생명과학회지 (Journal of Life Science)

  • 제15권5호
  • /
  • Pages.802-808
  • /
  • 2005
  • /
  • 1225-9918(pISSN)
  • /
  • 2287-3406(eISSN)
한국생명과학회 (Korean Society of Life Science)
권호 표지
DOI QR코드

DOI QR Code

사람의 유방암 세포주인 MDA MB-231 세포에서 CpG 메칠화에 의한 Disabled-2유전자의 발현억제

Silencing of Disabled-2 Gene by CpG Methylation in Human Breast Cancer Cell Line, MDA MB-231 Cells

  • 고명현 (원광대학교 자연대학 생명과학부) ;
  • 오유미 (원광대학교 자연대학 생명과학부) ;
  • 박준호 (원광대학교 자연대학 생명과학부) ;
  • 전병훈 (원광대학교 한의과대학 병리학교실) ;
  • 한동민 (원광대학교 자연대학 생명과학부) ;
  • 김원신 (원광대학교 자연대학 생명과학부)
  • Ko Myung Hyun (Division of Biological Science, Wonkwang University) ;
  • Oh Yu Mi (Division of Biological Science, Wonkwang University) ;
  • Park Jun Ho (Division of Biological Science, Wonkwang University) ;
  • Jeon Byung Hoon (College of Oriental Medicine, Wonkwang University) ;
  • Han Dong Min (Division of Biological Science, Wonkwang University) ;
  • Kim Won Sin (Division of Biological Science, Wonkwang University)
  • 발행 : 2005.10.01
PDF 다운로드
⟨ 이전 논문 다음 논문 ⟩

초록

사람의 Disabled-2 (Dab2)는 정상세포에서 c-Fos의 발현을 억제하여 세포 성장을 조절하는 암억제 유전자로 추정되어 지고 있다. 많은 암세포에서 Dab2는 유방과 난소의 정상세포에서는 발현이 되지만, 약 $85\%$의 유방과 난소의 종양세포에서는 발현이 줄어들거나, 발현이 억제되는 것으로 알려져 있다. 본 연구에서는 bisulfite 반응에 의한 염기서열 분석법과 MSP방법 등을 이용하여 유방암 세포주인 MDA MB-231세포에서 Dab2 유전자의 promoter상에 존재하는 Cpg island의 methylation된 상태를 분석하였다. 그 결과, 사람의 정상 자궁내막세포에서는 Dab2 promoter 부위가 완전하게 methylation되어 있지 않았다. 그러나 MDA MB-231세포에서는 TATA box 근처 의 CpG dinucleotide에서 비정상적으로 methylation되어 있었다. 이런 비정상적인 CpG dinucleotide의 methylation은 MDA MB-231세포를 5-azacytidine으로 처리하였을 때 methylation이 풀리고, Dab2의 발현이 회복되는 것으로 나타났다. 따라서 인간 유방암 세포주인 MBA MB-231세포에서 Dab2의 발현억제는 Dab2 유전자의 promoter부위의 CpG island의 비정상적인 methylation과 관련이 있는 것으로 여겨진다.

Human Disabled-2 (Dab2) is a candidate tumor suppressor gone that regulates cell growth by c-Fos suppression in normal cells. In many cancer cells, Dab2 expression is lost or greatly diminished in $\∼85\%$ of the breast and ovarian cancers. In this study, we have examined the methylation status of CpG island on Dab2 gene promoter using bisulfite-assisted genomic sequencing and methylation specific PCR (MSP) method in human breast cancer cell line, MDA MB-231 cells. In normal human uterus endometrial cells, Dab2 was completely unmethylated. In contrast, Dab2 was methylated on CpG dinucleotides near the TATA_ box in MDA MB-231 cells. following MDA MB-231 cells by treatment with 5-azacytidine, Dab2 gene were demethylated and reexpressed. Result of this study suggested that silencing of Dab2 gene is correlated to CpG island methylation in human breast cancer cell line, MBA MD-231 cells.

키워드

참고문헌

  1. Antequera, F., J. Boyes and A. Bird. 1990. High levels of de novo methylation and altered chromatin structure at epG islands in cell lines. Cell 62, 503-514 https://doi.org/10.1016/0092-8674(90)90015-7
  2. Beard, C., E. Li and R. Jaenisch. 1995. Loss of methylation activates Xist in somatic but not in embryonic cells. Genes Dev. 9, 2325-2334 https://doi.org/10.1101/gad.9.19.2325
  3. Becker, P. B., S. Ruppert and G. Schutz. 1987. Genomic footprinting. reveals cell type-specific DNA binding of ubiquitous factors. Cell 51, 435-443 https://doi.org/10.1016/0092-8674(87)90639-8
  4. Berger, J. and G. Daxenbichler. 2002. DNA methylation of nuclear receptor genes-possible role in malignancy. J Steroid Biochem. Mol. Biol. 80, 1-11 https://doi.org/10.1016/S0960-0760(01)00179-0
  5. Bird, A. 1992. The essentials of DNA methylation. Cell 70, 5-8 https://doi.org/10.1016/0092-8674(92)90526-I
  6. Clemson, C. M., J. A. McNeil, H. F. Willard and J. B. Lawrence. 1996. XIST RNA paints the inactive X chromosome at interphase: evidence for a novel RNA involved in nuclear/chromosome structure. J. Cell Biol. 132, 259-275 https://doi.org/10.1083/jcb.132.3.259
  7. Cooper, D. N. and M. Krawczak. 1990. The mutational spectrum of single base-pair substitutions causing human genetic disease: patterns and predictions. Hum. Genet. 85, 55-74
  8. Fazili, Z., W. Sun, S. Mittelstaedt, C. Cohen and X. X. Xu. 1999. Disabled-2 inactivation is an early step in ovarian tumorigenicity. Oncogene 18, 3104-3113 https://doi.org/10.1038/sj.onc.1202649
  9. Fulop, V., S. C. Mok, D. R. Genest, I. Gati, J. Doszpod and R. S. Berkowitz. 1998. p53, p21, Rb and mdm2 oncoproteins expression in normal placenta, partial and complete mole, and choriocarcinoma. J. Reprod. Med. 43, 119-127
  10. Gebara, M. M. and J. R. McCarrey. 1992. Protein-DNA interactions associated with the onset of testis-specific expression of the mammalian Pgk-2 gene. Mol. Cell. Biol. 12, 1422-1431 https://doi.org/10.1128/MCB.12.4.1422
  11. Grigg, G. and S. Clark, 1994. Sequencing 5-methylcytosine residues in genomic DNA. Bioessays 16, 431-436 https://doi.org/10.1002/bies.950160612
  12. Gertler, F. B., K. K. Hill, M. J. Clark and F. M. Hoffmann. 1993. Dosage-sensitive modifiers of Drosophila abl tyrosine kinase function: prospero, a regulator of axonal outgrowth, and disabled, a novel tyrosine kinase substrate. Genes Dev. 7, 441-453 https://doi.org/10.1101/gad.7.3.441
  13. Guo, S. X., T. Taki, H. Ohnishi, H. Y. Piao, K. Tabuchi, F. Bessho, R. Hanada, M. Yanagisawa and Y. Hayashi. 2000. Hypermethylation of p16 and pIS genes and RB protein expression in acute leukemia. Leuk. Res. 24, 39-46 https://doi.org/10.1016/S0145-2126(99)00158-7
  14. Hajkova, P., O. el-Maarri, S. Engemann, J. Oswald, A. Olek and J. Walter. 2002. DNA-methylation analysis by the bisulfite-assisted genomic sequencing method. Methods Mol. Biol. 200, 143-154
  15. He, J., E. R. Smith and X. X. Xu. 2001. Disabled-2 exerts its tumor suppressor activity by uncoupling c-Fos expression and MAP kinase activation. J. Biol. Chem. 276, 26814-26818 https://doi.org/10.1074/jbc.M101820200
  16. Herman, J. G. 1999. Hypermethylation of tumor suppressor genes in cancer. Semin Cancer Biol. 9, 359-367. Review
  17. lssa, J. P., S. B. Baylin and S. A. Belinsky. 1996. Methylation of the estrogen receptor CpG island in lung tumors is related to the specific type of carcinogen exposure. Cancer Res. 56, 3655-3658
  18. Jones, PA and P. W. Laird. 1999. Cancer epigenetics comes of age. Nat. Genet. 21, 163-167 https://doi.org/10.1038/5947
  19. Li, E., C. Beard and R. Jaenisch. 1993. Role for DNA methylation in genomic imprinting. Nature 366, 362-365 https://doi.org/10.1038/366362a0
  20. Lichtenstein, M, G. Keini, H. Cedar and Y. Bergman. 1994. B cell-specific demethylation: a novel role for the intronic chain enhancer sequences. Cell 76, 913-923 https://doi.org/10.1016/0092-8674(94)90365-4
  21. Maass, N., M Biallek, F. Rosel, C. Schem, N. Ohike, M. Zhang, W. Jonat and K. Nagasaki. 2002. Hypermethylation and histone deacetylation lead to silencing of the mas pin gene in human breast cancer. Biochem Biophys Res. Commun. 297, 125-128 https://doi.org/10.1016/S0006-291X(02)02136-8
  22. Mok, S. C., W. Y. Chan, K. K. Wong, K. K. Cheung, C. C. Lau, S. W. Ng, A. Baldini, C. V. Colitti, C. O. Rock and R. S. Berkowitz. 1998. DOC-2, a candidate tumor suppressor gene in human epithelial ovarian cancer. Oncogene 16, 2381-2387 https://doi.org/10.1038/sj.onc.1201769
  23. Razin, A. and H. Cedar. 1991. DNA methylation and gene expression. Microbiol. Rev. 55, 451-458
  24. Richardson, B. and R. Yung. 1999. Role of DNA methylation in the regulation of cell function. J. Lab Clin. Med. 134, 333-340 https://doi.org/10.1016/S0022-2143(99)90147-6
  25. Tseng, C. P., C. H. Huang, C. C. Tseng, M. H. Un, J. T. Hsieh and C. H. Tseng. 2001. Induction of disabIed-2 gene during megakaryocyte differentiation of k562 cells. Biochem. Biophys. Res. Commun. 285, 129-135 https://doi.org/10.1006/bbrc.2001.5133
  26. Xu, X. X., T. Yi, B. Tang and J. D. Lambeth. 1998. Disabled2 (Dab2) is an SH3 domain-binding partner of Grb2. Oncogene 16, 1561-1569 https://doi.org/10.1038/sj.onc.1201678
  27. Yeivin, A. and A. Razin. 1993. Gene methylation patterns and expression, p. 523-568. In J. P. Host and H. P. Saluz (ed.), DNA methylation: molecular biology and biological significance. Birkhauser Verlag, Basel, Switzerland
  28. Young, H. A. 1996. Regulation of interferon-gamma gene expression. J. Interferon Cytokine Res. 16, 563-568 https://doi.org/10.1089/jir.1996.16.563