A comprehensive assessment of p53-responsive genes following adenoviral-p53 gene transfer in Bcl-2-expressing prostate cancer cells

Spurgers, Kevin B.; Coombes, Kevin R.; Meyn, Raymond E.; Gold, David; Logothetis, Christopher J.; Johnson, Terry J.; McDonnell, Timothy J.

doi:10.1038/sj.onc.1207293

Cited by 13 publications

(17 citation statements)

References 90 publications

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“…However, involvement of this mechanism seems unlikely because our experiments demonstrate no relationship between ␤ 1 integrin signaling and Mdm2 levels and suggest an alternative pathway involving Skp2 (see below). Furthermore, PC-3 (and Capan-1) cells exhibit inactivating mutations in the p53 gene (36,37).…”

Section: Discussionmentioning

confidence: 99%

Up-regulation of Skp2 after Prostate Cancer Cell Adhesion to Basement Membranes Results in BRCA2 Degradation and Cell Proliferation

Moro

Arbini

Marra

et al. 2006

Journal of Biological Chemistry

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Aberrant interaction of carcinoma cells with basement membranes (BM) is a fundamental pathophysiological process that initiates a series of events resulting in cancer cell invasion and metastasis. In this report, we describe the results of our investigations pertaining to the events triggered by the adhesion of normal (PNT1A) and highly metastatic (PC-3) prostate cells onto BM proteins. Unlike PNT1A, PC-3 cells adhered avidly to Matrigel BM matrix as well as to isolated collagen type IV, laminin, and heparan sulfate proteoglycan perlecan, main BM components. This aberrantly increased cancer cell adhesion resulted in sustained BRCA2 protein depletion and vigorous cell proliferation, a cascade triggered by ␤ 1 integrin-mediated phosphatidylinositol 3-kinase activation leading to BRCA2 degradation in the proteasome. This latter effect was orchestrated by phosphatidylinositol 3-kinase-dependent up-regulation of Skp2, a subunit of the Skp1-Cul1-F-box protein ubiquitin complex that directly associates with BRCA2 as demonstrated by coimmunoprecipitation assays, determines its ubiquitination, and ultimately targets it for proteasomal degradation. Inhibition of Skp2 expression by small interference RNA prevented BRCA2 depletion and inhibited the trophic effect upon cell proliferation. These results provide additional evidence on the role of BRCA2 as a modulator of cancer cell growth and elucidate the molecular mechanisms involved in its downregulation in cancer cells when interacting with BM, a crucial step in the biology of metastasis. Furthering the understanding of this molecular pathway may prove valuable in designing new therapeutic strategies aimed at modifying the natural history of prostate carcinoma. Basement membranes (BM)2 are thin layers of specialized extracellular matrix (ECM) that surround and closely associate with epithelial and endothelial cells, muscle fibers, and nerves. They consist mostly of collagen type IV (COL4) admixed with laminins (LN), nidogens, and the heparan sulfate proteoglycan perlecan (PLN) and may contain small amounts of fibronectin (FN) (1, 2). Although the BM structural role in defining tissue architecture and compartmentalization has long been recognized, its dynamic role in the modulation of cell behavior has only recently been documented (1).Aberrant cancer cell interactions with BM proteins play a crucial role in the biology of metastasis (3). Cancer cells must be able to coordinately decrease cell-cell interactions and increase cell adhesion to an adjacent BM in order to become motile, which along with the capacity of degrading/remodeling a BM directly relates to their metastatic potential (4). This cell behavior is accompanied by changes in the expression and/or usage of various adhesion receptors, including integrins (5). Integrins are transmembrane adhesion receptors for ECM proteins that not only provide physical anchoring cell support but also play a pivotal role in triggering intracellular signaling in response to environmental changes through interactions with molecules ...

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Section: Discussionmentioning

confidence: 99%

Up-regulation of Skp2 after Prostate Cancer Cell Adhesion to Basement Membranes Results in BRCA2 Degradation and Cell Proliferation

Moro

Arbini

Marra

et al. 2006

Journal of Biological Chemistry

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“…Cell culture conditions, along with the generation and characterization of stable vector control (PC3-BN) and BCL2-overexpressing (PC3-BCL2) PC3 cells have been described (13,14). Where noted, the cells were treated with etoposide (Sigma) dissolved in Me 2 SO added directly to the complete media of monolayer cultures (10 g/ml).…”

Section: Methodsmentioning

confidence: 99%

“…Construction of the recombinant, replication-deficient, type 5 adenovirus expressing human p53 (Ad-p53) and the experimental parameters and procedures used in this study have been described (13,15). Briefly, PC3 cells were infected with control virus expressing ␤-galactosidase (Ad-LacZ) or with Ad-p53 at 1,000 virus particles/cell.…”

Section: Methodsmentioning

confidence: 99%

Identification of Cell Cycle Regulatory Genes as Principal Targets of p53-mediated Transcriptional Repression

Spurgers

Gold

Coombes

et al. 2006

Journal of Biological Chemistry

108

104

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Historically, most studies attribute p53 function to the transactivation of target genes. That p53 can selectively repress genes to affect a cellular response is less widely appreciated. Available evidence suggests that repression is important for p53-induced apoptosis and cell cycle arrest. To better establish the scope of p53-repressed target genes and the cellular processes they may affect, a global expression profiling strategy was used to identify p53-responsive genes following adenoviral p53 gene transfer (Ad-p53) in PC3 prostate cancer cells. A total of 111 genes, 0.77% of the 14,500 genes represented on the Affymetrix U133A microarray, were repressed more than 2-fold ( p < 0.05). Validation of the array data, using reverse transcription-PCR of 20 randomly selected genes, yielded a confirmation rate of >95.5% for the complete data set. Functional over-representation analysis revealed that cell cycle regulatory genes exhibited a highly significant enrichment ( p < 5 ؋ 10 ؊28 ) within the transrepressed targets. 41% of the repressed targets are cell cycle regulators. A subset of these genes exhibited repression following DNA damage, preceding cell cycle arrest, in LNCaP cells. The use of a p53 small interfering RNA strategy in LNCaP cells and the use of p53-null cell lines demonstrated that this repression is p53-dependent. These findings identify a set of genes not known previously to be down-regulated by p53 and indicate that p53-induced cell cycle arrest is a function of not only the transactivation of cell cycle inhibitors (e.g. p21) but also the repression of targets that regulate proliferation at several distinct phases of the cell cycle.

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“…Computational methods predict that thousands of genes possess p53 DNA binding elements. However, functional p53 enhancer identification requires experimental validation [5].…”

Section: Introductionmentioning

confidence: 99%

The direct p53 target gene, FLJ11259/DRAM, is a member of a novel family of transmembrane proteins

Kerley-Hamilton

Pike

Hutchinson

et al. 2007

Biochimica et Biophysica Acta (BBA) - Gene Structure and Expres

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The tumor suppressor p53 regulates diverse biological processes primarily via activation of downstream target genes. Even though many p53 target genes have been described, the precise mechanisms of p53 biological actions are uncertain. In previous work we identified by microarray analysis a candidate p53 target gene, FLJ11259/DRAM. In this report we have identified three uncharacterized human proteins with sequence homology to FLJ11259, suggesting that FLJ11259 is a member of a novel family of proteins with six transmembrane domains. Several lines of investigation confirm FLJ11259 is a direct p53 target gene. p53 siRNA prevented cisplatin-mediated up-regulation of FLJ11259 in NT2/D1 cells. Likewise in HCT116 p53+/+ cells and MCF10A cells, FLJ11259 is induced by cisplatin treatment but to a much lesser extent in isogenic p53-suppressed cells. A functional p53 response element was identified 22.3kb upstream of the first coding exon of FLJ11259 and is shown to be active in reporter assays. In addition, chromatin immunoprecipitation assays indicate that p53 binds directly to this element in vivo and that binding is enhanced following cisplatin treatment. Confocal microscopy showed that an FLJ-GFP fusion protein localizes mainly in a punctate pattern in the cytoplasm. Overexpression studies in Cos-7, Saos2, and NT2/D1 cells suggest that FLJ11259 is associated with increased clonal survival. In summary, we have identified FLJ11259/DRAM as a p53-inducible member of a novel family of transmembrane proteins. FLJ11259/DRAM may be an important modulator of p53 responses in diverse tumor types.

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A comprehensive assessment of p53-responsive genes following adenoviral-p53 gene transfer in Bcl-2-expressing prostate cancer cells

Cited by 13 publications

References 90 publications

Up-regulation of Skp2 after Prostate Cancer Cell Adhesion to Basement Membranes Results in BRCA2 Degradation and Cell Proliferation

Up-regulation of Skp2 after Prostate Cancer Cell Adhesion to Basement Membranes Results in BRCA2 Degradation and Cell Proliferation

Identification of Cell Cycle Regulatory Genes as Principal Targets of p53-mediated Transcriptional Repression

The direct p53 target gene, FLJ11259/DRAM, is a member of a novel family of transmembrane proteins

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