicles (receptosomes) derived from coated pits, in tubular elements of the trans-reticular Golgi system, and in microtubule-associated membranous elements thought to be part of the constitutive exocytic system. This distribution is the same as that previously shown for labeled transferrin in these same cells (Willingham MC, Hanover JA, Dickson BB, Pastan J: Proc Natl Acad Sci USA 81:175, 1984). No significant amounts of receptor were found in lysosomes. An aggregation of membranous elements containing this receptor was found in the pericentriolar region of cells during mitosis. Together with the previous data on the immunocytochemical localization of transferrin, these results suggest that the transferrin receptor may constitutively enter and exit KB cells by endocytosis and exocytosis, carrying bound transferrin into and out of the cell for the purpose of supplying iron from the extracellular environment for cell growth.

Antibodies, Monoclonal↗
PubMed1992

Circumvention of multidrug resistance in human carcinoma KB cells by polyether antibiotics.

We examined the effect of various polyether antibiotics on colchicine resistance in multidrug-resistant KB-C4 cells which exhibit about 4,000-fold resistance to colchicine. As a result, 4 out of 14 polyether antibiotics were found to reverse colchicine resistance. Among them, laidlomycin was the most potent. It potentiated colchicine cytotoxicity on KB-C4 cells about 700-fold at 1 microgram/ml. Degree of potentiation was calculated by dividing of the IC50 value of colchicine in the absence of a polyether antibiotic by the IC50 value of colchicine in the presence of the polyether antibiotic. Monensin, dianemycin, and leuseramycin at 3 micrograms/ml also potentiated the cytotoxicity, about 100-fold. We previously reported that inostamycin is a potent chemosensitizer in KB-C4 cells. Although lysocellin has a structure very similar to that of inostamycin, it didn't reverse colchicine resistance. It slightly increased [3H]vinblastine accumulation in KB-C4 cells and weakly inhibited the [3H]vinblastine binding to KB-C4 plasma membranes.

Anti-Bacterial Agents↗
PubMed1988

Stability and covalent modification of P-glycoprotein in multidrug-resistant KB cells.

An antipeptide antibody (P7) to P-glycoprotein has been produced by immunizing rabbits with a synthetic peptide. Antibody P7 is directed against the amino-terminal region of P170 (residues 28-35). The antibody immunoprecipitates a 170-kDa P-glycoprotein from extracts of drug-resistant KB-V1 cells that is not present in the drug-sensitive cell line KB-3-1. Antibody P7 was used to quantitate the amount of P-glycoprotein present in drug-resistant KB lines at various levels of resistance and to demonstrate the presence of P-glycoprotein in NIH 3T3 cells transfected with a cloned MDR1 cDNA or human genomic DNA encoding MDR1. Pulse-chase labeling experiments demonstrated that P-glycoprotein is synthesized as a 140-kDa precursor which is slowly converted over 2-4 h to a 170-kDa glycoprotein. Tunicamycin treatment blocks the conversion of the precursor to the mature form, and removal of N-linked oligosaccharides with Endo F reduces the relative molecular weight of P-glycoprotein to 140K. The mobility of mature P-glycoprotein is unaffected by treatment with neuraminidase and Endo H. These data indicate that P-glycoprotein is N-glycosylated and contains little or no neuraminic acid. P-Glycoprotein is also phosphorylated, and the extent of phosphate incorporated is proportional to the amount of protein present in drug-resistant cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1970

Transfer ribonucleic acid in KB cells infected with adenovirus type 2.

Populations of transfer ribonucleic acid (tRNA) extracted from control and type 2 adenovirus (Ad2)-infected KB cells were compared. No consistent differences in acceptor activity for 11 amino acids were observed. Comparison of methylated albumin-kieselguhr (MAK) elution profiles of arginyl-tRNA from control and infected cells revealed a minor modification in that the proportion of arginyl-tRNA eluting at high salt concentration was somewhat greater in infected cells. No similar differences were observed in MAK elution profiles of aspartyl-, isoleucyl-, leucyl-, phenylalanyl-, seryl-, tyrosyl-, and valyl-tRNA. Hybridization of 4S RNA from infected cells labeled by incorporation of (3)H-uridine with Ad2 deoxyribonucleic acid revealed the presence of a complementary species of RNA in this preparation. Hybridization of (3)H-arginyl-tRNA and of (3)H-aminoacyl-tRNA labeled by charging with (3)H-arginine or a (3)H-mixture of amino acids, respectively, failed to detect the presence of virus-specific tRNA in Ad2-infected cells.

Adenoviridae↗
PubMed1978

The deoxyribonuclease induced after infection of KB cells by herpes simplex virus type 1 or type 2. I. Purification and characterization of the enzyme.

The deoxyribonuclease induced in KB cells by herpes simplex virus (HSV) type 1 and type 2 has been purified. Both enzymes are able to completely degrade single- and double-stranded DNA yielding 5'-monophosphonucleotides as the sole products. A divalent cation, either Mg2+ or Mn2+, is an absolute requirement for catalysis and a reducing agent is necessary for enzyme stability. The maximum rate of reaction is achieved with 5 mM MgCl2 for both HSV-1 and HSV-2 DNase. The optimum concentration for Mn2+ is 0.1 to 0.2 mM and no exonuclease activity is observed when the concentration of Mn2+ is greater than 1 mM. The rate of reaction at the optimal Mg2+ concentration is 3- to 5-fold greater than that at the optimal Mn2+ concentration. In the presence of Mg2+, the enzymes are inhibited upon the addition of Mn2+, Ca2+, and Zn2+. The enzymatic reaction is also inhibited by spermine and spermidine, but not by putrescine. Crude and purified HSV-1 and HSV-2 DNase can degrade both HSV-1 and HSV-2 DNA, but native HSV-1 DNA is hydrolyzed at only 22% of the rate and HSV-2 DNA at only 32% of the rate of Escherichia coli DNA. Although HSV-1 and HSV-2 DNase were similar, minor differences were observed in most other properties such as pH optimum, inhibition by high ionic strength, activation energy, and sedimentation coefficient. However, the enzymes differ immunologically.

Cell Line↗
PubMedDate not supplied

The effects of N-methyl-N'-nitro-N-nitrosoguanidine on KB cells. I. Growth inhibition and cell killing in relation to inhibition of macromolecular synthesis.

Cytotoxic effects of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) on KB cells were analyzed taking into account cell killing and growth inhibition. Attempts to correlate these effects with macromolecular synthesis inhibition were monitored. At low doses (1 X 10(-6) at 1 X 10(-5) M), MNNG acting for 1 h only inhibited cell proliferation in a dose dependent manner and specifically inhibited DNA synthesis. At intermediate doses (1.6 X 10(-5) at 3.2 X 10(-5) M), treatment resulted in detachment and death of the cells during a 20-h post-incubation period. This effect was related to a dose-dependent inhibition of RNA and protein synthesis. At high doses (greater than 3.2 X 10(-5) M) MNNG treatment resulted in cell killing which was distinct from that produced by lower doses, since the cells did not detach from the glass. At these high doses strong inhibition of RNA and protein synthesis was observed early. When cells prelabelled with radioactive uridine or amino acids were treated with MNNG, the release of labelling into the culture medium increased, especially with uridine prelabelled cells. This increase was very high with the intermediate doses of the drug, but more modest with the strong doses. Furthermore, with the moderate doses, cell size showed greater reduction compared to the control than with the high doses. Thus, cell size reduction and loss of cellular material varied parallel to cell detachment. We hypothesize that moderate doses of MNNG may induce cellular degradation, whereas high doses may prevent it. The significance of this finding is discussed.

Carcinoma, Squamous Cell↗
PubMed1980

Structural and enzymological characterization of a deoxyribonucleic acid dependent adenosine triphosphatase from KB cell nuclei.

We have purified to near homogeneity the single DNA-dependent ATPase activity that we have identified in extracts of KB cell nuclei. The protein structure of the enzyme was defined by sodium dodecyl sulfate gel electrophoresis, which revealed a single protein band of 75000 daltons that was coincident with the profile of ATPase activity resolved by the final step of agarose-ATP chromatography or by isoelectric focusing. The enzyme has a pI of 8.5, a Stokes' radius by gel filtration of 3.8 nm, and a sedimentation coefficient in high salt of 5.3 S. At low ionic strength the enzyme activity sediments at 7.0 S, suggesting that it may dimerize under these conditions. The purified enzyme has a specific activity of 5.9 X 10(5) nmol of ATP hydrolyzed per h per mg of protein and is devoid of endonuclease, exonuclease, RNA or DNA polymerase, nicking-closing, and gyrase activities at exclusion limits of 10(-6)-10(-8) of the ATPase activity. The enzyme can hydrolyze only ATP or dATP, to generate ADP or dADP plus Pi, but the other NTPs and dNTPs are competitive inhibitors of the enzyme with respect to ATP. A divalent cation (Mg2+ greater than Mn2+ greater than Ca2+) as well as a nucleic acid cofactor is required for activity. Single-stranded DNA or deoxyhomopolymers are most effective, but blunt-ended linear and nicked circular duplex DNA molecules are also used at Vmax values approximately 20% of that obtained with single-stranded DNA. Intact duplex DNA and polyribonucleotides are unable to support ATP hydrolysis. Velocity gradient sedimentation studies corroborate the interpretations of the kinetic analyses and demonstrate enzyme binding to single-stranded DNA and nicked duplex DNA but not to intact duplex DNA. Although we have not succeeded directly in demonstrating DNA unwinding by this protein, preliminary results suggest that in the presence of ATP, the ATPase can stimulate the reactivity of homogeneous human DNA polymerases alpha and beta on nicked duplex DNA substrates.

Adenosine Triphosphatases↗
PubMed1984

DNA primase from KB cells. Characterization of a primase activity tightly associated with immunoaffinity-purified DNA polymerase-alpha.

A very highly purified fraction of KB cell DNA polymerase-alpha, prepared with a monoclonal antibody, contains DNA primase activity. The primase synthesizes oligonucleotide chains initiated with ATP in a reaction that is resistant to alpha-amanitin and strictly dependent on added template and ribonucleoside triphosphates (rNTPs). In the presence of added dNTPs and M13 DNA template, the primase produces a uniform population of oligoribonucleotides, predominantly hexamers to decamers, that are extended by polymerase-alpha into DNA chains up to 3000 nucleotides long. There is no evidence for nucleotide preferences at RNA/DNA junctions. In the absence of added dNTPs, the oligomeric products are heterogeneous in size and composition and susceptible to cleavage by pancreatic DNase I due to their content of short oligodeoxynucleotide tracts synthesized by primase from trace contaminant dNTPs in the rNTP substrates. The primase and polymerase-alpha activities are distinguishable by several physical and chemical criteria, and the primase reaction is only partially sensitive to two potent, independent monoclonal antibodies that neutralize polymerase-alpha. Although the presence of both primase and polymerase-alpha activities in a highly purified immune complex prepared with a monoclonal antibody argues for their tight physical association, the chemical, physical, and immunological discriminations indicate the two catalytic entities are functionally and structurally distinct.

Carcinoma↗
PubMed1988

Combined modalities of resistance in etoposide-resistant human KB cell lines.

The alkaloid derivative 4'-demethylepipodophyllotoxin 9-(4,6-O-ethylidene)-beta-D-glucopyranoside (etoposide, VP-16) is believed to exert cytotoxicity by causing double-stranded DNA breaks through interruption of the breaking-resealing reaction of topoisomerase II (topo II). Thus it was conceivable that cells could become resistant to VP-16 by a decrease in topo II enzyme level, since this would lead to fewer DNA breaks. As well, given the structure of VP-16, it was also possible that a pleiotropic mechanism of resistance could decrease sensitivity to this drug. To study these possibilities, a series of VP-16-resistant human KB cell lines was established by stepwise selection. The concentrations of VP-16 required to inhibit cell proliferation by 50% in the parent line and KB/1c, KB/7d, KB/20a, and KB/40a lines were, respectively, 0.16, 4.7, 24, 31, and 47 microM. These cell lines expressed cross-resistance to 4'-(9-acridinylamino)methanesulfon-m-anisidide, doxorubicin, vincristine, and methotrexate, although the pattern of relative drug sensitivity was quite different from that of pleiotropic resistant cell lines reported elsewhere. The resistance to vincristine and methotrexate did not increase above the level of the KB/1c cells, and resistance to VP-16, doxorubicin, and especially vincristine was unstable in VP-16-resistant cells cultured in the absence of drug. Although the drug resistance marker Mr 180,000 glycoprotein could not be detected in any of our cell lines, cellular accumulation of [3H]VP-16 was reduced 50-75% in the resistant lines compared with parent KB. With increasing VP-16 resistance, the level of topo II protein, detected by antibody staining, decreased at each step of selection, concomitant with a general decrease in topo II unknotting activity. Sensitivity of the topo II unknotting assay to inhibition by VP-16 was the same for the parent and all resistant cell lines. The level of topo I activity and enzyme increased slightly in the resistant cells. Thus, these cell lines are resistant to VP-16 by virtue of at least two mechanisms: (a) reduced levels of topo II, which confers cross-resistance to other compounds which are topo II-dependent cytotoxic agents; and (b) reduced accumulation of drug, which is likely also responsible for vincristine and methotrexate resistance. However, the possible existence of other mechanisms of resistance cannot be ruled out.

DNA Damage↗
PubMedDate not supplied

The effect of human alpha leukocyte interferon (IFN) on the frequency of sister chromatid exchanges (SCE) in the KB cell line.

Concentrations of 50, 500 and 5,000 iu/ml of natural human alpha leukocyte interferon (IFN) were added into the culture medium of KB cells, 4 h after serial passage, in the presence of 5-bromo-desoxyuridine (Brdu) at 10 micrograms/ml. Similar cultures without IFN were set up as controls. After 72 h of incubation, the harlequin technique (differential staining of sister chromatids) was applied in order to discriminate among the metaphases of different generations and to appreciate the frequency of sister chromatid exchanges (SCE) in the second generation cells. The incidence of sister chromatid exchanges was slightly increased following IFN treatment but no dose-effect relationships were observed. At the same time, cell cycle kinetics estimated as replication index (RI) and average generation time (AGT) was not modified in IFN-treated cells as against the controls.

Cell Division↗
PubMed1992

Folate-binding protein mRNA is decreased in methotrexate-resistant KB cells.

Folate-binding protein (FBP), a high-affinity folate receptor expressed in certain malignant and normal cell lines and tissues, is responsible for cellular transport of folate and structurally related antifolates such as methotrexate in human KB (nasopharyngeal carcinoma) cells. We have developed a sensitive polymerase chain reaction (PCR) assay to quantitate folate-binding protein mRNA levels. This assay was tested in KB and methotrexate (MTX)-resistant KB (KB1BT) cell lines. Based on assay results, we conclude there are 245 molecules of folate-binding protein (FBP) mRNA per KB cell, and there is an approximately 3-fold decrease in FBP mRNA levels per KB1BT cell. The data obtained by quantitative PCR correlate well with northern blot analysis of mRNA levels and membrane-associated folate-binding protein (M-FBP) levels. Moreover, when KB1BT cells were grown in the absence of MTX for 8 months, the levels of FBP and M-FBP mRNA did not change, and there was a substantial decrease in MTX uptake, compared with KB cells.

Base Sequence↗
PubMed1983

Characterization of a phosphoprotein associated with the SS-B/La nuclear antigen in adenovirus-infected and uninfected KB cells.

We have employed sera from patients with autoimmune disease to characterize the nuclear SS-B/La antigen in uninfected and adenovirus-infected KB cells. A 45,000-dalton phosphorylated polypeptide was specifically precipitated with anti-SS-B sera, and the level of phosphorylation was increased after infection. The increased phosphorylation appears to occur at the same amino acid residues phosphorylated in uninfected cells and results from increased phosphorylating activity rather than from altered enzyme specificity. A competition experiment between infected and uninfected cell extracts indicates that the antigen in the infected cell binds more strongly to SS-B/La antibodies. Fragments of adenovirus-induced virus-associated RNA as well as intact molecules complex with SS-B/La antigen and are immune precipitated with autoimmune sera.

Adenoviridae↗
PubMed1991

The role of glycosylation in the biosynthesis and acquisition of ligand-binding activity of the folate-binding protein in cultured KB cells.

The biosynthesis, processing, and ligand-binding function of the membrane-associated and soluble forms of the folate-binding protein (FBP) in KB cells, a cultured human cell line, were studied using pulse-chase labeling with [35S] methionine. The intermediary and mature forms of the protein were isolated by immunoprecipitation and affinity chromatography and analyzed by sodium dodecyl sulfate electrophoresis and autoradiography. The earliest species identified had an Mr of 32 Kd and disappeared over 5 hours concomitant with the appearance of a 38-Kd cellular FBP. As the 38-Kd species disappeared, a 40-Kd form appeared in the medium. When tunicamycin was added to the culture medium to inhibit core glycosylation, a 26-Kd aglycosylated species and minor 28-Kd and 30-Kd forms appeared. Endoglycosidase H, which cleaves high mannose but not complex oligosaccharides, reduced the 32-Kd species to 26-Kd but the enzyme had no effect on the 38-Kd form, indicating that this species is complex glycosylated. Monensin, which blocks complex glycosylation, also inhibited synthesis of the 38-Kd species. Although both the 32-Kd and 38-Kd forms had ligand-binding sites (as demonstrated by binding to a folate-Sepharose matrix), the 26-Kd aglycosylated species, labeled in the presence of tunicamycin, lacked similar binding sites because it did not bind to the affinity matrix. In contrast, the aglycosylated 26-Kd form, which was obtained by treatment of the 32-Kd species with endoglycosidase H, did bind to the folate affinity matrix, indicating that it retained ligand-binding function. Thus, the high mannose oligosaccharide moiety is not required for the folate-binding property of the FBP, but its addition to the polypeptide chain precedes a later step that is necessary for the mature protein to have ligand-binding function.

Acetylglucosaminidase↗
PubMed1979

Enzymological characterization of DNA polymerase alpha. Basic catalytic properties processivity, and gap utilization of the homogeneous enzyme from human KB cells.

This report describes the results of our initial enzymological characterization of a homogeneous preparation of DNA polymerase alpha that we have purified from cultured human KB cells. Although the enzyme is most reactive with duplex DNA substrates that contain short gaps (optimally activated) in incubations that require Mg2+, the polymerase possesses the intrinsic capacity to copy the initiated ribohomopolymer template, (A)-n, (dT)-200, at low rates in the presence of Mn2+. Because of the preponderance of DNA polymerase alpha in actively multiplying vertebrate cells, it is probable that this low level of activity comprises the majority of the ribopolymer copying activity that can be detected in crude tissue extracts. The presence of contaminating or associated deoxyribonuclease activities can be excluded from the purified enzyme to levels of 10(-4) to 10(-7) of the polymerase activity. The mechanism of polymerization on activated DNA under optimum conditions is moderately processive, with 11 +/- 5 nucleotides incorporated per polymerization cycle. The polymerase is unable to work at nicks or at short gaps of approximately 20 to 30 nucleotides in length, and it measures a surprisingly invariant effective template length on optimally activated DNA and on DNA molecules that have been gapped to varying extents with Escherichia coli exonuclease III. In the "Appendix" we present an amplification of the theoretical formulation of Bambara et al. (Bambara, R. A., Uyemura, D., and Choi, T. (1978) J. Biol. Chem. 253, 413--423) that permits the use of DNA polymerases with significant associated 3' leads to 5'-exonuclease activities for the accurate measurement of average template lengths (gap sizes) and titration of usable 3'-hydroxyl primer termini in gapped, duplex DNA substrates.

Binding, Competitive↗
PubMed1977

DNA polymerase-alpha. Purification and structural characterization of the near homogeneous enzyme from human KB cells.

In this report we describe the purification and structural characterization of a near homogeneous preparation of DNA polymerase-alpha that we have obtained from cultured human KB cells. When analyzed by nondenaturing gel electrophoresis, velocity gradient centrifugation, and isoelectric focusing, the enzyme activity demonstrates a constancy of gel mobility, sedimentation coefficient, and isoelectric point during the final three chromatographic steps of the purification. Native gel electrophoresis of the penultimate polymerase fraction at seven concentrations of acrylamide reveals co-migration of the enzyme activity with a single discernible protein band, at constant specific activity, and indicates that the polymerase protein is electrophoretically homogeneous at this stage. The purified enzyme has a sedimentation coefficient of 7.1 to 7.2 S at high or low ionic strength, a molecular weight gel filtration of 149,000, and an isoelectric point of pH 5.0 to 5.2. Sodium dodecyl sulfate gel analyses demonstrate that the polymerase-alpha molecule is a dimer comprised of two dissimilar subunits of 76,000 and 66,000 daltons that are present in equimolar ratio.

Cells, Cultured↗
PubMed1976

[Compared cytologic study of (in situ and isolated) metaphasic chromosomes from KB cells, after incorporation of 5 bromodeoxyuridine (5 BrdU)].

Substitution of thymidine by 5 BrdU indduces the same morphological and cytological changes in in situ and isolated metaphase chromosomes of KB cells: a dichroïc fluorescence and a difference in the affinity of each chromatid for Giemsa stain. However, the precursor does not seem to modify the appearance of G or Q bands in in situ and isolated chromosomes. The fibrilar unit which can be seen with electronic microscopy, shows no difference before and after 5-BrdU incorporation.

Bromodeoxyuridine↗