| Budget Amount *help |
¥2,600,000 (Direct Cost: ¥2,600,000)
Fiscal Year 2006: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 2005: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Research Abstract |
The aim of this study is to identify centrosome re-duplication suppressor gene(s) by chromosome-transfer technology. We carried out the following experiments.
(1) At first, we analyzed centrosome status of mouse A9 cells. Mouse A9 cells are normal origin but makes tumors in nude mice. Thus, A9 cells are precancerous cell line. We examined centrosome staus in A9 cells by immunostaining, and found that centrosome number is normal. We next expose hydroxyurea (HU), a DNA synthesis inhibitor, to A9 cells. The exposed cells were arrested at G1-S transition in cell cycle and were inhibited DNA synthesis. However, centrosome duplication cycle was dysregulated in A9 cells; multi-round of centrosome duplication occurred under HU treatment. Thus, A9 cells have a potent centrosome re-duplication activity.
(2) Next, we tried to examine centrosome re-duplication activity in A9 cells transferred with a human chromosome 1, 5, 8, 10, 11, 13, 15, 16, 17, and 18, respectively. As a result, we found that centrosome re-duplication activity was suppressed in A9-Ch. 8, A9-Ch. 15, and A9-Ch. 17 cells, respectively. These results suggest that the gene(s), which is related in suppression of centrosome re-duplication, is located on chromosome 8, chromosome 15, and chromosome 17. Especially, A9-Ch. 8 cells have high centrosome re-duplication suppressor activity. Therefore, we determined to isolate centrosome re-duplication suppressor gene(s) from chromosome 8.
(3) As it is found that human osteosarsoma cell line, U2OS has centrosome re-duplication activity, we transferred chromosome 8 into U2OS cells and examined centrosome re-duplication activity. As a result, we found that chromosome 8 had a centrosome re-duplication suppressor activity. Now, we tried to isolate the candidate gene by radiation hybrid mapping.
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