UMLS:C0029463 - FACTA Search

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Query: UMLS:C0029463 (osteosarcoma)
16,637 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cell cycle-dependent transcription factor, E2F-1, regulates the cyclin-like species of the DNA repair enzyme uracil-DNA glycosylase (UDG) gene in human osteosarcoma (Saos-2) cells. We demonstrate, through the deletion of the human UDG promoter sequences, that expression of E2F-1 activates the UDG promoter through several E2F sites. The major putative downstream site for E2F, located in the first exon, serves as a target for E2F-1/DP1 complex binding in vitro. We also provide evidence for the functional relationship between the cyclin-like UDG gene product and E2F. High levels of UDG expression in a transient transfection assay result in the down-regulation of transcriptional activity through elements specific for E2F-mediated transcription. Overexpression of UDG in Saos 2 cells was observed to delay growth late in G1 phase and transiently arrest these cells from progressing into the S phase. This hypothetical model integrates one mechanism of DNA repair with the cell cycle control of gene transcription, likely through E2F. This implicates E2F as a multifunctional target for proteins and enzymes, possibly, responsive to DNA damage through the negative effect of UDG on E2F-mediated transcriptional activity.
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PMID:E2F-1 and a cyclin-like DNA repair enzyme, uracil-DNA glycosylase, provide evidence for an autoregulatory mechanism for transcription. 753 93

The E2F family of transcription factors controls the expression of genes that are involved in cell cycle regulation. E2F DNA-binding activity is found in complex with the retinoblastoma protein, pRb, and with the pRb-related p107 and p130. To date, cDNAs for three members of the E2F gene family have been isolated. However, all three E2Fs associate in vivo exclusively with pRb. We report here the cloning and functional analysis of a fourth E2F family member. E2F-4 encodes a 413-amino-acid protein with significant homology to E2F-1. E2F-4 antibodies recognize a 60-kD protein in anti-p107 immunoprecipitates, indicating that E2F-4 associates with p107 in vivo. Like the other E2Fs, E2F-4 requires DP-1 for efficient DNA binding and transcriptional activation of E2F site-containing promoters. Increased expression of E2F-4 and DP-1 in SaoS-2 osteosarcoma cells causes a shift from G1-phase cells to S and G2/M-phase cells, suggesting a role for E2F-4 in regulation of cell-cycle progression. We show that expression of E2F-4 and DP-1 together with an activated ras oncogene in rat embryo fibroblasts, causes transformation, indicating that E2F-4 has oncogenic activity.
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PMID:E2F-4, a new member of the E2F gene family, has oncogenic activity and associates with p107 in vivo. 795 25

Glucocorticoids inhibit proliferation of many cell types, but the events leading from the activated glucocorticoid receptor (GR) to growth arrest are not understood. Ectopic expression and activation of GR in human osteosarcoma cell lines U2OS and SAOS2, which lack endogenous receptors, result in a G1 cell cycle arrest. GR activation in U2OS cells represses expression of the cyclin-dependent kinases (CDKs) CDK4 and CDK6 as well as their regulatory partner, cyclin D3, leading to hypophosphorylation of the retinoblastoma protein (Rb). We also demonstrate a ligand-dependent reduction in the expression of E2F-1 and c-Myc, transcription factors involved in the G1-to-S-phase transition. Mitogen-activated protein kinase, CDK2, cyclin E, and the CDK inhibitors (CDIs) p27 and p21 are unaffected by receptor activation in U2OS cells. The receptor's N-terminal transcriptional activation domain is not required for growth arrest in U2OS cells. In Rb-deficient SAOS2 cells, however, the expression of p27 and p21 is induced upon receptor activation. Remarkably, in SAOS2 cells that express a GR deletion derivative lacking the N-terminal transcriptional activation domain, induction of CDI expression is abolished and the cells fail to undergo ligand-dependent cell cycle arrest. Similarly, murine S49 lymphoma cells, which, like SAOS2 cells, lack Rb, require the N-terminal activation domain for growth arrest and induce CDI expression upon GR activation. These cell-type-specific differences in receptor domains and cellular targets linking GR activation to cell cycle machinery suggest two distinct regulatory mechanisms of GR-mediated cell cycle arrest: one involving transcriptional repression of G1 cyclins and CDKs and the other involving enhanced transcription of CDIs by the activated receptor.
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PMID:Glucocorticoid receptor-mediated cell cycle arrest is achieved through distinct cell-specific transcriptional regulatory mechanisms. 915 17

The effect of overexpression of p21waf1 on drug sensitivity was studied in an osteosarcoma cell line (SaOs-2) lacking both p53 and functional retinoblastoma protein using a tetracycline (TC)-inducible expression system. p21waf1 expression was barely detectable in SaOS-2 cells incubated in the presence of TC. After TC withdrawal, high levels of p21waf1 were induced in these cells. These p21waf1-induced cells showed increased sensitivity to doxorubicin, tomudex, and methotrexate as compared to uninduced cells; this condition is associated with increased apoptosis. Expression of p21waf1 reduced cyclin A-associated kinase activity and, surprisingly, resulted in inhibition of phosphorylation of E2F-1 and increased E2F-1 binding activity. An S-G2 cell cycle arrest/delay and an increase in expression of E2F-responsive genes (dihydrofolate reductase and thymidylate synthase) was correspondingly observed. Overexpression of p21waf1 in cells lacking functional retinoblastoma protein may mediate sensitivity to anticancer drugs by inhibiting E2F-1 phosphorylation, which may contribute to increased S-G2 cell cycle delay and increased cell susceptibility to apoptosis.
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PMID:Overexpression of p21waf1 leads to increased inhibition of E2F-1 phosphorylation and sensitivity to anticancer drugs in retinoblastoma-negative human sarcoma cells. 918 20

The retinoblastoma tumor suppressor gene product (pRb) is involved in controlling cell cycle progression from G1 into S. pRb functions, in part, by regulating the activities of several transcription factors, making pRb involved in the transcriptional control of cellular genes. Transient-transfection assays have implicated pRb in the transcription of several genes, including c-fos, the interleukin-6 gene, c-myc, cdc-2, c-neu, and the transforming growth factor beta2 gene. However, these assays place the promoter in an artificial context and exclude the effects of far 5' upstream regions and chromosomal architecture on gene transcription. In these experiments, we have studied the role of pRb in the control of cell cycle-related genes within a chromosomal context and within the context of the G1 phase of the cell cycle. We have used adenovirus vectors to overexpress pRb in human osteosarcoma cells and breast cells synchronized in early G1. By RNase protection assays, we have assayed the effects of this virus-produced pRb on gene expression in these cells. These results indicate that pRb is involved in the transcriptional downregulation of the E2F-1, E2F-2, dihydrofolate reductase, thymidine kinase, c-myc, proliferating-cell nuclear antigen, p107, and p21/Cip1 genes. However, it has no effect on the transcription of the E2F-3, E2F-4, E2F-5, DP-1, DP-2, or p16/Ink4 genes. The results are consistent with the notion that pRb controls the transcription of genes involved in S-phase promotion. They also suggest that pRb negatively regulates the transcription of two of the transcription factors whose activity it also represses, E2F-1 and E2F-2, and that it plays a role in downregulating the immediate-early gene response to serum stimulation.
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PMID:Regulation of cellular genes in a chromosomal context by the retinoblastoma tumor suppressor protein. 967 66

The retinoblastoma (RB) gene is one of the most extensively studied tumour-suppressor genes. Deletion or inactivation of both RB alleles is an essential, rate-limiting step in the formation of retinoblastoma and osteosarcoma that arise in families that carry mutant RB (ref. 2). RB inactivation is also found in other human tumours. Whereas loss of RB function is associated with the loss of cellular proliferative control, introduction of a wild-type RB can suppress cell growth and tumorigenicity. Thus, identification of factors that interfere with and/or control the function of the RB protein is critical for understanding both cell-cycle control and oncogenesis. Here we describe a new gene, Bog (for B5T over-expressed gene), which was identified and shown to be overexpressed in several transformed rat liver epithelial (RLE) cell lines resistant to the growth-inhibitory effect of TGF-beta1, as well as in primary human liver tumours. The Bog protein shares homology with other retinoblastoma-binding proteins and contains the Rb-binding motif LXCXE. Using the yeast two-hybrid system and co-immunoprecipitation, we demonstrated that Bog binds to Rb. In vivo, Bog/Rb complexes do not contain E2F-1, and Bog can displace E2F-1 from E2F-1/Rb complexes in vitro. Overexpression of Bog in normal RLE cells conferred resistance to the growth-inhibitory effect of TGF-beta1. Furthermore, normal RLE cells are rapidly transformed when Bog is continuously overexpressed and form hepatoblastoma-like tumours when transplanted into nude mice. These data suggest that Bog may be important in the transformation process, in part due to its capacity to confer resistance to the growth-inhibitory effects of TGF-beta1 through interaction with Rb and the subsequent displacement of E2F-1.
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PMID:A retinoblastoma-binding protein that affects cell-cycle control and confers transforming ability. 969 99

We previously demonstrated that P16Ink4a (p16) expression in p16-deficient U343 astrocytoma cells causes a G1 cell cycle arrest, profound changes in cytoskeletal proteins and alterations in expression and activity of the pRB and E2F family proteins. We examine here the effects of expressing wild type or mutant versions of the downstream targets of p16 in U343 astrocytomas. We first attempted to block proliferation of U343 cells using the dominant mutant of pRB, deltap34. Expression of this mutant in the human osteosarcoma, SAOS-2, potently blocked proliferation but did not affect the cell cycle of U343 cells. We next showed that expression of E2F-1, E2F-2, E2F-3 and E2F-4 are each able to overcome this p16-dependent cell cycle arrest but exhibit distinct biological activities. Adenoviral-mediated expression of E2F-1, E2F-2, E2F-3, or E2F-4 overcame the p16-dependent cell cycle block and induced alterations in cell morphology. E2F-5, only in conjunction with DP1, promoted cell cycle progression. For both E2F-1 and E2F-2, but not E2F-3 or E2F-5/DP1, cell cycle re-entry was associated with almost quantitative cell death. Only small numbers of dying cells were observed in E2F-4-expressing cultures. Expression of the different E2F's altered the expression of distinct sets of cell cycle regulatory proteins. E2F-1 induced endogenous E2F-4 expression and also caused an increase in pRB, p107 and cyclin E levels. Expression of E2F-4 caused a weak increase in E2F-1 levels but also strongly induced pRB, p107, p130 and cyclin E. However, E2F-1 and E2F-4 clearly regulate expression of distinct genes, demonstrated when E2F-4 caused a threefold increase in the levels of cdk2 whereas E2F-1 failed to increase in this cyclin dependent kinase. Similarly, expression of E2F-1 or E2F-2 were shown to have distinct effects on the expression of cdk2, cyclin E and pRB despite both of these closely related E2F-family members potently inducing cell death. Thus, E2F-1, E2F-2, E2F-3 and E2F-4 are able to overcome the p16-dependent proliferative block in U343 astrocytoma cells. While overcoming this cell cycle block, each of the E2F's uniquely affect the expression of a number of cell cycle regulatory proteins and have distinct abilities to promote cell death.
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PMID:The E2F-family proteins induce distinct cell cycle regulatory factors in p16-arrested, U343 astrocytoma cells. 978 3

Human cyclin A1, a newly discovered cyclin, is expressed in testis and is thought to function in the meiotic cell cycle. Here, we show that the expression of human cyclin A1 and cyclin A1-associated kinase activities was regulated during the mitotic cell cycle. In the osteosarcoma cell line MG63, cyclin A1 mRNA and protein were present at very low levels in cells at the G0 phase. They increased during the progression of the cell cycle and reached the highest levels in the S and G2/M phases. Furthermore, the cyclin A1-associated histone H1 kinase activity peaked at the G2/M phase. We report that cyclin A1 could bind to important cell cycle regulators: the Rb family of proteins, the transcription factor E2F-1, and the p21 family of proteins. The in vitro interaction of cyclin A1 with E2F-1 was greatly enhanced when cyclin A1 was complexed with CDK2. Associations of cyclin A1 with Rb and E2F-1 were observed in vivo in several cell lines. When cyclin A1 was coexpressed with CDK2 in sf9 insect cells, the CDK2-cyclin A1 complex had kinase activities for histone H1, E2F-1, and the Rb family of proteins. Our results suggest that the Rb family of proteins and E2F-1 may be important targets for phosphorylation by the cyclin A1-associated kinase. Cyclin A1 may function in the mitotic cell cycle in certain cells.
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PMID:Functions of cyclin A1 in the cell cycle and its interactions with transcription factor E2F-1 and the Rb family of proteins. 1002 26

We have focused on the roles of PARP and poly(ADP-ribosyl)ation early in apoptosis, as well as during the early stages of differentiation-linked DNA replication. In both nuclear processes, a transient burst of PAR synthesis and PARP expression occurs early, prior to internucleosomal DNA cleavage before commitment to apoptosis as well as at the round of DNA replication prior to the onset of terminal differentiation. In intact human osteosarcoma cells undergoing spontaneous apoptosis, both PARP and PAR decreased after this early peak, concomitant with the inactivation and cleavage of PARP by caspase-3 and the onset of substantial DNA and nuclear fragmentation. Whereas 3T3-L1, osteosarcoma cells, and immortalized PARP +/+ fibroblasts exhibited this early burst of PAR synthesis during Fas-mediated apoptosis, neither PARP-depleted 3T3-L1 PARP-antisense cells nor PARP -/- fibroblasts showed this response. Consequently, whereas control cells progressed into apoptosis, as indicated by induction of caspase-3-like PARP-cleavage activity, PARP-antisense cells and PARP -/- fibroblasts did not, indicating a requirement for PARP and poly(ADP-ribosyl)ation of nuclear proteins at an early reversible stage of apoptosis. In parallel experiments, a transient increase in PARP expression and activity were also noted in 3T3-L1 preadipocytes 24 h after induction of differentiation, a stage at which approximately 95% of the cells were in S-phase, but not in PARP-depleted antisense cells, which were consequently unable to complete the round of DNA replication required for differentiation. PARP, a component of the multiprotein DNA replication complex (MRC) that catalyzes viral DNA replication in vitro, poly(ADP-ribosyl)ates 15 of approximately 40 MRC proteins, including DNA pol alpha, DNA topo I, and PCNA. Depletion of endogenous PARP by antisense RNA expression in 3T3-L1 cells results in MRCs devoid of any DNA pol alpha and DNA pol delta activities. Surprisingly, there was no new expression of PCNA and DNA pol alpha, as well as the transcription factor E2F-1 in PARP-antisense cells during entry into S-phase, suggesting that PARP may play a role in the expression of these proteins, perhaps by interacting with a site in the promoters for these genes.
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PMID:Involvement of PARP and poly(ADP-ribosyl)ation in the early stages of apoptosis and DNA replication. 1033 50

The transcription factor complex E2F-1/DP-1 regulates the G1-to-S-phase transition and has been associated with sensitivity to the S-phase-specific anticancer agents camptothecin and etoposide, which poison DNA topoisomerase I and II, respectively. To investigate the relationship between E2F-1 and drug sensitivity in detail, we established human osteosarcoma U-20S-TA cells expressing full-length E2F-1/ DP-1 under the control of a tetracycline-responsive promoter, designated UE1DP-1 cells. Topoisomerase I levels and activity as well as the number of camptothecin-induced DNA single- and double-strand breaks were unchanged in UEIDP-1/tc- cells with >10-fold E2F-1/DP-1 overexpression. However, UE1DP-1/tc- cells were hypersensitive to camptothecin in both a clonogenic assay and four different apoptotic assays. This indicates that camptothecin-induced toxicity in this model is due to the activation of an E2F-1/ DP-1-induced post-DNA damage pathway rather than an increase in the number of replication forks caused by the S-phase initiation. In contrast, topoisomerase IIalpha levels (but not topoisomerase IIbeta levels), together with topoisomerase IIalpha promoter activity, increased 2--3-fold in UE1DP-1/tc-cells. Furthermore, the number of etoposide-induced DNA single- and double-strand breaks increased in UE1DP-1/tc-cells together with a rise in clonogenic sensitivity to etoposide, but an equal apoptotic sensitivity to etoposide. The increase in topoisomerase IIalpha promoter activity in UE1DP-1/tc--cells was shown to be due to S-phase initiation per se because it was blocked by ectopic expression of dominant negative cyclin-dependent kinase 2. In conclusion, overexpression of E2F-1/DP-1 in U-20S-TA cells is sufficient to increase clonogenic sensitivity to both topoisomerase I- and II-targeted anticancer drugs. However, the mechanism by which this occurs appears to be qualitatively different. The UE1DP-1 cell model may be used to elucidate post-DNA damage mechanisms of cell death induced by topoisomerase I-directed anticancer agents.
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PMID:Differential cytotoxic pathways of topoisomerase I and II anticancer agents after overexpression of the E2F-1/DP-1 transcription factor complex. 1077 81

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