Kaulich, Manuel. Functional characterization of PICH, a DNA-dependent ATPase required for faithful chromosome segregation during mitosis. 2010, PhD Thesis, University of Basel, Faculty of Science.
Official URL: http://edoc.unibas.ch/diss/DissB_9395
In the first part of this thesis, we identify the ATPase activity of PICH as being essential for its correct subcellular localization and show that ATPase-dead mutants of PICH localize to the chromosome arms. Rescue experiments with a mutant of PICH that is unable to interact with Plk1 imply that the kinase activity of Plk1 is only indirectly required to delocalize PICH from the chromosome arms. This suggests that an unknown Plk1 substrate regulates the localization of PICH. PICH-immunoprecipitation and mass spectrometry identified the uncharacterized protein BEND3 as a binding partner. BEND3 localizes to mitotic chromosomes and its depletion results in the loss of PICH and Plk1 from the centromere/KTs. BEND3 depleted cells are unable to align their chromosomes at the metaphase plate and undergo apoptosis within a short time, suggesting that BEND3 regulates not only the localization of PICH and Plk1 but has additional functions.
In the second part of this thesis, we demonstrate that PICH does not function in the spindle assembly checkpoint. Instead, we show that neutralization or depletion of PICH results in chromatin bridges during anaphase. We identify the ATPase activity of PICH as indispensable to prevent the formation of these bridges and show that they most likely arise from non-centromeric chromatin. PICH was previously shown to interact with the BTR complex (composed of BLM, TOP3A and RMI1) that has been linked to the faithful separation of chromosomes during mitosis. It is of great interest, therefore, that we discovered an association of PICH with Rif1, which has recently been shown to provide a DNA-binding interface for the BTR complex. In contrast to the colocalization of PICH and the BTR complex to non-centromeric UFBs, Rif1 localizes to centromeric UFBs in a PICH-dependent manner. Thus, we speculate that PICH, Rif1 and the BTR complex act together to prevent chromatin bridges during mitosis.
|Advisors:||Nigg, Erich A.|
|Committee Members:||Affolter, Markus and Jiricny, Josef|
|Faculties and Departments:||05 Faculty of Science > Departement Biozentrum > Growth & Development > Cell Biology (Nigg)|
|Bibsysno:||Link to catalogue|
|Number of Pages:||109 Bl.|
|Last Modified:||30 Jun 2016 10:41|
|Deposited On:||11 Mar 2011 08:59|
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