研究者詳細 - 時野　隆至

論文 - 時野　隆至

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Id2 protein is selectively upregulated by UVB in primary, but not in immortalized human keratinocytes and inhibits differentiation.

Simbulan-Rosenthal CM, Trabosh V, Velarde A, Chou FP, Daher A, Tenzin F, Tokino T, Rosenthal DS

Oncogene 24 ( 35 ) 5443 - 58 2005年08月 [査読有り] [国際誌]

Solar ultraviolet B (UVB) acts as both an initiator and promoter in models of multistage skin carcinogenesis. We found that, whereas UVB induces apoptosis in human papillomavirus-16 E6/7-immortalized keratinocytes, it inhibits markers of differentiation in human foreskin keratinocytes (HFK). Potential mechanisms for this differential response were examined by DNA microarray, which revealed that UVB alters the expression of three of the four human inhibitor of differentiation/DNA binding (Id) proteins that comprise a class of helix-loop-helix family of transcription factors involved in proliferation, differentiation, apoptosis, and carcinogenesis. These results were verified by RT-PCR and immunoblot analysis of control and UVB-irradiated primary and immortalized keratinocytes. Whereas Id1 was downregulated in both cell types, Id2 expression was upregulated in primary HFK, but not immortalized cells. In contrast, Id3 expression was significantly increased only in immortalized cells. The differential expression pattern of Id2 in response to UVB was recapitulated in reporter constructs containing the 5' regulatory regions of this gene. Id2 promoter activity increased in response to UVB in HFK, but not in immortalized cells. To identify the regulatory elements in the Id2 promoter that mediate transcriptional activation by UVB in HFK, promoter deletion/mutation analysis was performed. Deletion analysis revealed that transactivation involves a 166 bp region immediately upstream to the Id2 transcriptional start site and is independent of c-Myc. The consensus E twenty-six (ETS) binding site at -120 appears to mediate UVB transcriptional activation of Id2 because point mutations at this site completely abrogated this response. Chromatin immunoprecipitation and electrophoretic mobility-shift assays verified that the Id2 promoter interacts with known Id2 promoter (ETS) binding factors Erg1/2 and Fli1, but not with c-Myc; and this interaction is enhanced after UVB exposure. Similar to the effects of UVB exposure, ectopic expression of Id2 protein in primary HFK resulted in inhibition of differentiation, as shown by decreased levels of the terminal differentiation marker keratin K1 and inhibition of involucrin crosslinking. Reduction of Id2 expression by small interfering RNAs attenuated the UVB-induced inhibition of differentiation in these cells. These results suggest that UVB-induced inhibition of differentiation of primary HFK is at least, in part, due to the upregulation of Id2, and that upregulation of Id2 by UVB might predispose keratinocytes to carcinogenesis by preventing their normal differentiation program.

DOI PubMed

Identification of pigment epithelium-derived factor as a direct target of the p53 family member genes.

Yasushi Sasaki, Yasuyoshi Naishiro, Yuichiro Oshima, Kohzoh Imai, Yusuke Nakamura, Takashi Tokino

Oncogene 24 ( 32 ) 5131 - 6 2005年07月 [査読有り] [国際誌]

p63 and p73 show a high degree of structural homology to p53 and are members of a family of transcriptional factors that can activate transcription of p53-responsive genes. p53 is mutated in more than 50% of human cancers, whereas p63 and p73 are rarely mutated. Studies of knockout mice also revealed an unexpected functional diversity among the p53 family. To determine how p63 and p73 are involved in tumorigenesis and normal development, we used cDNA microarray to examine 9216 genes in human colorectal cancer cells. We discovered that the expression of pigment epithelium-derived factor (PEDF) was specifically induced by either p63 or p73, but not by p53. We also report here that the PEDF gene contains a response element specific for p63 and p73 in its promoter region and is a direct target of p63 and p73. Collectively, p63 and p73 may be involved in cell fate by inducing PEDF expression.

PubMed

The Ras effector RASSF2 is a novel tumor-suppressor gene in human colorectal cancer.

Akino K, Toyota M, Suzuki H, Mita H, Sasaki Y, Ohe-Toyota M, Issa JP, Hinoda Y, Imai K, Tokino T

Gastroenterology 129 ( 1 ) 156 - 69 2005年07月 [査読有り] [国際誌]

BACKGROUND & AIMS: Activation of Ras signaling is a hallmark of colorectal cancer (CRC), but the roles of negative regulators of Ras are not fully understood. Our aim was to address that question by surveying genetic and epigenetic alterations of Ras-Ras effector genes in CRC cells. METHODS: The expression and methylation status of 6 RASSF family genes were examined using RT-PCR and bisulfite PCR in CRC cell lines and in primary CRCs and colorectal adenomas. Colony formation assays and flow cytometry were used to assess the tumor suppressor activities of RASSF1 and RASSF2. Immunofluorescence microscopy was used to determine the effect of altered RASSF2 expression on cell morphology. Mutations of K- ras , BRAF, and p53 were identified using single-strand conformation analysis and direct sequencing. RESULTS: Aberrant methylation and histone deacetylation of RASSF2 was associated with the gene's silencing in CRC. The activities of RASSF2, which were distinct from those of RASSF1, included induction of morphologic changes and apoptosis; moreover, its ability to prevent cell transformation suggests that RASSF2 acts as a tumor suppressor in CRC. Primary CRCs that showed K- ras /BRAF mutations also frequently showed RASSF2 methylation, and inactivation of RASSF2 enhanced K- ras -induced oncogenic transformation. RASSF2 methylation was also frequently identified in colorectal adenomas. CONCLUSIONS: RASSF2 is a novel tumor suppressor gene that regulates Ras signaling and plays a pivotal role in the early stages of colorectal tumorigenesis.

DOI PubMed

Small interfering RNA-induced CHFR silencing sensitizes oral squamous cell cancer cells to microtubule inhibitors.

Ogi K, Toyota M, Mita H, Satoh A, Kashima L, Sasaki Y, Suzuki H, Akino K, Nishikawa N, Noguchi M, Shinomura Y, Imai K, Hiratsuka H, Tokino T

Cancer biology & therapy 4 ( 7 ) 773 - 80 2005年07月 [査読有り] [国際誌]

Alterations in the function of cell cycle checkpoints are frequently detected in oral squamous cell carcinomas (OSCCs), and are often associated with the sensitivity of the cancer cells to chemotherapeutic drugs. Recently, a mitotic checkpoint gene, Chfr, was shown to be inactivated by promoter methylation and point mutations in various human tumors. Here we show that the absence of its product, CHFR, is associated with mitotic checkpoint dysfunction, and that cancer cells lacking CHFR are sensitive to microtubule inhibitors. Checkpoint impairment appears to be caused by a prophase defect in this case, as OSCC cells lacking CHFR showed phosphorylation of histone H3 on Ser10 and translocation of cyclin B1 to the nucleus. When CHFR-deficient OSCC cells were treated with a microtubule inhibitor (docetaxel or paclitaxel), significant numbers of apoptotic cells were observed. Moreover, disruption of CHFR using small interfering RNA (siRNA) impaired the mitotic checkpoint, thereby reducing the ability of OSCC cells to arrest at G2/M phase and making them more sensitive to microtubule inhibitors. Our results suggest that CHFR could be a useful molecular target for chemotherapy.

PubMed

p53によるビタミンD受容体の発現誘導と癌治療への応用

丸山玲緒, 豊田実, 青木文夫, 佐々木泰史, 明石浩史, 見田裕章, 鈴木拓, 時野隆至, 今井浩三, 篠村恭久

日本臨床分子医学会学術総会プログラム・抄録集 ( 日本臨床分子医学会 ) 42回 112 - 112 2005年07月

Upregulation of BNIP3 by 5-aza-2'-deoxycytidine sensitizes pancreatic cancer cells to hypoxia-mediated cell death.

Abe T, Toyota M, Suzuki H, Murai M, Akino K, Ueno M, Nojima M, Yawata A, Miyakawa H, Suga T, Ito H, Endo T, Tokino T, Hinoda Y, Imai K

Journal of gastroenterology 40 ( 5 ) 504 - 10 2005年05月 [査読有り] [国内誌]

BACKGROUND: Pancreatic cancer cells often show resistance to hypoxia-mediated apoptosis, but the molecular mechanism underlying that resistance remains unknown. The purpose of the present study, therefore, was to examine the role of epigenetic gene alteration in the resistance to hypoxia-mediated apoptosis among pancreatic cancer cells. METHODS: Reverse transcription-polymerase chain reaction (RT-PCR) was used to examine the expression of five genes associated with hypoxia-mediated apoptosis (PUMA, Caspase-8 [CASP8], APAF-1, BNIP3, and BNIP3L) in a panel of pancreatic cancer cell lines. Protein expression was examined by Western blot analysis, using lysates from cells incubated under normoxic or hypoxic conditions. The methylation status of the genes was determined using bisulfite-PCR and sequencing. The percentages of cells that were apoptotic were determined using flow cytometry. RESULTS: Under normoxic conditions, the expression of the BNIP3 gene varied among the 12 pancreatic cancer cell lines tested, with 50% of them showing no BNIP3 expression at all, whereas expression of the other four genes was readily detected in all 12 cell lines. DNA methylation of BNIP3's CpG island in the region around the transcription start site of the gene was closely associated with its silencing. The expression of BNIP3 was restored by the methyltransferase inhibitor 5-aza-deoxycytidine (5-aza-dC), as was the hypoxia-mediated pancreatic cancer cell death. CONCLUSIONS: BNIP3 expression is silenced in some pancreatic cancer cells by the methylation of its CpG island. Demethylation of BNIP3, using a methyltransferase inhibitor, restores the gene's expression and induces hypoxia-mediated cell death. BNIP3 may thus be a useful target for new therapies aimed at treating pancreatic cancer.

DOI PubMed

Aberrant laminin beta3 isoforms downstream of EWS-ETS fusion genes in Ewing family tumors.

Hideto Irifune, Hiroyuki Nishimori, Goichi Watanabe, Kouichi Yoshida, Tatsuru Ikeda, Chihiro Matsui, Masaaki Morohashi, Satoshi Kawaguchi, Satoshi Nagoya, Takuro Wada, Toshihiko Yamashita, Yusuke Nakamura, Takashi Tokino

Cancer biology & therapy 4 ( 4 ) 449 - 55 2005年04月 [査読有り] [国際誌]

Ewing family tumors (EFTs) are associated with a chromosomal translocation resulting in a fusion of the amino-terminus of EWS with the DNA-binding domain of an ETS transcription factor. Although previous reports suggested that these chimeric proteins would act as aberrant transcription factors, their downstream targets have not been fully elucidated. To identify downstream targets of these EWS-ETS fusion proteins, we introduced EWS-ETS fusion constructs into a human fibrosarcoma cell line, HT-1080, by retroviral transduction. Here we report that the LAMB3 gene encoding the beta3 chain of basement membrane protein laminin-5 is induced to a significantly higher level in cells expressing EWS-ETSs than in cells expressing normal ETSs. Additionally through use of an antisense oligonucleotide for EWS-ERG in the W-ES EFT cell line, laminin beta3 protein was reduced coordinately with EWS-ERG fusion protein expression. Furthermore, we found small mRNAs were preferentially transcribed from the LAMB3 gene in EFT cell lines. Molecular cloning of the entire coding region shows that the alternative transcripts from different promoter(s) located within the intron 14, which encode small proteins, likely are major products of the LAMB3 gene in EFT cells. We show that the small isoforms conferred increased anchorage-independent proliferation to NIH3T3 cells. Together with previous studies showing that laminin-5 is involved in the invasive and malignant phenotype of several tumor types, our data suggest that the oncogenic effect of EWS-ETS may be mediated in part by upregulation of LAMB3 expression.

PubMed

Abnormal beta-catenin expression in oral cancer with no gene mutation: correlation with expression of cyclin D1 and epidermal growth factor receptor, Ki-67 labeling index, and clinicopathological features.

Tetsuyo Odajima, Yasushi Sasaki, Nobuyuki Tanaka, Yuko Kato-Mori, Hiroko Asanuma, Tatsuru Ikeda, Masaaki Satoh, Hiroyoshi Hiratsuka, Takashi Tokino, Norimasa Sawada

Human pathology 36 ( 3 ) 234 - 41 2005年03月 [査読有り] [国際誌]

Beta-Catenin not only acts as a regulator of E-cadherin-mediated cell-cell adhesion but also plays an important role in Wnt signaling. To assess the prevalence of Wnt signaling, we examined beta-catenin mutation and its immunohistochemical protein expression in oral cancers. The results were linked with expression of cyclin D1, one of the target genes of Wnt signaling, expression of epidermal growth factor receptor (EGFR) relevant to beta-catenin tyrosine phosphorylation, Ki-67 labeling index, clinicopathological features, and survival. In the analysis based on membranous expression of beta-catenin, 75 (68.2%) of 110 cases showed a reduced membranous pattern, and the remaining 35 (31.8%) had a preserved membranous pattern similar to that in oral epithelium. In the analysis of another category of beta-catenin expression, a cytoplasmic/nuclear pattern was observed in 21 (19.1%) of the 110 tumors. Most (19/21, 90.5%) of these tumors had a concomitant reduction of membranous expression of beta-catenin. The reduced membranous or cytoplasmic/nuclear pattern of beta-catenin was significantly associated with an invasive growth pattern, EGFR expression, an increased Ki-67 labeling index, and shorter survival but not with cyclin D1 expression. Mutational analyses of beta-catenin were performed for 39 cases, including the 21 tumors with a cytoplasmic/nuclear pattern, but no mutations in the beta-catenin gene exon 3 were detected in these samples. Our data indicate that altered expression of beta-catenin may play an important role in tumor progression through increased proliferation and invasiveness under EGFR activation. However, mutations of beta-catenin do not appear to be responsible for tumor development and abnormal expression of beta-catenin in oral cancers.

DOI PubMed

Aberrant DNA methylation associated with silencing BNIP3 gene expression in haematopoietic tumours

M Murai, M Toyota, A Satoh, H Suzuki, K Akino, H Mita, Y Sasaki, T Ishida, L Shen, G Garcia-Manero, JPJ Issa, Y Hinoda, T Tokino, K Imai

BRITISH JOURNAL OF CANCER ( NATURE PUBLISHING GROUP ) 92 ( 6 ) 1165 - 1172 2005年03月

Hypoxia is a key factor contributing to the progression of human neoplasias and to the development of resistance to chemotherapy. BNIP3 is a proapoptotic member of the Bcl-2 protein family involved in hypoxia-induced cell death. We evaluated the expression and methylation status of BNIP3 gene to better understand the role of epigenetic alteration of its expression in haematopoietic tumours. Methylation of the region around the BNIP3 transcription start site was detected in four acute lymphocytic leukaemia, one multiple myeloma and one Burkitt lymphoma cell lines, and was closely associated with silencing the gene. That expression of BNIP3 was restored by treatment with 5-aza2 '-deoxycytidine (5-aza-dC), a methyltransferase inhibitor, which confirmed the gene to be epigenetically inactivated by methylation. Notably, re-expression of BNIP3 using 5-aza2-dC also restored hypoxia-mediated cell death in methylated cell lines. Acetylation of histone H3 in the 5 ' region of the gene, which was assessed using chromatin immunoprecipitation assays, correlated directly with gene expression and inversely with DNA methylation. Among primary tumours, methylation of BNIP3 was detected in five of 34 (15%) acute lymphocytic leukaemias, six of 35 (17%) acute myelogenous leukaemias and three of 14 (21%) multiple myelomas. These results suggest that aberrant DNA methylation of the 5 ' CpG island and histone deacetylation play key roles in silencing BNIP3 expression in haematopoietic tumours.

DOI

Aberrant methylation and silencing of the BNIP3 gene in colorectal and gastric cancer.

Masafumi Murai, Minoru Toyota, Hiromu Suzuki, Ayumi Satoh, Yasushi Sasaki, Kimishige Akino, Masako Ueno, Fumihiko Takahashi, Masanobu Kusano, Hiroaki Mita, Kazuyoshi Yanagihara, Takao Endo, Yuji Hinoda, Takashi Tokino, Kohzoh Imai

Clinical cancer research : an official journal of the American Association for Cancer Research 11 ( 3 ) 1021 - 7 2005年02月 [査読有り] [国際誌]

BNIP3 protein is a proapoptotic member of the Bcl-2 family that is expressed in hypoxic regions of tumors. To examine its role in the progression of gastrointestinal cancer, we examined the expression and DNA methylation status of BNIP3 gene in a panel of colorectal and gastric cancer cell lines. BNIP3 was not expressed in 14 of the 24 cell lines tested, and its absence was not caused by gene mutation or by altered expression of hypoxia inducible factor-1, a key transcription factor that regulates BNIP3 expression. On the other hand, methylation of the 5' CpG island of BNIP3 was closely correlated with silencing the gene. Moreover, treating methylated cells with the methyltransferase inhibitor 5-aza-2'-deoxycytidine restored hypoxia-induced expression of BNIP3 mRNA and protein, which in turn led to cell death. Aberrant methylation of BNIP3 was also detected in 66% of primary colorectal and 49% of primary gastric cancers, but not in normal tissue samples collected from areas adjacent to the tumors. Apparently, epigenetic alteration of BNIP3 is a frequent and cancer-specific event, which suggests that inactivation of BNIP3 likely plays a key role in the progression of some gastrointestinal cancers and that it may be a useful molecular target for therapy.

PubMed

Enhanced Bax in oral SCC in relation to antitumor effects of chemotherapy.

Kanako Takemura, Makoto Noguchi, Kazuhiro Ogi, Takashi Tokino, Hiromi Kubota, Akihiro Miyazaki, Geniku Kohama, Hiroyoshi Hiratsuka

Journal of oral pathology & medicine : official publication of the International Association of Oral Pathologists and the American Academy of Oral Pathology 34 ( 2 ) 93 - 9 2005年02月 [査読有り] [国際誌]

BACKGROUND: Antitumor effects of chemotherapeutic agents are commonly associated with the induction of apoptosis. Bax belongs to the Bcl-2 family and induces apoptosis. The present study was conducted to investigate the relationship between enhanced Bax expression in oral squamous cell carcinoma (SCC; cell lines and clinical cases) and the antitumor effects of chemotherapy. METHODS: In three oral SCC cell lines, Bax expression before and after treatment with chemotherapeutic agents [docetaxel (TXT), cisplatin and 5-fluorouracil] was examined by reverse transcriptase-polymerase chain reaction and immunoblotting. The effects of treatment were assessed by counting the number of viable cells and determining sub-G1 cells. Tissue samples (both biopsy specimens before chemotherapy and surgically excised specimens after chemotherapy) from nine patients with oral SCC who underwent neoadjuvant chemotherapy were immunostained for Bax. The relationship between enhancement of Bax expression and chemotherapeutic effects was established. RESULTS: Two of three cell lines did not express Bax mRNA or protein before treatment. After treatment, Bax expression was enhanced only by TXT in one cell line, but by all chemotherapeutic agents in the other two cell lines. In three of nine patients, Bax expression was not found before chemotherapy. Two of these three patients showed enhanced Bax expression after chemotherapy including TXT, but one still failed to express Bax. Both in cell lines and clinical cases, enhancement of Bax after chemotherapy was associated with antitumor effects. CONCLUSION: Certain chemotherapeutic agents enhance Bax expression in oral SCC, and it is suggested that this contributes to the antitumor effects of chemotherapy.

DOI PubMed

Tight junction protein MAGI-1 is up-regulated by transfection with connexin 32 in an immortalized mouse hepatic cell line: cDNA microarray analysis.

Masaki Murata, Takashi Kojima, Toshinobu Yamamoto, Mitsuru Go, Ken-ichi Takano, Hideki Chiba, Takashi Tokino, Norimasa Sawada

Cell and tissue research 319 ( 2 ) 341 - 7 2005年02月 [査読有り] [国際誌]

Gap junctions are considered to play a crucial role in differentiation of epithelial cells, including hepatocytes. Recently, we found that Cx32 but not Cx26 was closely related to tight junctional proteins in primary cultured rat hepatocytes (Kojima et al., Exp Cell Res 263:193-201, 2001) and that Cx32 formation and/or Cx32-mediated intercellular communication could induce expression and function of tight junctions in a mouse hepatic cell line (Kojima et al., Exp Cell Res 276:40-51, 2002). In this study, to investigate the mechanisms of induction of tight junctions by transfection with Cx32, we performed cDNA microarray analysis of Cx32 transfectants, compared with parental cells derived from Cx32-deficient hepatocytes. In cDNA microarray analysis, a 2.5-fold increase in expression of membrane-associated guanylate kinase with inverted orientation-1 (MAGI-1), which is known to be localized at adherens and tight junction regions, was observed. High expression of MAGI-1 in Cx32 transfectants was confirmed by Western blotting and RT-PCR. MAGI-1 was colocalized with occludin, claudin-2, ZO-1, and F-actin, but not with E-cadherin in the apical-most regions at cell borders of Cx32 transfectants, similar to junctional adhesion molecule-1 (JAM-1), which may play a crucial role in formation and assembly of tight junctions. Treatment with the gap junction blocker 18beta-glycyrrhetinic acid did not affect expression of MAGI-1 and JAM-1 in Cx32 transfectants. These results suggest that Cx32 expression is in part related to induction of tight junctions through modulation of MAGI-1 expression in an immortalized mouse hepatic cell line.

DOI PubMed

Identification of a specific target sequence for p53 family in the Jagged2 gene

Yuichiro Oshima, Setsuko Ishida, Reo Maruyama, Minoru Toyota, Toshihiko Yamashita, Takashi Tokino, Yasushi Sasaki

Tumor Research 40 13 - 24 2005年 [査読有り]

Inactivation of the tumour suppressor p53 is the most common defect in cancer cells. The p53 family members, p73 and p63, have remarkable structural similarities with p53 and can activate transcription of p53-responsive genes. In contrast to p53 however, p73 and p63 are rarely mutated in human cancers. Mice that lack p53 are developmentally normal, while p73 and p63 appear to play critical roles in normal development. In our earlier study, we showed that both p73 and p63 regulate the ligands for the Notch receptor, Jagged1 (JAG1) and Jagged2 (JAG2). We also identified a p63-binding site in the second intron of the human JAG1 gene. Here, we have analyzed the responsive element within the human JAG2 gene for p73/p63-dependent induction of gene expression. Using a chromatin immunoprecipitation assay, we identified a p73/p63-binding site located in the second intron of the JAG2 gene. A heterologous reporter assay revealed that this binding site is a functional response element and is relatively specific for p73 and p63 among the p53 family. This binding site consists of four copies of 10-bp consensus p53-binding motif and is highly conserved between human and rodents. Furthermore, both JAG1 and JAG2 mRNA were specifically up-regulated by either p63 or p73, but not by p53 in mouse embryo fibroblasts. Our findings demonstrate that JAG2 is indeed a direct and evolutionarily conserved transcriptional target of p73 and p63, and suggest a potential molecular mechanism for the involvement of the p53 family genes in normal development.

SFRP遺伝子のepigeneticな不活化は大腸癌におけるWnt経路の異常活性化を補完する

鈴木拓, 豊田実, 時野隆至, 森満, 今井浩三

日本分子腫瘍マーカー研究会誌 ( Japanese Society for Molecular Tumor Marker Research ) 20 73 - 74 2005年

DOI CiNii

Dual role of p53 in DNA binding.

Takashi Tokino

Cancer biology & therapy 3 ( 12 ) 1322 - 3 2004年12月 [査読有り] [国際誌]

PubMed

Erratum: Inactivation of class II transactivator by DNA methylation and histone deacetylation associated with absence of HLA-DR induction by interferon-γ in haematopoietic tumour cells (British Journal of Cancer (2004) 90 (844-852) doi:10.1038/sj.bjc.6601602)

Y. Morimoto, M. Toyota, H. Ikeda, T. Tokino, K. Imai

British Journal of Cancer 91 ( 11 ) 1978 2004年11月 [査読有り]

DOI

Epigenetic inactivation of class II transactivator (CIITA) is associated with the absence of interferon-gamma-induced HLA-DR expression in colorectal and gastric cancer cells.

Ayumi Satoh, Minoru Toyota, Hideyuki Ikeda, Yoshikazu Morimoto, Kimishige Akino, Hiroaki Mita, Hiromu Suzuki, Yasushi Sasaki, Takayuki Kanaseki, Yukio Takamura, Hidenobu Soejima, Takeshi Urano, Kazuyoshi Yanagihara, Takao Endo, Yuji Hinoda, Masahiro Fujita, Masao Hosokawa, Noriyuki Sato, Takashi Tokino, Kohzoh Imai

Oncogene 23 ( 55 ) 8876 - 86 2004年11月 [査読有り] [国際誌]

Tightly regulated at the level of transcription, expression of MHC class II molecules varies significantly among gastrointestinal cancers. High levels of MHC class II expression are often associated with a better prognosis, which is indicative of the involvement of CD4+ lymphocytes in tumor suppression, but the molecular mechanism by which MHC class II expression is regulated remains unclear. In the present study, we investigated the expression of one inducible MHC class II molecule, HLA-DR, and its coactivators in a panel of colorectal and gastric cancer cell lines. Interferon-gamma induced expression of HLA-DR in 14 of 20 cell lines tested; the remaining six cell lines did not express HLA-DR. Analysis of the expression of transcription factors and coactivators associated with HLA-DR revealed that the loss of CIITA expression was closely associated with the absence of HLA-DR induction. Moreover, DNA methylation of the 5' CpG island of CIITA-PIV was detected in all cancer cells that lacked CIITA. The methylation and resultant silencing of CIITA-PIV depended on the activities of two DNA methyltransferases, DNMT1 and DNMT3B, and their genetic inactivation restored CIITA-PIV expression. It thus appears that CIITA methylation is a key mechanism that enables some gastrointestinal cancer cells to escape immune surveillance.

DOI PubMed

Identification of SCN3B as a novel p53-inducible proapoptotic gene.

Katsuya Adachi, Minoru Toyota, Yasushi Sasaki, Toshiharu Yamashita, Setsuko Ishida, Mutsumi Ohe-Toyota, Reo Maruyama, Yuji Hinoda, Tsuyoshi Saito, Kohzoh Imai, Ryuichi Kudo, Takashi Tokino

Oncogene 23 ( 47 ) 7791 - 8 2004年10月 [査読有り] [国際誌]

Tumor suppressor p53 is a transcription factor that induces growth arrest and/or apoptosis in response to cellular stress. To identify novel p53-inducible genes, we compared the expression of genes in normal mouse embryo fibroblasts (MEFs) to p53-null cells by cDNA representational difference analysis. We report here that expression of endogenous sodium channel subunit beta 3 (SCN3B) is upregulated in mouse embryonic fibroblasts by DNA damage in a p53-dependent manner. In addition, we found that SCN3B levels are upregulated in human cancer cell lines by DNA damaging agents, as well as by overexpression of p53, but not significantly by p63 or p73. Furthermore, we identified two putative p53-binding sites upstream of the first exon (RE1) and in the third intron (RE2). The p53 protein can directly interact with the putative p53-binding sites in vivo, as assessed by chromatin immunoprecipitation. A reporter gene assay revealed that these two p53-binding sites are functional response elements. The SCN3B protein appears to be localized to the endoplasmic reticulum (ER). Introduction of the SCN3B gene into T98G and Saos2 cells potently suppressed colony formation. Furthermore, we found that adenovirus-mediated transfer of SCN3B induced apoptosis when combined with anticancer agents. The results presented here suggest that SCN3B mediates a p53-dependent apoptotic pathway and may be a candidate for gene therapy combined with anticancer drugs.

DOI PubMed

T-fimbrin遺伝子のG2/M期チェックポイントへの関与と抗がん剤感受性

池田博, 佐々木泰史, 見田裕章, 豊田実, 時野隆至, 今井浩三

日本癌学会総会記事 ( (一社)日本癌学会 ) 63回 471 - 471 2004年09月

Reply: CIITA methylation and decreased levels of HLA-DR in tumour progression

Y Morimoto, M Toyota, H Ikeda, T Tokino, K Imai

BRITISH JOURNAL OF CANCER ( NATURE PUBLISHING GROUP ) 91 ( 4 ) 814 - 815 2004年08月

DOI

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