The study on function of PAX3-FOXO1 in cell cycle

• Main Authors: Chun-Yen Wang, 王竣彥
• Other Authors: Wen-Ming Yang
• Format: Others
• Language: zh-TW
• Published: 2017
• Online Access: http://ndltd.ncl.edu.tw/handle/70561901933415091716

碩士 === 國立中興大學 === 分子生物學研究所 === 105 === Fusion protein PAX3-FOXO1 involves in the pathogenesis of alveolar rhabdomyosarcoma, a pediatric skeletal muscle cancer that occur in children and teenager. The PAX3-FOXO1, that contains the DNA binding domain of PAX3 and transactivation domain of FOXO1, is caused by the gene translocation of chromosome 2 and chromosome 13. Previous studies showed that PAX3 localizes on heterochromatin region and chromosome during S phase and M phase through its’ DNA binding domain. In this thesis, It is found that the PAX3-FOXO1 co-localized with PAX3 on heterochromatin region and chromosome, which implied a novel function of PAX3-FOXO1 in S phase and M phase. In addition, previous studies showed that PAX3-FOXO1 induces cell proliferation through down-regulation of cyclin-dependent kinase inhibitor. However, the function of PAX3-FOXO1 in cell cycle remains unclear. Therefore, this thesis studied the function of PAX3-FOXO1 in cell cycle. In order to study the function of PAX3-FOXO1 in cell cycle, cell cycle synchronization and flow cytometry were used to analyze whether PAX3-FOXO1 impair progression of G1, S, and G2/M phase. The results showed that PAX3-FOXO1 prolonged G2/M phase progression but not G1 and S phase, suggesting PAX3-FOXO1 was a gain of function mutant, which prolonged G2/M phase progression. To further confirm this phenomenon, flow cytometry was further used to analyze the cell cycle progression of alveolar rhabdomyosarcoma cell line, that contain PAX3-FOXO1, and embryonal rhabdomyosarcoma cell line, that do not contain PAX3-FOXO1. It was found that alveolar rhabdomyosarcoma cell line got longer cell cycle, supporting PAX3-FOXO1 prolonged cell cycle progression. According to these findings, It was hypothesized that PAX3-FOXO1 causes the genome instability through mitotic defect. The genome instability contains the phenomena of chromatin bridge, multipolar, and micronuclei. In this thesis, It was found that PAX3-FOXO1 causes the phenomena of mitotic bridge and multipolar but not micronuclei or cell death. Taken together, the results showed that PAX3-FOXO1 co-localized with PAX3 on mitotic chromosome and delayed the progression of G2/M phase through causing genome instability during M phase but not cell death.