Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In this study we analyzed the covalent binding to proteins of 17 beta-estradiol (E2), retinoic acid (RA), and progesterone in MCF-7 and MCF-7/AdrR cells. MCF-7 cells have receptors for E2 and progesterone. MCF-7/AdrR cells do not have these receptors. After a 1-day incubation period with either [3H]E2, [3H]progesterone, or [3H]RA the levels of covalently bound radioactivity was between 1.4- to 2-fold greater in MCF-7 cells than in MCF-7/AdrR cells. We analyzed the labeled proteins with two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and fluorography. About 40 proteins were labeled by E2 in MCF-7 cells and about 10 of these proteins were the only proteins labeled by E2 in MCF-7/AdrR cells. We saw that the same 8 proteins were labeled by RA in both cell lines. Progesterone labeled 2 proteins with M(r) values of 37,000 and 20,000 in MCF-7 cells. These 2 proteins had mobilities that were the same as proteins that were labeled by either E2 or RA in both MCF-7 and MCF-7/AdrR cells. Besides these 2 proteins, we saw proteins of M(r) 51,000 (p51) and 55,000 that were covalently labeled by E2 in MCF-7 cells and by RA in both MCF-7 and MCF-7/AdrR cells. The p51 had the same mobility on 2D-PAGE as an 8-azido-[32P]cAMP-labeled protein. This protein is probably RII alpha, the type II cAMP-binding regulatory subunit of type II cAMP-dependent protein kinase. These results suggest that the estrogen receptor, while not obligatory, might still modulate the covalent linkage of E2 to protein. In addition, our results raise the possibility that some effects of some ligands of the thyroid/steroid hormone receptor family may involve the covalent linking of these hormones to proteins, including RII alpha.
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PMID:The covalent labeling of proteins by 17 beta-estradiol, retinoic acid, and progesterone in the human breast cancer cell lines MCF-7 and MCF-7/AdrR. 132 24

The biological activity of interferons (IFNs) is presumed to be mediated through the induction of a number of IFN-inducible genes. IFN-mediated gene induction was examined in two human breast cancer cell lines, MCF-7 and BT-20. Both these cell lines were remarkably responsive to IFNs as a number of IFN inducible genes were rapidly induced. We examined the sensitivity of these genes towards 2-aminopurine (2-AP), a known inhibitor of double-stranded (ds) RNA dependent protein kinase. 2-AP has also been reported to inhibit the induction of IFN-beta 1 in response to dsRNA and the genes c-myc and c-fos in fibroblasts. In both MCF-7 and BT-20 cell lines, 2-AP selectively inhibited the IFN-induced gene responses. 2-AP did not affect levels of the oncogene, HER-2/neu. Tamoxifen (TAM), an antiestrogenic drug, which is known to inhibit the activity of protein kinase C at high concentrations, did not affect IFN-mediated gene induction. Our data is consistent with the concept that the 2-AP sensitive kinase is primarily associated with the IFN-induced gene systems and that positive and negative growth regulating stimuli in breast cancer may require the participation of distinct kinases.
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PMID:A distinct kinase modulates the expression of IFN-inducible genes in human breast cancer cells. 171 33

We have examined the ability of estradiol (E2) to regulate the expression of three mRNAs [for pS2, progesterone receptor (PR), and estrogen receptor (ER)], known to be under E2 regulation in the parental E2 growth-responsive MCF-7 cells, in an E2 growth-independent MCF-7 K3), previously isolated from the parental estrogen-dependent MCF-7 K1 human breast cancer cells after long term growth in vitro in the absence of estrogen, acquired estrogen-independent growth in vitro as well as the ability to form tumors in nude mice in vivo without estrogen. We find that the content of pS2 mRNA and the transcription rate of the pS2 gene, while being markedly increased by E2 in MCF-7 K1 cells, are no longer stimulated by E2 in this subline, although protein kinase activators tremendously increase (greater than 10-fold) pS2 mRNA in both K1 and K3 cells. In fact, basal pS2 mRNA levels are elevated 2.8 +/- 0.4-fold in MCF-7 K3 cells, and E2 evokes a concentration-dependent suppression of the pS2 mRNA level. In contrast, PR mRNA in the K3 subline, as in the parental K1 cells, is still up-regulated by E2, and ER mRAN content and the ER mRNA transcription rate are still down-regulated by E2 and show normal E2 dose-response relationships, implying that the ER in this subline is functional. These results demonstrate that the progression to estrogen-independent growth in K3 cells is accompanied by a change in the regulation of some estrogen-induced genes by estrogen. While PR and ER retain normal patterns of regulation by E2, the pS2 gene in the estrogen growth-independent K3 subline is differentially affected and is no longer stimulated by E2. Our data suggest that this altered regulation of the pS2 gene is probably not caused by a defect of the ER or ER regulation in this subline.
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PMID:Differential regulation of gene expression by estrogen in estrogen growth-independent and -dependent MCF-7 human breast cancer cell sublines. 172 71

A novel serine/threonine protein kinase (termed rac-PK) has recently been identified and cloned from cDNA libraries derived from the human cell lines MCF-7 and WI38. A second form of this protein kinase, termed rac protein kinase beta, has been identified from cDNAs derived from the same cell lines. These two closely related forms show 90% homology, although the beta form with a predicted Mr 60,200 has a carboxyl terminal extension of 40 amino acids in comparison to the alpha form. This extension has a high serine content with 11 serine residues in the last 30 amino acids. The beta form of the protein has been shown by both in vitro translation and bacterial expression to be approximately 5000 Da larger than the alpha form. rac protein kinase beta is encoded by a 3.4-kb transcript and the alpha form is encoded by a 3.2-kb mRNA. Using gene-specific probes both transcripts were detected in all cell types analyzed, although levels of expression were different for the two forms. The catalytic domain of rac protein kinase beta shows a high degree of homology to both the protein kinase C and cyclic AMP-dependent protein kinase families, and hence rac protein kinases appear to represent a new subfamily of the second messenger serine/threonine protein kinases.
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PMID:Molecular cloning of a second form of rac protein kinase. 180 21

A partial cDNA was isolated that encoded a protein kinase, termed rac (related to the A and C kinases). This cDNA was subsequently used to screen libraries derived from the human cell lines MCF-7 and WI38 and led to the isolation of full-length cDNA clones. DNA sequence analysis identified an open reading frame of 1440 base pairs encoding a protein of 480 amino acids (Mr, 55,716). This result was supported by the synthesis of a Mr 58,000 protein in an in vitro translation system that used RNA transcribed from cloned cDNAs with SP6 RNA polymerase. The predicted protein contains consensus sequences characteristic of a protein kinase catalytic domain and shows 73% and 68% similarity to protein kinase C and the cAMP-dependent protein kinase, respectively. Northern (RNA) analysis revealed a single mRNA transcript of 3.2 kilobases that varied up to 300-fold between different cell lines. Specific antisera directed towards the carboxyl terminal of the rac protein kinase were prepared and used to identify that phosphorylated several substrates in immunoprecipitates prepared with the rac-specific antisera.
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PMID:Molecular cloning and identification of a serine/threonine protein kinase of the second-messenger subfamily. 185 97

Purified L-asparaginase of Tetrahymena pyriformis is a multi-subunit enzyme exhibiting protein kinase activity as well. The enzyme's L-asparaginase activity is affected by its phosphorylation state. Both native and dephosphorylated L-asparaginase show antiproliferative activity on three breast cancer cell lines (T47D, BT20 and MCF-7) and on Walker 256 cells. These cells do not possess measurable L-asparaginase or L-asparagine synthetase activity. When T47D cells are treated for different times with L-asparaginase and then placed in fresh medium, the growth of cells treated for 1, 3, or 6 hours is initiated and parallels control curve, while the growth of cells treated for 24 or 48 hours with L-asparaginase stays at the same inhibitory level (24 h treatment) or continues to drop (48 h treatment). Addition of D-asparagine, a competitive inhibitor of T. pyriformis L-asparaginase, counteracts the antiproliferative activity of L-asparaginase, indicating that L-asparaginase and not the kinase activity is responsible for that effect.
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PMID:Antiproliferative activity of L-asparaginase of Tetrahymena pyriformis on human breast cancer cell lines. 212 95

We have examined the synthesis of the protease inhibitors alpha 1-antitrypsin (alpha 1-AT) and alpha 1-antichymotrypsin (alpha 1-ACHY) by variants of the MCF-7 human breast cancer cell line. Spent medium from MCF-7 203P cells, grown in the absence of serum, was found to contain immunoreactive alpha 1-AT and alpha 1-ACHY by Western blotting. In the presence of 10(-8) M estradiol, levels of both inhibitors were increased 3- to 6-fold. Incubation of spent medium with [125I]trypsin or [125I]chymotrypsin resulted in the formation of stable 75- and 90-kDa complexes identical to the complexes formed between these proteases and the protease inhibitors in plasma, showing the release of active protease inhibitors by MCF-7 cells in culture. Immunoprecipitation of 35S-labeled proteins from the medium of cells grown in the presence of [35S]methionine yielded comparable results, confirming hormonally sensitive synthesis of both protease inhibitors. Northern blot analysis suggests that stimulation of estradiol occurs at the level of transcription. Tetradecanoyl phorbol acetate (50 ng/ml) also stimulated alpha 1-AT and alpha 1-ACHY synthesis 2- to 4-fold, suggesting the involvement of protein kinase-C. Comparison studies with MCF-7 cell sublines ML, BK, 203P, and 300P (a variant spontaneously appearing after 100 passages of 203P) show a wide variation in synthesis of alpha 1-AT and alpha 1-ACHY proteins; sublines 203P and 300P synthesize both inhibitors, the ML subline synthesizes detectable amounts only of alpha 1-ACHY, while no detectable synthesis of either inhibitor was seen in the BK subline. Similar results were obtained for protease inhibitor mRNA transcription by Northern blotting, although low levels of alpha 1-AT mRNA transcription by the ML subline and of alpha 1-AT and alpha 1-ACHY mRNA transcription by the BK subline could be detected.
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PMID:Regulation of antitrypsin and antichymotrypsin synthesis by MCF-7 breast cancer cell sublines. 220 35

We investigated the effects of the antiestrogen tamoxifen on MCF-7 cell protein kinase C either by using the in vitro histone kinase assay or by studying the phosphorylation of its endogenous Mr 28,000 protein substrate in intact cells. In the in vitro assay, tamoxifen inhibited the enzyme competitively with respect to phospholipid, whereas estradiol and morpholinobenzyl phenoxy ethanamine, a specific ligand for antiestrogen binding sites, were considerably less efficient. In contrast, tamoxifen did not affect phosphorylation of the Mr 28,000 protein induced by the phorbol ester 12-O-tetradecanoylphorbol 13-acetate in intact MCF-7 cells. Estradiol and morpholinobenzyl phenoxy ethanamine also had no effect. At high concentration (100 microM), tamoxifen itself stimulated specific phosphorylation of this Mr 28,000 protein. Estradiol and morpholinobenzyl phenoxy ethanamine neither mimicked nor interfered with this effect. Our data suggest that the effect of tamoxifen on protein kinase C activity depends on the phospholipid environment of the enzyme, and opposite effects may be observed in intact cells to those seen in disrupted cells. The action of tamoxifen on endogenous protein phosphorylation was thought to be due to direct interaction with the phospholipid binding domain of the enzyme rather than by interaction with the estrogen receptor or the antiestrogen binding site. Nevertheless, our results do not rule out a possible activation by tamoxifen of specific protein kinase(s) and phosphatase(s). In any case, the antiproliferative activity of tamoxifen on MCF-7 cells cannot be attributed to its effects on protein kinase C.
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PMID:Opposite effects of tamoxifen on in vitro protein kinase C activity and endogenous protein phosphorylation in intact MCF-7 cells. 239 53

Cyclic AMP (cAMP) analogues that selectively bind to either one of the two binding sites of cAMP-dependent protein kinase demonstrate a potent inhibition of the growth stimulated by estrogen in MCF-7 human breast-cancer cells in culture. The site-selective analogues, which are more potent activators of protein kinase than the analogues studied earlier, exhibit growth inhibition at micromolar concentrations. Among the analogues tested, 8-Cl-cAMP (Site I-selective) and N6-benzyl-cAMP (Site 2-selective) are the 2 most potent inhibitors, causing 40-70% inhibition of the estrogen-stimulated growth at 10-20 microM concentrations with no sign of toxicity. 8-Cl-cAMP (1 microM) in combination with N6-benzyl-cAMP (0.5 microM) almost completely blocks estrogen-stimulated growth, demonstrating synergism between the Site 1- and Site 2-selective analogues. The growth inhibition parallels an increase in the R11 cAMP receptor protein with a decrease in the R1 receptor as well as reduction of c-myc and c-ras oncoproteins, whereas growth inhibition by tamoxifen does not affect the levels of the cAMP receptor proteins or the c-myc and c-ras protein levels. Site-selective cAMP analogues are antagonistic to estrogen stimulation of breast-cancer cell growth through a mechanism different from that of tamoxifen.
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PMID:Site-selective cyclic AMP analogues are antagonistic to estrogen stimulation of growth and proto-oncogene expression in human breast-cancer cells. 283 20

Quantitative polyacrylamide gel electrophoresis analysis of Ca2+, phospholipid-dependent protein kinase (PKC) of human mammary tumor cell lines (MCF-7, ZR-75, T-47-D, MDA-MB-231, BT-20, and HBL-100) revealed that 80% of the total cellular PKC resided in the cytosol. The tumor cells with no detectable levels of estrogen receptors (MDA-MB-231, HBL-100, and BT-20 cells) exhibited significantly larger (P less than 0.001) cytosolic PKC activities than those cells that contained estrogen receptors (MCF-7, T-47-D, and ZR-75 cells). In addition, in estrogen receptor-negative cell lines, relatively high levels of specific low-affinity (apparent Kd = 700 pM) epidermal growth factor (EGF) binding activities were found as compared with estrogen receptor-positive cells with significantly (P less than 0.001) lower levels of specific high-affinity (apparent Kd = 90 pM) EGT binding. A significant positive correlation (P less than 0.01) was observed between the number of EGF receptor (Rs = 0.50) and/or the EGF receptor dissociation constants (Rs = 0.78) with the cytosolic PKC activity levels. These data indicate that, in human breast cancer cells, a positive relationship may exist between PKC activity, estrogen, and EGF receptors.
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PMID:Epidermal growth factor binding and protein kinase C activities in human breast cancer cell lines: possible quantitative relationship. 300 98


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