Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
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Impaired Spindle Checkpoint Response of Brca1-deficient Mouse Embryonic Fibroblasts (MEFs) to Nocodazole Treatment

Brca1 결손 세포주에서 nocodazole 처리에 의한 spindle checkpoint 활성화 연구

  • Kim Myoung-Ae (Medical Research Center for Cancer Molecular Therapy, College of Medicine, Dong-A University, Department of Biochemistry, College of Medicine, Dong-A University) ;
  • Kim Hyunju (Medical Research Center for Cancer Molecular Therapy, College of Medicine, Dong-A University, Department of Biochemistry, College of Medicine, Dong-A University) ;
  • Yun Jeanho (Medical Research Center for Cancer Molecular Therapy, College of Medicine, Dong-A University, Department of Biochemistry, College of Medicine, Dong-A University)
  • 김명애 (동아대학교 의과대학 생화학교실, 동아대학교 암분자치료연구센터) ;
  • 김현주 (동아대학교 의과대학 생화학교실, 동아대학교 암분자치료연구센터) ;
  • 윤진호 (동아대학교 의과대학 생화학교실, 동아대학교 암분자치료연구센터)
  • Published : 2006.02.01
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Abstract

Genetic alternation of Brca1 predispose of breast and ovarian cancer. Brca1 plays critical role in cell cycle regulation following DNA damage. Previous studies revealed that Brca1 plays an important role in S phase and G2/M checkpoint regulation. However, whether Brca1 involves in spindle checkpoint is unclear. In this study, the role of Brca1 in cell cycle response following nocodazole, which is a reagent that depolymerizes microtubules and activates the spindle checkpoint, has been examined using wild type $p53^{-/-}\;and\;p53^{-/-}Brca1^{-/-}$ mouse embryonic fibroblasts (MEFs). While wild type and Brca1-proficient MEFs showed an acute mitotic arrest, Brca1-deficient MEFs failed to arrest at mitotic phase in response to nocodazole treatment. In double-thymidine block and nocodazole treatment experiment, a portion of $p53^{-/-}\;Brca1^{-/-}$ MEFs were clearly by-passed nocodazole induced mitotic arrest. Consistent with this, in morphologic analysis, $p53^{-/-}\;Brca1^{-/-}$ MEFs showed growing cell morphology after nocodazole treatment. Taken together, these results suggest that Brca1 protein is an important component for normal induction of spindlecheckpoint and impairment of Brca1 function could induce dysregulation of mitotic cell cycle that ultimately results in genomic instability.

항암유전자 Brca1의 변이는 유방암 및 난소암에 대한 감수성을 증가시키며, Brca1은 DNA손상신호후 세포주기 조절에 필수적인 역할을 한다. 연구결과, Brca1이 세포주기 S기와 G2/M 조절점에서 중요한 역할을 담당함이 밝혀졌다. 그러나, Brca1의 spindle checkpoint 관여여부는 알려져 있지 않다. 본 연구에서는 spindle checkpoint를 활성화시키는 nocodazole를 처리하여 야생형, $p53^{-/-}$ 그리고 $p53^{-/-}\;Brca1^{-/-}$ 세포주의 세포주기 변화를 조사하였다. 야생형과 $p53^{-/-}$ 세포주는 신속한 mitosis기 정지가 나타난 반면, $p53^{-/-}\;Brca1^{-/-}$ 세포주의 경우 모든 세포가 M기에서 정지하지 않았다. Double-thymidine block 기법에 의한 세포주기 동조화후 nocodazole 처리시에도 $p53^{-/-}\;Brca1^{-/-}$ 세포주에서는 일부세포가 M기 조절점을 통과하여 계속 G1기로 진행하였다. 형태학적 분석에서도 nocodazole 함유배지에서 계속 증식하는 세포형태가 관찰되었다. 이와 같은 결과들은 Brca1이 spindle checkpoint가 정상적으로 작동하는데 중요한 역할을 담당한다는 것을 의미하고 있다.

Keywords

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