UC Santa Cruz
Worst case scenario: exiting metaphase with a broken chromosome
- Author(s): Karg, Travis
- Advisor(s): Sullivan, William T
- et al.
Unrepaired double strand DNA breaks can have deleterious consequences at mitosis due to the generation of acentric chromosome fragments that lack centromeres. Although acentrics are common in cancer cells, surprisingly little is known about the mechanisms regulating kinetochore-independent chromosome segregation. Acentrics are efficiently induced in Drosophila neuroblasts by expressing an endonuclease called I-CreI. Previous reports in Drosophila have found that while acentrics lack a kinetochore, they still move to the metaphase plate and segregate to daughter cells. The delayed but ultimately successful segregation of acentrics requires a chromatin tether that connects the acentrics to the main chromosome mass. The tethers are decorated with BubR1, Aurora B and Polo kinases. Disruptions in BubR1 and Polo functions led to defects in acentric segregation and a decreased rate of survival from larvae into adulthood (synthetic lethality). This led to the conclusion that the tether provides the driving force for acentric segregation. Here, I demonstrate that acentrics are preferentially bundled with microtubules during anaphase suggesting that microtubules and motor proteins exert the force for driving acentric segregation. Interestingly, the orientation of acentrics during anaphase is random with half the acentrics segregating telomere first (telomere facing pole), while the other half segregates telomere last (telomere facing metaphase plate). This is in accord with a model in which acentrics behave as cargo capable of associating in any orientation with microtubules. By conducting a synthetic lethality screen, I found that I-CreI expressing larvae with disruptions in Klp3a, which encodes for a chromosome and microtubule-associated motor protein (chromokinesin) led to an increased rate of synthetic lethality. Through live imaging, I found defects in acentric segregation in neuroblast depleted of Klp3a. Taken together, these results suggest that the cell maintains a microtubule and Kp3a dependent mechanism to drive acentric segregation.
Delayed acentric segregation during anaphase raises the question of how acentrics affect nuclear envelope reformation (NER). Here I demonstrate that acentrics enter daughter nuclei by inducing a local delay in NER, which leads to the formation of a channel in the nuclear envelope. The formation of the nuclear envelope channels is dependent on Aurora B kinase.