Friday, November 22, 2024
11/22/2024
The goal of this proposal is to elucidate the molecular and cellular basis of a newly discovered diverse set of adaptive mechanisms that maintain the integrity of the genome during anaphase and telophase. Eukaryotic cells have evolved sophisticated mechanisms to maintain genomic integrity. In response to insults to the genome, checkpoints delay progression through the cell cycle, providing time for either repair of the damaged DNA or induction of apoptosis to eliminate the compromised cell. The majority of previous studies have focused on checkpoint mechanisms operating during interphase and metaphase. In spite of these safeguards, cells occasionally enter and progress through metaphase with damaged DNA, lagging chromatin, or a chromosome which has failed to secure a stable association with a kinetochore microtubule. Focusing on these errors led to the discovery of the abscission checkpoint, which delays the final stages of cytokinesis, providing time to clear chromatin from the path of the cytokinesis furrow. This discovery changed the prevailing view that, once the spindle-assembly checkpoint is satisfied, anaphase and telophase proceed rapidly without error correction mechanisms. Clearly, this is not the case and a number of labs, included ours, investigate the compensatory mechanisms that operate during anaphase to maintain genomic integrity. Here, we propose that in addition to the abscission checkpoint, the eukaryotic cell maintains a sophisticated and diverse set of mechanisms that function during the anaphase-telophase transition to ensure that chromosome fragments lacking a kinetochore (acentrics) are successfully transmitted and incorporated into daughter nuclei. This conclusion is based on our finding that in Drosophila, acentric chromosome fragments successfully congress, undergo delayed but proper sister separation, and efficiently segregate to opposing poles and incorporate into daughter nuclei. This was surprising as all of these chromosome dynamics were thought to be largely driven by kinetochore-microtubule associations. Our studies revealed a number of previously undescribed mechanisms and cellular adaptations such as DNA tethers, cell and spindle elongation, and channels in the nuclear envelope. We view these as revealing novel mechanisms operating during anaphase/telophase to maintain genomic integrity. In addition, observing the behavior of acentric chromosome fragments reveals forces acting on chromosomes that are independent of the kinetochore-microtubule attachments. We view these as key mechanisms that maintain genomic fidelity when cells exit metaphase with damaged chromosomes. Given that transmission of acentric chromosomes and DNA bridges have been observed across the phyla, many of these mechanisms are likely conserved. The studies described here rely on a combination of genetic, fluorescent and electron microscope studies as well as on high throughput cell-based screens to determine the molecular basis of these adaptive mechanisms functioning at the final stages of the eukaryotic cell cycle.
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