Mitosis-specific role of ATR: The ATR (ataxia telangiectasia mutated and Rad3-related) kinase plays important roles in the S phase and during the DNA damage response to safeguard genome integrity. Kabeche et al. identified a distinct ATR activation pathway in mitosis that is also critical for suppressing genome instability (see the Perspective by Saldivar and Cimprich). ATR is recruited by Aurora A and activated by R loops at centromeres of mitotic chromosomes; this leads to Aurora B activation, which is necessary for accurate chromosome segregation. This mitotic, R loop–driven ATR signaling pathway could potentially be exploited in the search for cancer therapeutics. Science , this issue p. 108 ; see also p. 30

Cyclin A is shown to maintain unstable kinetochore–microtubule (k–MT) attachments in prometaphase in order to allow for error correction; at the prometaphase–metaphase switch, k-MT attachments are stabilized when cyclin A drops below threshold levels. During metaphase, chromosomes become aligned at the mitotic spindle through the formation of bi-oriented attachments of kinetochores to spindle microtubules. However, kinetochore–microtubule attachment errors can arise in early prometaphase even in cells that correctly segregate chromosomes. In this study, Lilian Kabeche and Duane Compton demonstrate that cyclin A maintains unstable kinetochore–microtubule attachments at prometaphase in order to allow for error correction. At the switch from prometaphase to metaphase, cyclin A is degraded resulting in stable kinetochore–microtubule attachments. These results suggest that cyclin A functions as a timer in prometaphase to ensure efficient error correction and prevent aneuploidy. The most conspicuous event in the cell cycle is the alignment of chromosomes in metaphase. Chromosome alignment fosters faithful segregation through the formation of bi-oriented attachments of kinetochores to spindle microtubules. Notably, numerous kinetochore–microtubule (k–MT) attachment errors are present in early mitosis (prometaphase)1, and the persistence of those errors is the leading cause of chromosome mis-segregation in aneuploid human tumour cells that continually mis-segregate whole chromosomes and display chromosomal instability2,3,4,5,6,7. How robust error correction is achieved in prometaphase to ensure error-free mitosis remains unknown. Here we show that k–MT attachments in prometaphase cells are considerably less stable than in metaphase cells. The switch to more stable k–MT attachments in metaphase requires the proteasome-dependent destruction of cyclin A in prometaphase. Persistent cyclin A expression prevents k–MT stabilization even in cells with aligned chromosomes. By contrast, k–MTs are prematurely stabilized in cyclin-A-deficient cells. Consequently, cells lacking cyclin A display higher rates of chromosome mis-segregation. Thus, the stability of k–MT attachments increases decisively in a coordinated fashion among all chromosomes as cells transit from prometaphase to metaphase. Cyclin A creates a cellular environment that promotes microtubule detachment from kinetochores in prometaphase to ensure efficient error correction and faithful chromosome segregation.

Many cancers display both structural (s-CIN) and numerical (w-CIN) chromosomal instabilities. Defective chromosome segregation during mitosis has been shown to cause DNA damage that induces structural rearrangements of chromosomes (s-CIN). In contrast, whether DNA damage can disrupt mitotic processes to generate whole chromosomal instability (w-CIN) is unknown. Here, we show that activation of the DNA-damage response (DDR) during mitosis selectively stabilizes kinetochore–microtubule (k-MT) attachments to chromosomes through Aurora-A and PLK1 kinases, thereby increasing the frequency of lagging chromosomes during anaphase. Inhibition of DDR proteins, ATM or CHK2, abolishes the effect of DNA damage on k-MTs and chromosome segregation, whereas activation of the DDR in the absence of DNA damage is sufficient to induce chromosome segregation errors. Finally, inhibiting the DDR during mitosis in cancer cells with persistent DNA damage suppresses inherent chromosome segregation defects. Thus, the DDR during mitosis inappropriately stabilizes k-MTs, creating a link between s-CIN and w-CIN. Significance: The genome-protective role of the DDR depends on its ability to delay cell division until damaged DNA can be fully repaired. Here, we show that when DNA damage is induced during mitosis, the DDR unexpectedly induces errors in the segregation of entire chromosomes, thus linking structural and numerical chromosomal instabilities. Cancer Discov; 4(11); 1281–9. ©2014 AACR. See related commentary by Jelluma and Kops, p. 1256 This article is highlighted in the In This Issue feature, p. 1243

TGF-β secreted by tumor stroma induces epithelial-to-mesenchymal transition (EMT) in cancer cells, a reversible phenotype linked to cancer progression and drug resistance. However, exposure to stromal signals may also lead to heritable changes in cancer cells, which are poorly understood. We show that epithelial cells failing to undergo proliferation arrest during TGF-β-induced EMT sustain mitotic abnormalities due to failed cytokinesis, resulting in aneuploidy. This genomic instability is associated with the suppression of multiple nuclear envelope proteins implicated in mitotic regulation and is phenocopied by modulating the expression of LaminB1. While TGF-β-induced mitotic defects in proliferating cells are reversible upon its withdrawal, the acquired genomic abnormalities persist, leading to increased tumorigenic phenotypes. In metastatic breast cancer patients, increased mesenchymal marker expression within single circulating tumor cells is correlated with genomic instability. These observations identify a mechanism whereby microenvironment-derived signals trigger heritable genetic changes within cancer cells, contributing to tumor evolution.

Accurate chromosome segregation during mitosis relies on a dynamic kinetochore (KT)–microtubule (MT) interface that switches from a labile to a stable condition in response to correct MT attachments. This transition is essential to satisfy the spindle-assembly checkpoint (SAC) and couple MT-generated force with chromosome movements, but the underlying regulatory mechanism remains unclear. In this study, we show that during mitosis the MT- and KT-associated protein CLASP2 is progressively and distinctively phosphorylated by Cdk1 and Plk1 kinases, concomitant with the establishment of KT–MT attachments. CLASP2 S1234 was phosphorylated by Cdk1, which primed CLASP2 for association with Plk1. Plk1 recruitment to KTs was enhanced by CLASP2 phosphorylation on S1234. This was specifically required to stabilize KT–MT attachments important for chromosome alignment and to coordinate KT and non-KT MT dynamics necessary to maintain spindle bipolarity. CLASP2 C-terminal phosphorylation by Plk1 was also required for chromosome alignment and timely satisfaction of the SAC. We propose that Cdk1 and Plk1 mediate a fine CLASP2 “phospho-switch” that temporally regulates KT–MT attachment stability.