Lee, Chang-Woo, PhD, Associate Professor Lab. of Tumor Cell Biology Department of Molecular Cell Biology Sungkyunkwan University School of Medicine 300 Chunchun-dong, Jangan-gu Suwon 440-746 Korea Tel: +82-31-299-6121 e-mail: cwlee@med.skku.ac.kr

Background:

Research:

Molecular mechanism of mitotic checkpoint activation: BubR1 mitotic checkpoint protein plays an essential role in the maintenance of genomic integrity, and thus, defects in the BubR1-mediated signaling not only eliminate checkpoint control, but are also linked to human cancers. Despite the importance of BubR1 mitotic checkpoint kinase in genomic integrity, very little is known in regards to what the target substrates of BubR1 kinase are, how BubR1 is phosphorylated during mitosis, and how BubR1 is involved in the prevention of the adaptation. Thus, a critical task in understanding the molecular basis of the BubR1-mediated signaling which is responsible for the mitotic checkpoint may be defining the genes required for establishing this checkpoint. In our approach to answering these questions, we have adopted the complex-proteomic and yeast two-hybrid assays, and screened two candidates as novel BubR1-interacting proteins.

Screening of the upstream kinase and downstream substrate of oncogenic mitotic kinases, Aurora-A, Aurora-B & Polo-like kinase 1: Aurora kinases and polo-like kinase 1 are aberrantly and highly overexpressed in the variety of human cancer cells. The deregulation of these mitotic kinases cause severe mitotic defects and anueuploidy and/or genetic instability, leading to cell death or disease. These kinases could be useful therapeutic targets against human cancers. To investigate the molecular signaling pathway of these mitotic kinases in tumorigenesis, we have performed the immuno-complex proteomics and MALDI-TOF analyses. We have screening upstream regulatory kinases and downstream substrate proteins of these mitotic kinases.

Molecular mechanism of the formation and resolution of cohesin complex: In vertebrates,sister chromatid cohesion depends on chromosomal protein complexes, consisting of four subunits (SMC1, SMC3, SCC3 and SCC1/Rad21). Cohesin is removed from chromosome arms in prophase and prometaphase by a mechanism that depends on phosphorylation of the cohesin complexes involved in SA2 and SCC1/Rad21 by mitotic kinases Plk1 and Aurora B. In addition to cohesin at centromeres is removed by the activated separase via SCC1/Rad21 cleavage-pathway and then initiates sister chromatid separation. We are interested in understanding how cohesin is dissociated at chromosome arms during prophase and prometaphase, how phosphorylation of cohesion complexes results in the removing of cohesin from chromosomes in mitosis, and how separase activity is regulated. Our research will contribute to a better understanding of human cancer because chromosome abnormality, such as aneuploidy and translocations, is tightly associated with tumor development and birth defects.

Screening of novel-Separase binding proteins: During metaphase, cohesin is required for holding sister chromatids together. Anaphase is triggered when separase cleaves the Rad21 subunit of cohesion. Destroying the cohesin complex by separase, the spindle pull sister chromatids into opposite poles of the cell. Because of the irreversible nature of Rad21 cleavage, separase is tightly regulated by phosphorylation and inhibitor protein-PTTG, auto-cleavage. In yeast, other functions of separase were reported, including mitotic exit and cleavage substrate. But, the precise regulation and the other function of separase in mammalian remain to be solved. We have been identified separase binding candidates using yeast two hybrid system and tried to find the function of the regulatory protein.

Molecular mechanism of mitotic chromatin remodeling: Eukaryotic cells must possess mechanisms for condensing and decondensing chromatin. Moreover, chromatin condensation is particularly evident during mitosis and apoptotic cell death, whereas chromatin relaxation is necessary for replication, repair, recombination and transcription. The post-translational modifications of histone tails such as reversible acetylation, phosphorylation and methylation play a critical role in dynamic condensation/relaxation that occurs during the cell cycle. Nonetheless, the potential role of histone acetylation, methylation, phosphorylation in maintaining the active configuration during the mitotic stage of the cell cycle is not completely understood. Therefore, we believe that this study may provides cooperative roles for multiple histone modifications, acetylation, methylation and phosphorylation etc, in regulating mitotic chromatin remodeling.

The cell cycle and division of adult stem cells: The goals of this proposal are to establish the tissue culture condition, to understand the molecular mechanisms of cell cycle control, self-renewal, asymetric cell division and differentiation lineage, and to screen the differentiation surface marker proteins of adipose-derived stem cells (ASCs). Therefore, this study may provides substantial evidences to know the molecular mechanisms of proliferation, division and differentiation of adult stem cells, and may be the basis for future clinical applications.

Publications (selected):

1. Ha GH, Baek KH, Cheong SJ, Kim HS, Kim CM, McKeon F, and Lee CW . 2007. p53 activation in response to mitotic spindle damage requires signaling via BubR1-mediated phosphorylation. Cancer Research (In press) .
2. Baek KH, Kang CM, Park HY, Kim SJ, Jeong SJ, Suzuki T, Kim CM, and Lee CW . 2006. Overexpression of hepatitis C virus NS5A protein induces chromosome instability via mitotic cell cycle dysregulation. Journal of Molecular Biology 359: 22-34.
3. Shin HJ, Park HY, Jeong SJ, Kim YG, Cho SH, Kim YY, Cho ML, Kim HY, Min KU, and Lee CW . 2005. STAT4 expression in human T cells is regulated by DNA methylation but not with promoter polymorphisms.Journal of Immunology 175: 7143-7150.
4. Jeong SJ, Shin HJ, Kim SJ, Ha GH, Baek KH, Kim CM, and Lee CW . 2004. Transcriptional abnormality of hsMAD2 mitotic checkpoint gene is a potential link to hepatocellular carcinogenesis. Cancer Research 64:8666-8673.
5. Shin HJ & Lee CW . et al. 2004. Dual roles of human BubR1, a mitotic checkpoint kinase, in the monitoring of chromosomal instability. Nature Reviews Cancer, 4:91.
6. Shin HJ, Baek KH, Jeon AH, Chung DH, Lee SJ, Lee HS, Sung YC, McKeon F, and Lee CW . 2003. Dual roles of human BubR1, a mitotic checkpoint kinase, in the monitoring of chromosomal instability. Cancer Cell (Cell press), 4:483-497.
7. Shin HJ, Baek KH, Jeon AH, Kim SJ, Jang KL, Sung YC, and Lee CW . 2003. Inhibition of histone deacetylase activity increases the chromosomal instability by the aberrant regulation of mitotic checkpoint activation. Oncogene 22:3853-3858.
8. Lee CW*, Shikama N*, France S, Delavaine L, Lyon J, Krstic-Demonacos M, and La Thangue NB. 1999. A new co-factor for p300/CBP that regulates the p53 response. Molecular Cell (Cell press) 4:365-376.