Inactivation of Crk SH3 domain-binding guanine nucleotide-releasing factor (C3G) in cervical squamous cell carcinoma

Okino, K.; Nagai, Hisaki; Nakayama, Hiroki; Doi, Daisuke; Yoneyama, Kihei; Konishi, Hideyuki; Takeshita, Toshiyuki

doi:10.1111/j.1525-1438.2006.00352.x

Cited by 33 publications

(35 citation statements)

References 35 publications

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“…The physiological pro-apoptotic role of P2X 7 in these cells was previously demonstrated [7,8,13]. Previous studies in cervical cells have also identified tumor suppressor genes [33][34][35][36][37][38][39][40][41][42][43][44][45][46] and apoptosis-related genes [35,39,43,44,[46][47][48][49] that are regulated by changes in DNA methylation. However, until recently, little was known about the effects of DNA methylation on P2X 7 gene expression.…”

Section: Introductionmentioning

confidence: 99%

Regulation of P2X7 gene transcription

Zhou

Luo

et al. 2009

Purinergic Signalling

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The pro-apoptotic P2X 7 receptor regulates growth of epithelial cells. The objectives of the study were to understand P2X 7 gene transcription; to identify the active promoter and the transcription initiation site (TpIS); and to begin understanding regulation of P2X 7 gene transcription. Experiments in vitro utilized normal and cancerous cultured human uterine cervical epithelial cells, and HEK293 cells overexpressing P2X 7 -luciferase reporters. Experiments in vivo used surgical specimen of normal and cancerous uterine cervix. Assays involved DNA, RNA, and protein techniques. (a) The P2X 7 TpIS was localized to adenine (+1) at nt 1683 of the human P2X 7 gene [GenBank Y12851]), with a TTAAA sequence at nt −32/−28 and an active promoter region within nt −158/+32. (b) P2X 7 transcription was found to be regulated by two enhancers located at nt + 222/+232 and +401/+573 regions downstream of the active P2X 7 promoter. (c) The putative enhancer regions formed four DNA-protein complexes. (d) P2X 7 transcription was found to be controlled by hypermethylated cytosines at cytosine-phosphodiesterguanosines (CpG) that cluster or co-localize with the enhancers' sites. (e) We identified nine CpGs as inhibitory cis elements, and three CpG sites that are hypermethylated in cultured cervical epithelial cells and in cervix epithelia in vivo. (f) In cancer cervical cells, the degree of hypermethylation of the CpG sites was greater than in the normal cervical cells. Expression of the P2X 7 receptor is controlled by hypermethylated CpGs that flank transcription enhancers located within a 547-nt region downstream of the promoter.

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Section: Introductionmentioning

confidence: 99%

Regulation of P2X7 gene transcription

Zhou

Luo

et al. 2009

Purinergic Signalling

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“…We further analyzed a 373-bp region within CGI-A covering the CpG sites previously analyzed by Okino et al, in cervical squamous carcinomas (10). We found no traces of methylation in any of 29 tumor samples (Fig.…”

Section: Methylation Status Of Rapgef1 In Cervical and Ovarian Cancersmentioning

confidence: 63%

“…Recently, promoter hypermethylation of RAPGEF1 was suggested as a gene silencing mechanism in cervical squamous cell carcinomas (10). A frequency of hypermethylation of 38% (5/13) in cervical tumors of the strict-criterion CpG island (CGI-A), was reported using methylation-sensitive PCR (MSP) (11).…”

Section: Discussionmentioning

confidence: 99%

“…On the one hand, it has been found that overexpression of exogenous RAPGEF1 blocks the anchorage-independent growth induced by cotransfected, activated, sis, ras and v-raf oncogenes in fibroblasts (8); on the other hand, amplification, upregulation and overexpression of endogenous RAPGEF1 have been reported in small cell lung carcinomas, suggesting that this gene can play an oncogenic role through derangement of the CRK-Rap1 signaling pathway (9). More recently, this gene has been reported to undergo silencing through promoter hypermethylation in squamous cervical cancers (10).…”

Section: Introductionmentioning

confidence: 99%

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Frequent somatic demethylation of RAPGEF1/C3G intronic sequences in gastrointestinal and gynecological cancer

Samuelsson

Alonso²,

Ruiz-Larroya³

et al. 2011

Int J Oncol

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Abstract. RAPGEF1 (also known as C3G and GRF2) is a guanine nucleotide exchange factor that releases GDP from the inactive Rap1 protein, facilitating its subsequent activation by the binding of GTP. Rap1 plays regulatory roles in proliferation, differentiation and apoptosis. Amplification and overexpression of RAPGEF1 have been found in small cell lung cancers, suggesting an oncogenic role. In contrast, hypermethylation of a promoter CpG island (CGI-A) of RAPGEF1 has been reported in squamous cervical tumors, suggesting an anti-oncogenic role in these gynecological cancers. In our studies of DNA methylation alterations in gastrointestinal cancer we found somatic demethylation of a relaxed-criterion CpG island (CGI-B) located in the first intron of RAPGEF1 in 40% of colon cancers and 8% of gastric cancers relative to their matching normal tissues that were always methylated. We also found somatic demethylation in 47% of squamous cervical carcinomas as well as 33% of ovarian cancers. This somatic change in methylation, however, did not extend to the strict-criterion CpG island located in the promoter region (CGI-A) that was unmethylated in all normal and tumor tissues analyzed. Thus, promoter hypermethylation of RAPGEF1 seems insignificant in colorectal, cervical and ovarian cancers.In contrast, tumor-specific hypomethylation of the gene appears to be frequent in gastrointestinal and gynecological cancers. IntroductionOur previous analysis of gastrointestinal cancer by methylationsensitive amplification fragment length polymorphism (MS-AFLP) (1,2) identified several genomic sequences that exhibited methylation alterations in colorectal and gastric cancers (1). One of them (named band CA-8) appeared hypomethylated in colorectal and gastric cancers. Band CA-8 contains a NotI site located in the first intron of RAPGEF1 (GeneID: 2889, chr 9q34.3). This gene functions as a guanine nucleotide exchange factor (GEF) of Rap1 that is involved in multiple effector pathways. Rap1 is known to inhibit MAPK activity by blocking the signal transduction from Ras to c-Raf-1 (3-5), activate the MAPK cascades via B-Raf (6), and activate other members of the MAPK family of proteins (7). Thus, RAPGEF1 can exert pleiotropic effects, promoting or inhibiting growth in different cellular contexts.The role of RAPGEF1 in cancer progression is unclear. On the one hand, it has been found that overexpression of exogenous RAPGEF1 blocks the anchorage-independent growth induced by cotransfected, activated, sis, ras and v-raf oncogenes in fibroblasts (8); on the other hand, amplification, upregulation and overexpression of endogenous RAPGEF1 have been reported in small cell lung carcinomas, suggesting that this gene can play an oncogenic role through derangement of the CRK-Rap1 signaling pathway (9). More recently, this gene has been reported to undergo silencing through promoter hypermethylation in squamous cervical cancers (10).We therefore extended the methylation analysis to a panel of squamous cervical tumors to compare the apparent disparity...

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“…Our results indicate that paxillin phosphorylation in human colon cancer cells in response to pressure facilitates paxillin-Crk binding, which is also required for the stimulation of adhesion. Further study may clarify whether pressure-induced signaling prior to adhesion also involves such known Crk-binding partners in adherent cells as DOCK180 or C3G [50,51].…”

Section: Discussionmentioning

confidence: 99%

Pressure activates colon cancer cell adhesion via paxillin phosphorylation, Crk, Cas, and Rac1

Downey

Craig

Basson

2008

Cell. Mol. Life Sci.

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Physical forces can activate colon cancer cell adhesion, critical for metastasis. Paxillin is phosphorylated by FAK and required for pressure-stimulated adhesion. However, whether paxillin acts as an inert scaffolding protein or whether paxillin phosphorylation is required is unknown. Transfection with paxillin point-phosphorylation mutants demonstrated that phosphorylation at tyrosines 31 and 118 together is necessary for pressure-stimulated adhesion. We further evaluated potential paxillin partners. Reducing the adaptor protein Crk or the focal adhesion protein p130 Cas blocked pressure-stimulated adhesion. Furthermore, Crk and p130 Cas both displayed increased coimmunoprecipitation with paxillin in response to increased pressure, except in cells transfected with a Y31Y118 paxillin mutant. Inhibiting the small GTPase Rac1 also abolished pressurestimulated adhesion, and reducing paxillin by siRNA blocked Rac1 phosphorylation by pressure. Thus, paxillin phosphorylation at tyrosines 31 and 118 together is necessary for pressure-induced adhesion. Paxillin, Crk and Cas form a trimeric complex that activates Rac1 and mediates this effect.

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Inactivation of Crk SH3 domain-binding guanine nucleotide-releasing factor (C3G) in cervical squamous cell carcinoma

Cited by 33 publications

References 35 publications

Regulation of P2X7 gene transcription

Regulation of P2X7 gene transcription

Frequent somatic demethylation of RAPGEF1/C3G intronic sequences in gastrointestinal and gynecological cancer

Pressure activates colon cancer cell adhesion via paxillin phosphorylation, Crk, Cas, and Rac1

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