TopBP1 contains repeats of the BRCA1 C-terminal (BRCT) domain and plays important roles in DNA damage response, DNA replication, and other cellular regulatory functions during the interphase. In prometaphase, metaphase, and anaphase, TopBP1 localizes to the mitotic centrosomes, which function as spindle-poles for the bipolar separation of sister chromatids. The localization of TopBP1 to the mitotic centrosomes is mediated by amino acid residues 1259 to 1420 in the TopBP1 C-terminal region (TbpCtr). GST and DsRed2 tags fused to TbpCtr were localized in the mitotic centrosomes, thereby suggesting that TbpCtr functions as a mitosis-specific centrosome localization signal (CLS). Mutations of Ser 1273 and/or Lys 1317, which were predicted to interact with a putative phosphoprotein, inhibited CLS function. Ectopic expression of TbpCtr specifically eliminated endogenous TopBP1 from the mitotic centrosomes, whereas mutant TbpCtr derivatives, containing substitutions at Ser 1273 and/or Lys 1317, did not. The specific elimination of TopBP1 from the mitotic centrosomes prolonged the durations of prometaphase and metaphase and shortened the inter-kinetochore distances of metaphase sister chromatids while maintaining the spindle assembly checkpoint. These results suggest that the localization of TopBP1 to the mitotic centrosomes is necessary for proper mitotic progression.
Bibliographical noteFunding Information:
We thank Dr. Yunje Cho for the 7th BRCT domain structure prediction, Dr. Chang-Woo Lee for providing BubR1 antibody, Dr. Hyunsook Lee for providing CREST antibody, Dr. Kunsoo Rhee for providing the HeLa GFP-Centrin cell line, and Jeena Jiyeon Yoo for technical assistance. This work was supported by grants from the Korea Research Foundation ( KRF-2008-313-C00672 and 2009-0053226 ) and the Nuclear R&D Program of the National Research Foundation of Korea ( 2009-0078699 ).
All Science Journal Classification (ASJC) codes
- Cell Biology