The Role of the Kinetochore in Chromosome Signaling and Segregation
Error-free chromosome segregation is crucial for the maintenance of genomic stability and prevention of tumorigenesis. The spindle assembly checkpoint (SAC) prevents exit from mitosis as long as not all chromosome pairs are properly attached to the mitotic spindle. Microtubules attach to chromosomes...
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| Format: | Dissertation |
| Language: | English |
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ProQuest Dissertations & Theses
01-01-2010
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| Online Access: | Get full text |
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| Summary: | Error-free chromosome segregation is crucial for the maintenance of genomic stability and prevention of tumorigenesis. The spindle assembly checkpoint (SAC) prevents exit from mitosis as long as not all chromosome pairs are properly attached to the mitotic spindle. Microtubules attach to chromosomes through binding at the kinetochore, a multi-protein structure that is organized over centromeric chromatin. The kinetochore plays an important role in the generation of the SAC signal that prevent mitotic exit while at the same time it is involved in chromosome movements during congression and segregation. The work reported in this thesis aims to further our understanding on how the combination of such functions ensures faithful chromosome segregation. The first part focuses on the phenotypic analysis of Drosophila melanogaster S2 cells depleted of the SAC protein BubR1 and the microtubule motor protein CENP-meta/CENP-E. The results show that while BubR1-depleted cells exit mitosis prematurely due to loss of SAC activity, CENPmeta-depleted cells are delayed in mitosis. Furthermore, a detailed analysis of microtubule-kinetochore interactions after depletion of BubR1 or CENP-meta clearly shows that without BubR1 cells contain a high frequency of monoriented and fully unattached chromosomes while most CENP-meta-depleted cells have chromosomes attached to spindle microtubules. Moreover, when both proteins are simultaneously depleted absence of CENP-meta is able to partly rescue microtubule-kinetochore attachments lost due to BubR1 depletion. The data shows that BubR1 is required to promote stable microtubule-kinetochore attachment. However, CENP-E appears to promote kinetochore-microtubule instability, suggesting a role of CENP-E in the mechanism that corrects inappropriate kinetochore attachment. Accordingly, the second part of the thesis concentrates in determining whether CENP-E and Aurora B kinase activities cooperate to promote efficient chromosome biorientation. The results, obtained in HeLa cells, do not support a regulation of Aurora B kinase activity by CENP-E. However, strongly suggest that Aurora B kinase destabilizes spindle pole proximal kinetochore-microtubule interactions keeping the SAC active to allow CENP-E-mediated congression of monoriented chromosomes. Taken together, the work presented in this thesis, through the study of two bona fidekinetochore proteins, provides insight in how chromosomes congress and how can they delay mitotic exit when errors occur thus avoiding aneuploidy. |
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| ISBN: | 9798382380735 |