Cell cycle responses to Topoisomerase II inhibition: Molecular mechanisms and clinical implications.

DNA Topoisomerase IIA (Topo IIA) is an enzyme that alters the topological state of DNA and is essential for the separation of replicated sister chromatids and the integrity of cell division. Topo IIA dysfunction activates cell cycle checkpoints, resulting in arrest in either the G2-phase or metaphase of mitosis, ultimately triggering the abscission checkpoint if non-disjunction persists. These events, which directly or indirectly monitor the activity of Topo IIA, have become of major interest as many cancers have deficiencies in Topoisomerase checkpoints, leading to genome instability. Recent studies into how cells sense Topo IIA dysfunction and respond by regulating cell cycle progression demonstrate that the Topo IIA G2 checkpoint is distinct from the G2-DNA damage checkpoint. Likewise, in mitosis, the metaphase Topo IIA checkpoint is separate from the spindle assembly checkpoint. Here, we integrate mechanistic knowledge of Topo IIA checkpoints with the current understanding of how cells regulate progression through the cell cycle to accomplish faithful genome transmission and discuss the opportunities this offers for therapy.

Topoisomerase II SUMOylation activates a metaphase checkpoint via Haspin and Aurora B kinases.

Topoisomerase II (Topo II) is essential for mitosis since it resolves sister chromatid catenations. Topo II dysfunction promotes aneuploidy and drives cancer. To protect from aneuploidy, cells possess mechanisms to delay anaphase onset when Topo II is perturbed, providing additional time for decatenation. Molecular insight into this checkpoint is lacking. Here we present evidence that catalytic inhibition of Topo II, which activates the checkpoint, leads to SUMOylation of the Topo II C-terminal domain (CTD). This modification triggers mobilization of Aurora B kinase from inner centromeres to kinetochore proximal centromeres and the core of chromosome arms. Aurora B recruitment accompanies histone H3 threonine-3 phosphorylation and requires Haspin kinase. Strikingly, activation of the checkpoint depends both on Haspin and Aurora B. Moreover, mutation of the conserved CTD SUMOylation sites perturbs Aurora B recruitment and checkpoint activation. The data indicate that SUMOylated Topo II recruits Aurora B to ectopic sites, constituting the molecular trigger of the metaphase checkpoint when Topo II is catalytically inhibited.

A noncatalytic function of the topoisomerase II CTD in Aurora B recruitment to inner centromeres during mitosis.

Faithful chromosome segregation depends on the precise timing of chromatid separation, which is enforced by checkpoint signals generated at kinetochores. Here, we provide evidence that the C-terminal domain (CTD) of DNA topoisomerase IIα (Topo II) provides a novel function at inner centromeres of kinetochores in mitosis. We find that the yeast CTD is required for recruitment of the tension checkpoint kinase Ipl1/Aurora B to inner centromeres in metaphase but is not required in interphase. Conserved CTD SUMOylation sites are required for Ipl1 recruitment. This inner-centromere CTD function is distinct from the catalytic activity of Topo II. Genetic and biochemical evidence suggests that Topo II recruits Ipl1 via the Haspin-histone H3 threonine 3 phosphorylation pathway. Finally, Topo II and Sgo1 are equally important for Ipl1 recruitment to inner centromeres. This indicates H3 T3-Phos/H2A T120-Phos is a universal epigenetic signature that defines the eukaryotic inner centromere and provides the binding site for Ipl1/Aurora B.