吳益群臺灣大學：分子與細胞生物學研究所李以如Lee, I-JuI-JuLee2007-11-252018-07-062007-11-252018-07-062007http://ntur.lib.ntu.edu.tw//handle/246246/49921In Eukaryotes, the cell cycle is under delicate control. Different phosphorylation state of a protein often leads to different activity, thus a number of protein kinases play important roles in mitosis. The C. elegans tousled-like kinase TLK-1 is a serine/threonine kinase that is important for DNA condensation and segregation. The tlk-1(RNAi) embryos arrested at approximately 55 cells stage and showed aneuploidy. To study the expression pattern of TLK-1, we generated TLK-1 antibodies. We show that TLK-1 is localized in the interphase nucleus but diffuses to the cytosol after nuclear envelope breakdown during mitosis. We found that TLK-1 is likely to act in chromosome segregation through DLC-1. DLC-1, a subunit of cytoplasmic dynein complex, interacts with TLK-1 in a yeast 2-hybrid system. By using time-course microscopic analysis we show that the depletion of dlc-1 causes defects in spindle assembly and chromosome segregation. Using immuno-staining we found that DLC-1 is enriched in mitotic centrosomes and spindle in wild-type but not tlk-1(RNAi) embryos. We conclude TLK-1 may mediate chromosome segregation by controlling the localization of DLC-1 to the mitotic centrosomes and spindles.Abstract...1 摘要...2 Introduction...3 Materials and Methods...8 Results...15 tlk-1(RNAi) embryos exhibit defects in DNA condensation and segregation...15 The depletion of tlk-1 affects kinetochore alignment...16 TLK-1 diffuses to cytosol when a cell processes from prophase to metaphase...17 The localization of TLK-1 is not regulated through APC-targeted ubiquitination...18 dlc-1(RNAi) causes embryonic lethality...19 dlc-1(RNAi) does not affect the localization of TLK-1...20 dlc-1(RNAi) embryos exhibit defects in mitotic spindle assembly and chromosome segregation...20 DLC-1 gathers around centrosomes during mitosis...22 The localization of DLC-1 is impaired in tlk-1(RNAi)...22 Discussion...24 The condensation defect in tlk-1(RNAi...24 TLK-1 and the holocentric proteins...25 The segregation defect in tlk-1(RNAi)...25 The regulation of TLK-1 during cell cycle...26 The embryonic lethality of dlc-1(RNAi) mutants in different strains...27 The interaction between TLK-1 and DLC-1...27 The functions of DLC-1 and dynein complex in mitosis...29 Future perspectives...31 Figures and Tables...32 Reference...50 Appendix...543471946 bytesapplication/pdfen-US線蟲有絲分裂細胞分裂C. elegansmitosisotherhttp://ntur.lib.ntu.edu.tw/bitstream/246246/49921/1/ntu-96-R94b43007-1.pdf