Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P04637 (p53)
 77,613 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Twenty-five mouse lung tumors induced by a single urethan treatment in female A/J, BALB/c, and (A/J x C3H/He)F1 (AC3) mice were analyzed for the presence of mutations at codon 61 of the Ki-ras gene and for the expression of the surfactant protein A (SP-A), retinoblastoma (Rb), growth arrest-specific-3 (gas-3), p53, c-myc, and thymidylate synthase (TS) genes. Ki-ras codon 61 mutations were detected in 22 of 25 tumor samples without differences among strains. In comparison with normal lungs, all the tumors showed increased SP-A mRNA levels, indicating their derivation from alveolar type II pneumocytes or Clara cells. Rb and gas-3 transcripts were instead found in all tumors at about tenfold and about 20-fold reduced levels, respectively. No apparent structural alterations or loss of heterozygosity at the Rb locus was detected in any tumors. The p53 mRNA was observed without variation in quantity or size in lung tumors and normal tissue. A threefold to fivefold c-myc overexpression was observed, without amplification of the gene. TS expression was only slightly increased, indicating no great differences in cell proliferation between lung tumors and normal tissue. Our data suggest that the pathogenesis of urethan-induced lung tumors in mice involves specific and recurrent molecular alterations (Ki-ras mutations, decrease of Rb and gas-3 expression, and increase of c-myc expression) that could represent different steps in lung carcinogenesis.
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PMID:Multiple molecular alterations in mouse lung tumors. 155 14

Asynchronous populations of Ehrlich ascites tumor cells grown in vivo were separated by centrifugal elutriation into fractions of G1-, S-, and G2/M-phase cells with less than 10% cross-contamination. Cytoplasmic mRNA from phase-synchronous cells was used to prepare cDNA which was ligated with bacteriophage lambda gt10 arms and amplified in Escherichia coli C600 hfl-. EcoRI digests of DNA isolated from the sublibraries (G1, S, G2/M) were submitted to Southern hybridizations with radiolabeled probes either (a) for genes whose phase-specific expression is clearly documented, thymidine kinase, dihydrofolate reductase, and thymidylate synthase, or (b) for genes whose change of expression during the cell cycle is likely, lamin C, beta-actin, alpha- and beta-tubulin, c-myc, c-fos, p53. The cDNA sequences for genes of group (a) were found to be significantly enriched in DNA of the S-phase library indicating that the cell cycle phase-specific patterns of the respective mRNA levels are conserved in the sublibraries. Sequences belonging to group (b) were also found to be enriched in DNA isolated from the sublibraries: c-fos in G1 phase, lamin C, beta-actin, tubulins, c-myc in S phase, and p53 in G1/S phase. The unexpected prevalence of c-myc and alpha-tubulin in the S-phase library is supported by Northern analysis of RNA from phase-synchronous cells. Non-phase-specific, randomly chosen sequences hybridized equally strong with DNA isolated from the different sublibraries. No significant changes of the patterns of hybridization signals were observed with DNA from different amplifications of the sublibraries when analyzed with the same DNA probe indicating that the cDNA complexities are well conserved during amplifications. Consequently, the sublibraries are useful to obtain information about the cell cycle phase-specific expression of mRNAs for other genes of interest. Since the sublibraries reflect mRNA levels of the cells growing in vivo they supply data on the physiological in vivo pattern of gene expression undisturbed by potentially unphysiological in vitro conditions.
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PMID:Cell cycle phase-specific cDNA libraries reflecting phase-specific gene expression of Ehrlich ascites cells growing in vivo. 333 23

The retinoblastoma susceptibility gene (Rb) participates in controlling the G1/S-phase transition, presumably by binding and inactivating E2F transcription activator family members. Mouse embryonic fibroblasts (MEFs) with no, one, or two inactivated Rb genes were used to determine the specific contributions of Rb protein to cell cycle progression and gene expression. MEFs lacking both Rb alleles (Rb-/-) entered S phase in the presence of the dihydrofolate reductase inhibitor methotrexate. Two E2F target genes, dihydrofolate reductase and thymidylate synthase, displayed elevated mRNA and protein levels in Rb- MEFs. Since absence of functional Rb protein in MEFs is sufficient for S-phase entry under growth-limiting conditions, these data indicate that the E2F complexes containing Rb protein, and not the Rb-related proteins p107 and p130, may be rate limiting for the G1/S transition. Antineoplastic drugs caused accumulation of p53 in the nuclei of both Rb+/+ and Rb-/- MEFs. While p53 induction led to apoptosis in Rb-/- MEFs, Rb+/- and Rb+/+ MEFs underwent cell cycle arrest without apoptosis. These results reveal that diverse growth signals work through Rb to regulate entry into S phase, and they indicate that absence of Rb protein produces a constitutive DNA replication signal capable of activating a p53-associated apoptotic response.
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PMID:Deficiency of retinoblastoma protein leads to inappropriate S-phase entry, activation of E2F-responsive genes, and apoptosis. 777 26

The endogenous expression of p53 and p53-regulated genes has been examined in a thymidylate synthase-deficient colon carcinoma cell line (TS-) and a derived mutant clone (Thy4) that exhibit acute or delayed apoptotic responses, respectively, when released from G0 synchrony under conditions of dThd starvation. These cell clones demonstrate heterozygosity in p53, thereby expressing one wt allele and one with an A-->C point mutation at codon 240. Following release from G0, upregulated expression of both alleles occurred. During apoptosis in TS-, a wtp53 phenotype was expressed and in Thy4 during cytostasis, a mp53 phenotype was manifested, as determined from the ratios of wtp53/mp53 proteins, transactivation of p50-2 (a wtp53-responsive CAT reporter construct) and the endogenous expression of MDM2. Neither cytotoxicity nor cytostasis correlated with expression of p21Waf1/Cip1 Thy4 cells sustained accumulation of high levels of Bax in a wtp53-independent and dThd-independent manner and survival was associated with upregulated expression of Bcl-2. In contrast, Bax expression decreased in TS- during apoptosis, except in a highly resistant subpopulation that retained high levels of Bax. Data suggest that resistant cells (Thy4) can sustain high Bax expression and that Bcl-2 is upregulated in response to an apoptotic stimulus due to the absence of negative regulation by wtp53.
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PMID:Acute and delayed apoptosis induced by thymidine deprivation correlates with expression of p53 and p53-regulated genes in colon carcinoma cells. 866 31

Fluoropyrimidines radiosensitize human colon cancer cells that progress into S phase in the presence of drug (M.A. Davis, H-Y. Tang, J. Maybaum, and T.S. Lawrence. Int. J. Radiat. Biol. 67. 509-512, 1995). We hypothesized that progression occurs in cells that generate elevated levels of cyclin E-dependent kinase activity despite the presence of the fluoropyrimidine. To test this hypothesis, we treated HT29 and SW620 human colon cancer cells with fluorodeoxyuridine under conditions that produced nearly complete inhibition of thymidylate synthase but which sensitized only the HT29 cells. We found that, whereas HT29 cells progressed into S phase and demonstrated increased cyclin E-dependent kinase activity, SW620 cells arrested just past the G1-S boundary and showed no change in kinase activity. Because these cell lines have the same p53 mutation, these findings suggest that there is a p53-independent G1-S checkpoint that mediates radiosensitization produced by fluorodeoxyuridine.
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PMID:Fluoropyrimidine-mediated radiosensitization depends on cyclin E-dependent kinase activation. 876 7

Using four complementary approaches, ie., cell synchronization, bromodeoxyuridine labeling, and DNA and Western blot analyses, we investigated the underlying mechanism of cell cycle perturbation in response to ZD1694, a quinazoline-based antifolate thymidylate synthase inhibitor. With a single exposure at a concentration of 1 microM for 2 h, ZD1694 completely inhibits thymidylate synthase over 72 h and causes a sustained growth for at least 120 h, DNA damage, and p53 induction in human carcinoma cells. Although these cells displayed an S-phase block with the precise terminal arrest point depending on the timing of drug treatment in the cell cycle, their DNA-replicating machinery associated with polymerase alpha was preserved intact. When supplemented with exogenous dThd, these cells resumed an apparently normal S-phase progression for at least 4 h. Kinetic analyses based on synchronized cells indicate that S-phase arrest occurs first, preceding the induction of DNA double strand breaks and p53/p21. SW480 cells, in which p53mu failed to transduce p21, also exhibited the mode of S-phase arrest, essentially indistinguishable from that displayed by HCT-8 cells expressing the functional p53 (p53wt). That the DNA replication process is prerequisite for DNA double strand breaks was indicated by the following: (a) DNA damage occurred only when cells treated with ZD1694 progressed through S phase; and (b) the inhibition of DNA polymerase alpha by aphidicolin-blocked DNA damage. Based on the above, we conclude that S-phase arrest by ZD1694, with a subsequent damage of DNA double strands, is caused by the block of DNA synthesis in the middle of replication due to dTTP depletion and not by p53-mediated G1-G2 checkpoint mechanisms or p21-induced inactivation of the DNA-replicating machinery.
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PMID:DNA damage and p53 induction do not cause ZD1694-induced cell cycle arrest in human colon carcinoma cells. 884 Sep 89

Thymidylate synthase plays a central role in the biosynthesis of thymidylate, an essential precursor for DNA biosynthesis. In addition to its role in catalysis and cellular metabolism, studies from our laboratory have shown that thymidylate synthase functions as an RNA binding protein. Specifically, thymidylate synthase binds with high affinity to its own mRNA resulting in translational repression. An extensive series of experiments have now been performed to elucidate the molecular elements underlying the interaction between thymidylate synthase and its own mRNA. These studies have shed new light into the critical nucleotide sequences and/or secondary structure that are important for protein recognition. As well, studies to define the domains on the protein essential for RNA binding are currently underway. In addition to the characterization of the cis- and trans-acting elements underlying the interaction between thymidylate synthase and its own mRNA, we have recently shown that thymidylate synthase has the capacity to specifically bind in vitro and in vivo to other cellular RNA species. In this regard, thymidylate synthase interacts with the mRNAs of the c-myc onocogene and the p53 tumor suppressor gene. These two genes have been shown to play critical roles in cell cycle control, DNA biosynthesis, and apoptosis. In vitro studies reveal that the interaction of TS with these cell-cycle related mRNAs results in their translational repression. While the biological significance of these cellular RNA/TS protein interactions remains to be defined, these studies suggest a potential role for TS as a mediator in the coordinate regulation of several critical aspects of cellular metabolism.
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PMID:The role of thymidylate synthase in cellular regulation. 886 45

The epithelia from the crypts of the intestine are exquisitely sensitive to metabolic perturbation and undergo cell death with the classical morphology of apoptosis. Administration of 40 mg/kg 5-fluorouracil (5-FU) to BDF-1 p53+/+ mice resulted in an increase in p53 protein at cell positions in the crypts that were also those subjected to an apoptotic cell death. In p53-/- mice apoptosis was almost completely absent, even after 24 hr. 5-FU is a pyrimidine antimetabolite cytotoxin with multiple mechanisms of action, including inhibition of thymidylate synthase (TS), which gives rise to DNA damage, and incorporation into RNA. The inhibition of TS can be increased by coadministration of folinic acid and can be abrogated by administration of thymidine. The incorporation of 5-FU into RNA is inhibited by administration of uridine. p53-Dependent cell death induced by 5-FU was only inhibited by administration of uridine. Uridine had no effect on the apoptosis initiated by 1 Gy of gamma-radiation. Although thymidine abrogated apoptosis induced by the pure TS inhibitor Tomudex, it had no effect on 5-FU-induced apoptosis, and coadministration of folinic acid did not increase apoptosis. The data show that 5-FU-induced cell death of intestinal epithelial cells is p53-dependent and suggests that changes in RNA metabolism initiate events culminating in the expression of p53.
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PMID:Inhibition by uridine but not thymidine of p53-dependent intestinal apoptosis initiated by 5-fluorouracil: evidence for the involvement of RNA perturbation. 905 Aug 58

In a previous study, we found that treatment of HCT-8 cells with ZD1694, a specific antifolate-based thymidylate synthase inhibitor, resulted in DNA fragmentation. In this study, we have demonstrated the dose- and time-dependent induction of DNA fragmentation accompanied by elevation of p53 and WAF1 protein expression by ZD1694. WAF1 mRNA showed a time-dependent increase, whereas p53 mRNA was not found to be significantly overexpressed. The initial increase in WAF1 mRNA was detected at 4 hr, but increased WAF1 protein expression was detected 8-24 hr after a 2-hr exposure. The amount of total and hypophosphorylated pRb seems to be rising greatly after ZD1694 exposure. The effects of ZD1694 on the expression of E2F1 and formation of the E2F1-Rb complex were investigated after a 2-hr drug exposure (IC90). The results showed a time-dependent decrease in E2F1 mRNA and protein expression; an increase in the abundance of the E2F-Rb complex could be demonstrated beginning 4 hr after drug exposure by a gel shift assay. Kinetic analysis showed increased availability of hypophosphorylated pRb for inhibition of E2F, which could indirectly result from WAF1-induced inhibition cyclin-dependent kinase activity. Whereas thymidylate synthase inhibition by ZD1694 was rapid in onset and maintained for at least 24 hr after drug treatment, drug-induced cellular growth inhibition was significant 24 hr after drug exposure. The increased abundance of hypophosphorylated pRb and binding to transcription factor E2F-1 is consistent with ZD1694-induced cell growth inhibition in HCT-8 cells. Therefore, the observed effect on downstream events after effective inhibition of thymidylate synthase may offer the critical determinants of response to ZD1694.
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PMID:p53 and WAF1 are induced and Rb protein is hypophosphorylated during cell growth inhibition by the thymidylate synthase inhibitor ZD1694 (Tomudex). 910 28

The E2F family of transcription factors play a key role in G1-S progression. A dominant negative mutant (E2F97) of E2F1 containing the DNA binding domain of E2F1 under the control of a tetracycline-responsive promoter was constructed. Stable transfectants were produced in the pRb-lacking SaOS-2 cell line and SV40-transformed VA-13 cell line, respectively. Induction of E2F97 by tetracycline withdrawal resulted in strong inhibition of the E2F transcriptional activity and a decreased percentage of cells in S-phase. To understand the mechanism(s) by which E2F97 exerts its effect on the cell cycle, the effect of E2F97 on expression of various cell cycle proteins was examined. Upon induction of E2F97, a significant decrease in the levels of both dihydrofolate reductase and thymidylate synthase was observed in transfectants derived from both cell lines. Induction of E2F97 also led to a decrease in cyclin A and D1 protein levels. Regulation of cyclin A by E2F97 occurred at the transcriptional level. In addition, in VA13 cells, induction of E2F97 resulted in down-regulation of the tumor suppressor protein p53. These data suggest that E2F regulates both G1 and S-phase cyclins and that there may be a potential positive feedback regulatory loop between E2F and cyclin D1.
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PMID:Functional roles of E2F in cell cycle regulation. 912 68


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