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:2.7.1.21 (thymidine kinase)
7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Although a number of transfection experiments have suggested potential targets for the action of the E2F1 transcription factor, as is the case for many transcriptional regulatory proteins, the actual targets in their normal chromosomal environment have not been demonstrated. We have made use of a recombinant adenovirus containing the E2F1 cDNA to infect quiescent cells and then measure the activation of endogenous cellular genes as a consequence of E2F1 production. We find that many of the genes encoding S-phase-acting proteins previously suspected to be E2F targets, including DNA polymerase alpha, thymidylate synthase, proliferating cell nuclear antigen, and ribonucleotide reductase, are indeed induced by E2F1. Several other candidates, including the dihydrofolate reductase and thymidine kinase genes, were only minimally induced by E2F1. In addition to the S-phase genes, we also find that several genes believed to play regulatory roles in cell cycle progression, such as the cdc2, cyclin A, and B-myb genes, are also induced by E2F1. Moreover, the cyclin E gene is strongly induced by E2F1, thus defining an autoregulatory circuit since cyclin E-dependent kinase activity can stimulate E2F1 transcription, likely through the phosphorylation and inactivation of Rb and Rb family members. Finally, we also demonstrate that a G1 arrest brought about by gamma irradiation is overcome by the overexpression of E2F1 and that this coincides with the enhanced activation of key target genes, including the cyclin A and cyclin E genes.
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PMID:Cellular targets for activation by the E2F1 transcription factor include DNA synthesis- and G1/S-regulatory genes. 762 16

E2F has been implicated in growth control because of its association with the retinoblastoma protein and the presence of E2F binding sites in the promoters of several growth-regulated genes. Proteins that bind to an E2F site have been cloned from human and mouse cells. However, these two proteins (human E2F1 and mouse DP-1) are quite different in sequence. We have now cloned a mouse cDNA encoding a protein 86% identical to the human E2F1 protein. The mouse E2F1 cDNA encodes a 430-amino-acid protein with a predicted molecular weight of 46,322 and detects mRNAs of 2.7 and 2.2 kb. Using primers complementary to sequences in the mouse E2F1 3' untranslated region, we mapped the mouse E2F1 gene to chromosome 2, near the Agouti and c-src loci. To understand the role of the different E2F family members in the growth of mouse NIH 3T3 cells, we examined the levels of E2F1 and DP-1 mRNAs in different stages of the cell cycle. Since the levels of E2F1 but not DP-1 mRNA correlated with changes in transcription from the dhfr promoter, we examined whether E2F1 could activate various growth-regulated promoters. We found that E2F1 could activate some (dhfr, thymidine kinase, and DNA polymerase alpha) but not all (thymidylate synthase, cad, and c-myc) of these promoters. On the basis of changes in levels of E2F1 and its ability to transactivate growth-regulated promoters, we propose that E2F1 may mediate growth factor-initiated signal transduction.
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PMID:Cloning, chromosomal location, and characterization of mouse E2F1. 811 19

Within the region around 150 bp upstream of the initiation codon, which was previously shown to suffice for growth-regulated expression, the murine thymidine kinase gene carries a single binding site for transcription factor Sp1; about 10 bp downstream of this site, there is a binding motif for transcription factor E2F. The latter protein appears to be responsible for growth regulation of the promoter. Mutational inactivation of either the Sp1 or the E2F site almost completely abolishes promoter activity, suggesting that the two transcription factors interact directly in delivering an activation signal to the basic transcription machinery. This was verified by demonstrating with the use of glutathione S-transferase fusion proteins that E2F and Sp1 bind to each other in vitro. For this interaction, the C-terminal part of Sp1 and the N terminus of E2F1, a domain also present in E2F2 and E2F3 but absent in E2F4 and E2F5, were essential. Accordingly, E2F1 to E2F3 but not E2F4 and E2F5 were found to bind sp1 in vitro. Coimmunoprecipitation experiments showed that complexes exist in vivo, and it was estabilished that the distance between the binding sites for the two transcription factors was critical for optimal promoter activity. Finally, in vivo footprinting experiments indicated that both the sp1 and E2F binding sites are occupied throughout the cell cycle. Mutation of either binding motif abolished binding of both transcription factors in vivo, which may indicate cooperative binding of the two proteins to chromatin-organized DNA. Our data are in line with the hypothesis that E2F functions as a growth- and cell cycle regulated tethering factor between Sp1 and the basic transcription machinery.
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PMID:Interaction of Sp1 with the growth- and cell cycle-regulated transcription factor E2F. 865 41

The hallmark of cellular aging is the failure of senescent cells to initiate the DNA synthesis during the progression of cell cycle. Since most, if not all, of the G1/S genes exhibit a significant down-regulation during aging, an alteration of gene regulation at late G1/S boundary could be a major contributing factor for the loss of dividing potential during cell senescence. The underlying cause for the apparent global attenuation of gene expression at late G1/S boundary is not clear. Since we have shown that thymidine kinase (TK) and dihydrofolate reductase (DHFR) are transcriptionally regulated during aging, we suspect that a similar mechanism may be operative in the age-dependent down-regulation of other G1/S genes. DNA binding activities using Y-box containing sequence in TK promoter or E2F containing sequence in DHFR promoter show prominent serum-responsiveness in low passage cells and dramatic attenuation in senescent cells. Promoter analysis using GCG program reveals striking similarities in promoter organization of twelve age-dependent G1/S genes. Specifically, these genes can be divided into two groups, one group contains tandem multiple CCAAT element, similar to that in TK promoter and the other contains E2F site, similar to that in DHFR promoter. Further analysis shows that the promoter of transcription factor, NF-Y, which recognizes CBP/tk site contains a tandem, two Y-box motif, similar to that in TK promoter and that the promoter of E2F1 contains four E2F motifs and two tandem CCAAT elements. Thus, these two important transcription factors could undergo autoregulatory control themselves. It is possible that regulation of only a few of transcription factors such as CBP/tk (NF-Y) and E2F1 may be sufficient to cause a global attenuation of most of G1/S genes in human diploid fibroblasts during senescence.
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PMID:Transcription factors and the down-regulation of G1/S boundary genes in human diploid fibroblasts during senescence. 928 3

Cell cycle progression is subject to several regulatory controls, of which the p53 protein plays a major role in growth arrest, subsequent to the detection of cellular aberrations. It is well documented that p53 has the ability to inhibit transcription driven by several promoters, possibly via distinct mechanisms. In this report, we show that expression of the cell cycle regulatory transcription factor DP1 is strongly inhibited by p53, at the level of transcription and probably through the basal TATA-less promoter. This inhibitory activity has a relative specificity for the DP1 promoter compared with the functionally related E2F1 promoter or unrelated promoters such as those of the transcription factor ATFa or the thymidine kinase gene. Inhibition of DP1 transcription has implications in one of the several possible mechanisms through which p53 induces cell cycle arrest.
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PMID:The p53 tumor suppressor inhibits transcription of the TATA-less mouse DP1 promoter. 955 76

The members of the Sp1 transcription factor family can act as both negative and positive regulators of gene expression. Here we show that Sp1 can be a target for histone deacetylase 1 (HDAC1)-mediated transcriptional repression. The histone deacetylase inhibitor trichostatin A activates the chromosomally integrated murine thymidine kinase promoter in an Sp1-dependent manner. Coimmunoprecipitation experiments with Swiss 3T3 fibroblasts and 293 cells demonstrate that Sp1 and HDAC1 can be part of the same complex. The interaction between Sp1 and HDAC1 is direct and requires the carboxy-terminal domain of Sp1. Previously we have shown that the C terminus of Sp1 is necessary for the interaction with the transcription factor E2F1 (J. Karlseder, H. Rotheneder, and E. Wintersberger, Mol. Cell. Biol. 16:1659-1667, 1996). Coexpression of E2F1 interferes with HDAC1 binding to Sp1 and abolishes Sp1-mediated transcriptional repression. Our results indicate that one component of Sp1-dependent gene regulation involves competition between the transcriptional repressor HDAC1 and the transactivating factor E2F1.
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PMID:Histone deacetylase 1 can repress transcription by binding to Sp1. 1040 40

The E2F1 transcription factor plays a pivotal role in driving cells out of a quiescent state and into the S phase of the cell cycle, in part by transactivating genes needed for DNA replication including DHFR, thymidine kinase, and DNA Polymerase alpha. E2F1 has also been implicated in regulating an S phase checkpoint, however its role in this checkpoint is not well defined. To determine how E2F1 affects such a checkpoint, we utilized an in vivo replication assay employing a plasmid based SV40 origin of replication, transfected into cells expressing SV40 large T antigen. Here we show that expression of full length E2F1, or only its N terminus, represses replication from plasmids containing the SV40 origin, while N terminal deletions of E2F1 do not. E2F1 appears to inhibit the elongation phase of replication and not the initiation phase since it does not affect the replication of other cotransfected plasmids containing only the SV40 origin. Further, inhibition of replication is dependent on both the amino-terminus of the E2F1 protein and on a DNA sequence that is contained within the 3' end of the E2F1 cDNA. Additionally, both full-length E2F1, or just its N-terminus, form protein complexes with two portions of the 3' end of the E2F1 cDNA. These data provide a clue to the mechanism by which E2F1 regulates transit through the S phase checkpoint, by acting on a specific DNA sequence via its amino-terminal region, to inhibit elongation of DNA replication.
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PMID:The amino-terminus of the E2F-1 transcription factor inhibits DNA replication of autonomously replicating plasmids in mammalian cells. 1203 40

The functional effect of the interaction of E2F1 and hepatitis B virus X protein (HBx) on the promoter of human p53 gene was studied using chloramphenicol acetyl transferase (CAT) assay. E2F1 activated the p53 promoter through E2F1 binding site. As previously reported, HBx repressed the p53 promoter through E-box. When E2F1 was cotransfected with HBx, E2F1 overcame the repressive effect of HBx on the p53 promoter through the E2F1 site. However, in the thymidine kinase (tk) heterologous promoter system with the E2F1 binding sites, cotransfection of E2F1 and HBx showed a strong synergistic activation. An in vitro interaction assay showed that E2F1 and HBx physically bind with each other. Analyses of the interaction domain with the GAL4 fusion protein showed that the pRb-binding domain of E2F1 was necessary for the functional interaction of these two proteins. Taken together, these results imply the functional inhibitory action of E2F1 on the HBV life cycle and HBV-mediated hepatocellular carcinogenesis (HCC). Therefore, the normal or enhanced function of E2F1 gene would be important in controlling the HBx function in HCC.
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PMID:E2F1 activates the human p53 promoter and overcomes the repressive effect of hepatitis B viral X protein (Hbx) on the p53 promoter. 1262 70

Various studies point to the potential role of combinatorial action of transcription factors as a mechanism to achieve the complexity of eukaryotic gene control with a finite number of regulatory proteins. Our previous work has focused on interactions involving the E2F family of transcription factors as an example of combinatorial gene control, leading to the identification of TFE3 and YY1 as transcription partners for several E2F proteins. We now show that additional E2F target genes share a common promoter architecture and are also regulated by the combined action of TFE3 and E2F3. In contrast, the thymidine kinase (TK-1) promoter is also regulated by E2F3 but independent of TFE3. Other promoters exhibit distinct specificity in the interaction with E2F proteins that includes a role for E2F1 but not E2F3, examples where both E2F1 and E2F3 are seen to interact, and promoters that are regulated by TFE3 but independent of an E2F. We propose that these examples of combinatorial interactions involving E2F proteins provide a basis for the specificity of transcription control in the Rb/E2F pathway.
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PMID:Combinatorial gene control involving E2F and E Box family members. 1501 47

The IkappaB kinase (IKK) complex consists of the catalytic subunits IKKalpha and IKKbeta and a regulatory subunit, IKKgamma/NEMO. Even though IKKalpha and IKKbeta share significant sequence similarity, they have distinct biological roles. It has been demonstrated that IKKs are involved in regulating the proliferation of both normal and tumor cells, although the mechanisms by which they function in this process remain to be better defined. In this study, we demonstrate that IKKalpha, but not IKKbeta, is important for estrogen-induced cell cycle progression by regulating the transcription of the E2F1 gene as well as other E2F1-responsive genes, including thymidine kinase 1, proliferating cell nuclear antigen, cyclin E, and cdc25A. The role of IKKalpha in regulating E2F1 was not the result of reduced levels of cyclin D1, as overexpression of this gene could not overcome the effects of IKKalpha knock-down. Furthermore, estrogen treatment increased the association of endogenous IKKalpha and E2F1, and this interaction occurred on promoters bound by E2F1. IKKalpha also potentiated the ability of p300/CBP-associated factor to acetylate E2F1. Taken together, these data suggest a novel mechanism by which IKKalpha can influence estrogen-mediated cell cycle progression through its regulation of E2F1.
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PMID:IKK alpha regulates estrogen-induced cell cycle progression by modulating E2F1 expression. 1640 16


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