The “Big Picture”: For an overview of the cool chromosome biology that motivates our work, check out our Research Overview page.
The big question: How are the final steps in cell division coordinated with the completion of DNA replication and the “resolution” of sister chromatids?
Early mitotic events (i.e., prior to anaphase) ensure that sister chromatids are properly aligned so that each daughter cell receives the same number of chromatids. Anaphase events must ensure that the final regions of the chromosome have replicated completely and that sister chromatids are “untangled” from one another to avoid chromosome damage. Work from Ian Hickson’s lab (and others; see figure below) highlight the importance of anaphase for maintaining chromosome integrity. In studies of patients’ cells with defects in a DNA helicase, BLM (Bloom’s syndrome helicase), Hickson and colleagues demonstrated that these cells are prone to chromosome damage and Bloom’s Syndrome patients exhibit a high incidence of cancer. The BLM helicase is critical for “untangling” sister chromatids in anaphase. Tangled sister chromatids are a natural consequence of the topological linkages formed in DNA during the replication process. These linkages can be made worse when cells experience “replication stress”. Unresolved sister chromatids (aka, lagging chromosomes) form anaphase “bridges” that if left unresolved can form micronuclei. Micronuclei are associated with high rates of genome rearrangement and with the most malignant forms of cancer.
Anaphase surveillance: We propose that there are a series of regulatory pathways that link chromosome replication to anaphase in order to ensure sister chromatid resolution is coordinated with cell division to suppress chromosome damage and micronuclei formation.
Anaphase regulators: Our interest in anaphase grew out of our experiments on kinetochore assembly. We noticed that a surprising number of kinetochore proteins also associated with the anaphase spindle, similar to those proteins in the Aurora B kinase complex whose behavior in transitioning from kinetochores to anaphase spindles earned them the name, “chromosome passenger complex” (CPC). To date, CPC function in anaphase has been linked to sister chromatid resolution, spindle dynamics (elongation in anaphase B and breakdown) as well as changes in membrane dynamics involved in de novo formation plasma membranes and nuclear envelopes at the end of mitosis. These functions clearly implicate the CPC in anaphase surveillance.
CPC septin regulation: Septin filaments assemble with cellular membranes to create a diffusion barrier for proteins that must be enriched in specific membrane regions. This function can limit the diffusion of membrane proteins thus creating concentrated membrane-protein domains (e.g., primary cilium and the cytokinetic furrow). We showed that CPCs can modulate the dynamics of septin filaments during cell division. These findings are summarized in the figure below. The fluorescence image shows how septin rings form in anaphase, the cartoon shows the how septins disassemble in the mother cell after mitotic exit and the kymographs show the distinct effects of different septin mutants on disassembly. Understanding how CPCs regulate septins remains on ongoing line of inquiry in the lab, but our findings connect Aurora B and CPCs to controlling membrane protein dynamics during cell division.
Tangled sister chromatids and anaphase spindle regulation: The Aurora B kinase complex, or CPCs, have also been implicated in regulating microtubule-associated proteins. In animal cells CPCs are linked to formation of the spindle midzone, while in yeast it has been suggested to help breakdown the anaphase spindle at the end of cell division. In unpublished work, we have found replication stress, or mutants that contribute to tangled sister chromatids, alter the substrate targeting of the Aurora B kinase. As a result, there is an increase in inter-polar spindle microtubules (see Figure below) that contributes to an increase in anaphase spindle elongation rate required for damage free resolution of sister chromatids. Thus, Aurora B and the CPC form a feedback loop that coordinates DNA replication with spindle dynamics and sister chromatid resolution.
Future directions and an anaphase surveillance model: In summary, our findings are consistent with a series of pathways contributing to anaphase surveillance that together ensure sister chromatid resolution is coordinated with the final steps of cell division. We are currently focused on pathways that link sister chromatid resolution to the regulation of membrane dynamics during cell division. This line of work includes understanding how septins contribute to autophagy membrane dynamics both during starvation and in anaphase, as well as addressing how regulation of autophagy is involved in preserving chromosome integrity after replication stress.