Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The activation of cyclic AMP-dependent protein kinase has been found to be the predominant mode by which cyclic AMP (cAMP) leads to alterations of a large variety of cellular functions. The activation of the kinase results in the release of the catalytic subunit which as the free enzyme possesses phosphotransferase activity for a variety of specific protein substrates. Using a sensitive and specific cytofluorometric technique we monitored the appearance of free catalytic subunit in Reuber H35 hepatoma cells in culture after incubation with N6-1'-O-dibutyryl-cyclic AMP (DBcAMP), 8-bromoadenosine-3':5'-cyclic monophosphate (8-BrcAMP), and glucagon. The cytochemical method employs the heat-stable inhibitor of the free catalytic subunit which has been conjugated to fluorescein isothiocyanate (F:PKI) and was validated as described in the companion paper (Fletcher and Byus. 1982. J. Cell Biol. 93:719-726). Here we studied the temporal and spatial kinetics of the free catalytic subunit following activation of cAMP-dependent protein kinase by increasing concentrations of DBcAMP,8-BrcAMP, and glucagon. Under similar conditions protein kinase activation was also assessed biochemically in H35 cell supernatants by assaying the protein kinase activity ratio. Incubation of the hepatoma cells with DBcAMP (0.1 mM) led to an increase in the activity ratio from 0.2 in control cultures to a value of nearly 1.0 within a 1- to 2-h period. During this same period using the F:PKI probe, a significant increase in cytoplasmic and nucleolar fluorescence indicative of the release of the free catalytic subunit was coincidentally observed. In contrast to the rapid appearance of catalytic subunit in the cytoplasm and nucleolus of the cell within 5-15 min of the addition of DBcAMP, discernible nucleoplasmic fluorescence did not occur until after 1 h. H35 cell cultures incubated with 8-BrcAMP (0.01-1.0 mM) exhibited a more rapid activation of the protein kinase measured cytochemically compared to the cells treated with DBcAMP. Cultures incubated with 8-BrcAMP had significantly increased cytoplasmic and nucleolar fluorescence compared to unstimulated cells within 1 min of the addition of the analogue and reached a maximal level within 15 min. By employing a microspectrophotometer a distinct dose-dependent increase in cellular fluorescence (i.e., free catalytic subunit) was observed as the concentration of 8-BrcAMP was increased from 0.01 to 1.0 mM at 1, 5, 15, and 60 min following stimulation. The addition of glucagon (10(-6) M) to the culture also led to the activation of cAMP-dependent protein kinase as determined by an increase in the activity ratio. This increase was paralleled throughout the incubation period by a marked elevation in cytoplasmic and nucleolar fluorescence. The results reported herein suggest that both cyclic nucleotide analogues and a polypeptide hormone lead to the activation of cAMP-dependent protein kinase in similar intracellular compartments in Reuber H35 hepatoma cells...
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PMID:Direct cytochemical localization of catalytic subunits dissociated from cAMP-dependent protein kinase in Reuber H-35 hepatoma cells. II. Temporal and spatial kinetics. 628 33

A cyclic nucleotide-independent, polyamine-responsive protein kinase from the cytosol of Morris hepatoma 3924A, which phosphorylated heat-stable endogenous substrates and casein in the presence of polyamines (Criss, W.E., Yamamoto, M., Takai, Y., Nishizuka, Y. and Morris, H.P. (1978) Cancer Res. 38, 3540-3545) was observed to be stimulated by an endogenous protein activator. This protein activator was identified to be calmodulin. the polyamine-responsive protein kinase was also stimulated by purified calmodulin, but only in the presence of polyamines such as polylysine. This action of calmodulin did not require Ca2+ for activation of the enzyme; and activation occurred in the presence of EGTA. DNA and RNA inhibited the polyamine-responsive protein kinase, either in the presence or absence of Ca2+. Purified calmodulin, in the presence of cyclic AMP or cyclic GMP, did not activate the protein kinase. Therefore, polyamines such as polylysine are an absolute requirement for this expression of calmodulin action. The increased enzyme activity by calmodulin was accompanied with an increased Vmax and with no changes in the Km (ATP). High levels of cation, up to 100 mM Mg2+, did not effect the action of calmodulin. These results indicate that tumor cytosolic polyamine-responsive protein kinase is regulated by calmodulin, the latter being increased in the tumor tissue.
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PMID:Calmodulin stimulates polyamine-responsive protein kinase in the absence of Ca2+. 629 10

The purified Novikoff hepatoma nuclear phosphoprotein with a molecular weight of 110 kdalton and pI 8.4, was found to be a type I topoisomerase. When isolated from 32P-labeled Novikoff ascites cells or incubated in vitro with protein kinase, phosphoserine was found to be its major phosphorylated amino acid. The enzymatic activity of topoisomerase I was altered by changes in phosphorylation. Its activity was increased by protein kinase and it was decreased by alkaline phosphatase.
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PMID:Phosphorylation of purified Novikoff hepatoma topoisomerase I. 630 89

The ability of the phorbol ester tumor promoter 12-O-tetradecanoylphorbol- 13-acetate (TPA) to lead to the activation of cyclic adenosine 3':5'-monophosphate (cAMP)-dependent protein kinase was investigated in three cell lines. In the Reuber H35 hepatoma cells, TPA (1.6 microM) led to an increase in the protein kinase activity ratio from a value of 0.13 in control cultures to 0.40 in the stimulated cells within 30 min of addition. A specific fluorescent cytochemical procedure was further utilized to study the intracellular localization of the free catalytic subunit following activation of the holoenzyme by TPA. A marked increase in cytoplasmic and nucleolar fluorescence indicative of the appearance of the free catalytic subunit (activation of cAMP-dependent protein kinase) in these cellular compartments was observed in both Reuber H35 hepatoma and Chinese hamster ovary 10001 cells incubated with TPA (1.6 microM) for 30 min. In the H35 cells monitored with the cytochemical probe for the catalytic subunit, protein kinase was activated within 5 min of the addition of TPA (1.6 microM) with 0.16 microM TPA in the culture medium resulting in the maximal degree of activation. No increase in intracellular fluorescence throughout a 40-min period was observed in a cAMP-dependent protein kinase-deficient mutant strain Chinese hamster ovary 10260 cells incubated with TPA. These studies indicate that, while TPA does lead to the marked activation of cAMP-dependent protein kinase in at least two cell types shown to be responsive to TPA, this activation may be only one of several rapid events which mediate the many specific effects of the phorbol esters.
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PMID:Activation of cyclic adenosine 3':5'-monophosphate-dependent protein kinase in H35 hepatoma and Chinese hamster ovary cells by a phorbol ester tumor promoter. 630 80

Using lectin affinity-purified receptor preparations from human hepatoma cells, insulin (10(-7)M) specifically stimulated phosphorylation of the 95,000 dalton (beta) subunit of its own receptor. Phospho-amino acid analysis of the receptor subunit revealed that insulin increased at least 2.5-fold the content of phosphoserine and of phosphotyrosine. In intact cells, the major effect of insulin is to increase the phosphoserine content of its receptor. These findings are the first demonstration of an insulin-stimulated serine kinase in a cell-free system.
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PMID:Insulin stimulates phosphorylation of serine residues in soluble insulin receptors. 631 66

Poly(A) polymerases purified from rat liver nuclei consisted of two distinct species, a predominant enzyme of Mr = 38,000 and a minor one of Mr = 48,000. Prior to extensive purification, the minor enzyme constituted approximately 1% of the total liver poly(A) polymerase. Poly(A) polymerase purified from a rat tumor, Morris hepatoma 3924A, was comprised of a single species of Mr = 48,000 which was identical to the minor liver enzyme with respect to chromatographic and immunological characteristics. Gel filtration on Sephacryl S-200 using 0.3 M NaCl for elution showed that the major liver poly(A) polymerase had a molecular weight of 156,000, which corresponded to a tetramer of the 38-kDa polypeptide, whereas the hepatoma and minor liver 48-kDa species existed as dimers with a molecular weight of 96,000. Fractionation by Sephacryl S-200 resulted in complete loss of both liver poly(A) polymerase activities which could be restored by exogenous N1-type protein kinase. Following CNBr cleavage, the 48-kDa poly(A) polymerase from liver and hepatoma exhibited nearly identical peptide maps which were distinct from that of the major liver enzyme (38 kDa). Antibodies raised against tumor poly(A) polymerase reacted with the 48-kDa polypeptide but not with the 38-kDa liver enzyme. Immune complex formation was observed between seven of the eight CNBr cleavage products derived from the 48-kDa polypeptide of both liver and hepatoma. It is concluded that distinct genes in rat liver code for two structurally and immunologically unique nuclear poly(A) polymerases, one of which is identical to the enzyme from the hepatoma.
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PMID:Structurally and immunologically distinct poly(A) polymerases in rat liver. Occurrence of a tumor-type enzyme in normal liver. 632 12

Insulin stimulates the growth and proliferation of a variety of somatic cells in culture, and evidence suggests that insulin is also an important regulator of growth in vivo. In cell culture, insulin interacts synergistically with other hormones and growth factors such as platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), tumor-promoting phorbol esters, and thrombin, to stimulate progression through the cell cycle of cells that have been arrested in G1 by deprivation for serum. In addition, insulin is required by most cells for optimal long term growth in hormone-supplemented serum-free media. In some cells, such as human skin fibroblasts, the growth-promoting effects of insulin appear to be mediated primarily by its low affinity interaction with receptors for insulin-like growth factor I (IGF-I). In other cells, such as hepatocytes, hepatoma cells, adrenocortical tumor cells, mammary carcinoma cells, and F9 embryonal carcinoma cells, insulin appears to stimulate growth by binding to high affinity insulin receptors. The insulin and IGF-I receptor proteins, like the receptor proteins for other growth-promoting hormones such as EGF and PDGF, are closely associated with tyrosine-specific protein kinase activities. The mechanism by which the binding of insulin to its receptor and activation of the receptor-associated tyrosine protein kinase activity control intracellular protein phosphorylation and dephosphorylation reactions, such as the phosphorylation of ribosomal protein S6, is a subject of considerable current interest. The phosphorylation of ribosomal protein S6 may be related mechanistically to the activation by insulin of protein synthesis, and hence the passage of cells through the G1 phase of the cell cycle. Malignant transformation does not generally result in a total loss of the growth requirement of cells for insulin or insulin-like growth factors, although transformation is accompanied in some cases by a qualitative reduction in the insulin/IGF requirement. Abnormalities in insulin production or sensitivity in vivo are accompanied by abnormalities in growth; thus, insulin appears to be an important regulator of growth in vivo. Some of the growth-promoting effects of insulin in vivo may be attributable to direct action of insulin, while other effects may be caused by the regulatory effect of insulin on somatomedin production, and possibly on somatomedin action.
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PMID:Growth-stimulatory actions of insulin in vitro and in vivo. 637 81

A protein kinase-substrate complex was precipitated by adding Ca2+ to the cytosol fraction of AH-66 ascites hepatoma cells. The amount of the precipitated complex was increased with increasing concentrations of Ca2+ and reached a plateau at about 5 mM Ca2+. In the presence of [gamma-32P]ATP, extensive uptake of radioactive phosphate into this complex occurred. The phosphorylation reaction was little affected by addition of cyclic nucleotides, Ca2+-phospholipid, Ca2+-calmodulin. When the complex after phosphorylation was analyzed by SDS-PAGE, a protein with molecular weight of 33,000 was most heavily phosphorylated. These phenomena were also observed for mouse myeloid leukemia cells (M1 cells). By contrast, the addition of Ca2+ to the cytosol fractions of regenerating rat liver, normal rat liver or brain caused little precipitation of the complex.
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PMID:Characterization of a protein kinase-substrate complex precipitable with Ca2+ from the cytosol fraction of AH-66 hepatoma cells. 650 98

Two major cyclic nucleotide-independent protein kinases, NI and NII, have been identified in Morris hepatoma 3924A and rat liver. When expressed per unit DNA, the activities of protein kinase NI and NII were 1.3 and 12 times greater, respectively, in the hepatoma than in liver. Protein kinase NII, but not NI, was capable of phosphorylating and activating the DNA-dependent RNA polymerases I and II. Phosphorylation of RNA polymerase I was accompanied by an increase in average size of the RNA synthesized in vitro, whereas phosphorylation of RNA polymerase II was concomitant with an elevation in the number of RNA chains initiated. RNA polymerase I polypeptides of Mr 120,000, 65,000 and 25,000 were phosphorylated by protein kinase NII; RNA polymerase II polypeptides of Mr 214,000, 140,000 and 21,000 were modified by this kinase. In contrast to the purified hepatoma enzyme, RNA polymerase I activity in nuclear lysates was not affected by addition of protein kinase NII. In vitro phosphorylation of the tumor lysate followed by immunoprecipitation of RNA polymerase I polypeptides indicated little or no phosphate transfer to the 65,000 Mr polypeptide of the enzyme. These data suggested that the tumor enzyme, particularly the 65,000 Mr polypeptide, was highly phosphorylated in vivo, but becomes dephosphorylated during purification. Unlike the tumor enzyme, RNA polymerase I in the liver lysate responded to protein kinase addition; phosphorylation of the liver polymerase I polypeptides of Mr 120,000, 65,000 and 25,000 was observed. These observations indicate that the liver enzyme is not completely phosphorylated (activated) in vivo and that the relatively rapid rate of ribosomal RNA synthesis in the rapidly growing hepatoma may result, at least in part, from a polymerase I which is maximally phosphorylated.
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PMID:RNA polymerase I in hepatoma 3924A: mechanism of enhanced activity relative to liver. 654 82

Relative to resting liver, Morris hepatomas with different growth rates (3924A, 5123D, 7800, and 7777) all had higher (two to eightfold) levels (activity/gm tissue) of RNA polymerase I. Only the most rapidly growing tumor (hepatoma 3924A) showed a substantial increase (fivefold) in RNA polymerase III activity. RNA polymerase II activity/gm tissue in the hepatomas was similar to that in resting liver. The elevation in the hepatoma RNA polymerase I activity resulted from both an increase in the number of transcriptionally active enzyme molecules and an increase in the specific activity of the enzyme as a result of phosphorylation. Phosphorylation of RNA polymerase I from Morris hepatoma 3924A could be catalyzed either by an endogenous protein kinase or by a highly purified preparation of NII protein kinase from the same tumor. Three out of eight polypeptides (Mr 120,000, 65,000, and 25,000) or RNA polymerase I were phosphorylated. Phosphorylation resulted in enhanced RNA synthesis at the level of chain elongation. Another nuclear protein kinase, NI, had no significant effect on RNA polymerase I. The activity and/or amount of the NII protein kinase was significantly reduced in resting liver, which correlated with decreased specific activity of the liver RNA polymerase I. Anti-RNA polymerase I antibodies were found in the sera of patients with rheumatic autoimmune diseases such as systemic lupus erythematosus (SLE), mixed connective tissue disease (MCTD), and rheumatoid arthritis (RA). Sera from these patients were capable of specifically inhibiting RNA polymerase I activity in vitro. Antibodies were produced predominantly against three of the polypeptides--S3 (Mr 65,000), S4 (Mr 42,000), and S5 (Mr 25,000) of RNA polymerase I. The spectrum and proportion of the antibodies against these three subunits differ with each patient and with the type of the autoimmune disease. These observations indicate that (1) the NII kinase can regulate RNA polymerase I activity, (2) protein kinase NII is associated with the polymerase I enzyme complex, and (3) certain polypeptides of this enzyme complex may be the target antigens in rheumatic autoimmune disease.
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PMID:Regulation of RNA polymerase I by phosphorylation and production of anti-RNA polymerase I antibodies in rheumatic autoimmune diseases. 660 44


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