SAURINLAB
THE ART OF SCIENCE
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Main research articles
2013
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Started Lab
In2TheScience
(short video or audio descriptions of each manuscript)
- 2025-
Pareri et al (2025) investigates a fundamental mechanism that may contribute to drug resistance in breast cancer. They show that CDK4/6 inhibitors - which typically cause cell enlargement that leads to senescence – can cause mitotic errors and chromosomal changes in enlarged cells that evade senescence, such as those lacking p53. Mechanistically, the authors demonstrate that the increased cytoplasmic volume weakens the mitotic checkpoint (SAC), as the inhibitory complexes produced at kinetochores are diluted and cannot efficiently halt cell division. Furthermore, the enlargement impairs sister chromatid cohesion by causing the delocalization of the protective protein Sgo1 from centromeres, resulting in attachment errors. Crucially, these mitotic and genetic defects are size-dependent, as co-treatment with mTOR inhibitors successfully prevents cell overgrowth and rescues chromosome segregation fidelity. This suggests a pathway by which p53-null cells use their enlarged size to acquire drug-resistant karyotypes and genotypes. This may be relevant for the emergence of drug resistance in the clinical since this is frequently associated with p53 loss.
Allen at al (2025b) investigates how the tumour suppressor p53 restricts the growth of cells that have undergone whole genome doubling (WGD), a frequent event in cancer evolution. P53 was known to sense extra centrosomes to WGD cells, but inducing WGD without centrosome duplication the authors show p53 still arrests these cells but via the mitotic stopwatch, a p53/53BP1/USP28-dependent pathway that typical arrested cells after extended mitotic delays. They found that WGD significantly reduces the amount of mitotic delay required to trigger the pathway, effectively lowering the stopwatch threshold. This heightened sensitivity is associated with elevated basal p21 protein levels in G2 cells, which amplify the stopwatch signal to enforce G1 arrest. Similar sensitization is observed in enlarged diploid cells caused by CDK4/6 inhibitor treatment, confirming that the stopwatch is a key sensor for both increased genome size and cell enlargement. The findings suggest that the mitotic stopwatch is a critical mechanism for restraining abnormal proliferation, which has important implications for cancer therapy.
Allen at al (2025a) details the development and application of a new chemical-genetic strategy to rapidly and specifically inhibit the PP2A-B56 phosphatase complex, designated "directSLiMs”. Researchers overcame the issues of targeting the enzymes catalytic core – which inhibits many other related phosphatase complexes - by targeting the enzyme's mechanism of substrate recognition instead. This relies on the drug-inducible binding of Short Linear Motifs (SLiMs) to the B56 regulatory subunit, which block the substrate binding pocket and compete of endogenous substrates with seconds. Using phosphoproteomic analysis, the study identified a wide range of mitotic substrates regulated by PP2A-B56. The system was used to reveal that this phosphatase complex plays an essential role in stabilizing chromosome alignment during metaphase by counteracting Aurora B activity at the kinetochore-microtubule interface. This strategy provides a generalizable method for studying other enzymes whose function depends on similar SLiM-based interactions.
- 2024 -
Coates et al (2024) examines the cytotoxic mechanism of Elraglusib, a drug currently in clinical trials and originally conceived as a glycogen synthase kinase-3 (GSK3) inhibitor. The study concludes that the anti-cancer activity of Elraglusib is independent of GSK3 inhibition, as structurally distinct GSK3 inhibitors did not produce similar effects across various tumor cell lines. Instead, the compound acts as a direct microtubule destabilizer, significantly impairing microtubule (MT) polymerization and increasing soluble tubulin fractions. This destabilization prevents proper chromosome alignment and attachment, inducing a cell-cycle delay known as mitotic arrest. Following this prolonged arrest, the cells undergo mitotic slippage, which results in catastrophic DNA damage and subsequent apoptosis. These findings are highly significant for optimizing clinical trial design and developing appropriate biomarkers, which should shift focus away from GSK3-related mechanisms.
Foy et al. (2024) argues the search for CDK4/6 inhibitor biomarkers has been fundamentally flawed due to reliance on inappropriate proliferation assays. The authors demonstrate that while CDK4/6 inhibitors cause cells to arrest in the G1 phase of the cell cycle, the cells often continue to grow in size and produce more mitochondria, which leads metabolic, ATP-based assays (like CellTiter-Glo) to incorrectly report cell proliferation. Using DNA-based assays (like CyQuant), which accurately measure cell number regardless of size, the researchers re-analyzed existing data, uncovering expected co-dependencies (such as Cyclin D1, CDK4, and CDK6 for sensitive cells) and potential biomarkers that were obscured by previous methods. The findings stress the urgent need to re-screen anti-cancer drugs that halt cell division but permit cell growth using appropriate, size-independent proliferation assays to accurately identify biomarkers and inform clinical trials.
- 2023 -
A short video overview is above but click below for a more detailed podcast audio description.
To read commentary articles for Foy and Crozier et al (2023) in Molecular Cell: click here
Foy et al (2023) and Crozier et al (2023) outlines the cellular mechanisms by which CDK4/6 inhibitors, used in breast cancer treatment, promote long-term cellular senescence. The key finding is that these inhibitors block proliferation but not cell growth, leading to detrimental cellular overgrowth during the G0/G1 arrest. This enlarged state triggers a biphasic stress response mediated by the inhibitor protein p21, which drives permanent cell cycle withdrawal. Initially, overgrowth causes osmotic stress and p38-dependent activation of the p53-p21 pathway, maintaining the G0/G1 arrest. However, for cells that escape and re-enter S-phase, the hypertrophy leads to debilitating replication stress, which prompts a second wave of p21 expression forcing cycle exit from G2. These data suggest that high p21 protein levels integrate signals from both osmotic and replication stress to determine the fate of overgrown cells. Importantly, these effect are exacerbate by oncogenic signals that drive constitutive growth signals in cancer cells. So, CDK4/6 inhibits turn oncogenic signals against cancer cells by causing them to drive toxic cell overgrowth that leads to senescence.
Corno et al (2023) investigates a fundamental mechanism regulating cell division, focusing on a bifunctional kinase–phosphatase module that is conserved throughout the animal kingdom and is essential for preserving genome stability. This module consists of binding motifs for the kinase PLK1 and the phosphatase PP2A-B56, which localized these enzymes within 50 aminoc acids of each other on the BUBR1 protein at kinetochores. The authors demonstrate that these enzymes regulate each other through an intramolecular negative feedback loop necessary for generating precise and dynamic signals during mitosis. This critical interplay ensures the correct timing and strength of the mitotic checkpoint (SAC) while also stabilizing necessary kinetochore–microtubule attachments (KT-MT). Furthermore, the number of KNL1's MELT motifs, which acts as a scaffold for the module by recruiting BUBR1 to kinetochores, dictates the overall levels of these regulatory enzymes, thereby tuning checkpoint strength. When this balance is disturbed, either by genetic manipulation or by altering the KNL1 scaffold, cells exhibit severe defects in chromosome alignment and segregation. This article therefore provides a fascinating example of how kinases and phosphatase can both be needed at the same protein to regulate different processes.
- 2022 -
To read a commentary article for Crozier et al (2022) in EMBO J: click here
Crozier et al (2022) details the mechanisms by which CDK4/6 inhibitors induce long-term cell cycle withdrawal in cancer cells, a process essential for their clinical efficacy. The study explains that while these drugs initially cause a temporary G1 arrest, prolonged exposure leads to significant replication stress due to the loss of key DNA replication components, such as the MCM complex. The cell's response to this stress is dependent on its tumor-suppressing pathways, specifically p53 status. Cells with functional p53 undergo permanent cell cycle withdrawal, whereas p53-deficient cells fail to arrest and instead enter mitosis with under-replicated DNA, often resulting in catastrophic cell division and genomic instability. This mechanistic understanding suggests new therapeutic approaches, including the combination of CDK4/6 inhibitors with agents that increase replication stress to enhance tumor cell destruction.
- 2020 -
Commentary article for Cordeiro et al (2020) in J Cell Biol: click here
Cordeiro et al (2020) examines the precise function of kinetochore phosphatases PP1 and PP2A-B56 in suppressing the mitotic spindle assembly checkpoint (SAC) once chromosome attachment is achieved. A crucial event in SAC silencing is known to be the dephosphorylation of the KNL1-MELT repeats, which is controlled by PP1 and PP2A-B56. This paper demonstrates that the primary role of PP1 and PP2A-B56 is to prevent PLK1 association with the BUB complex, thereby preventing this kinase from a self-amplifying loop that keeps the MELT motifs phosphorylated. Strikingly, the study demonstrates that the MELT repeats can be dephosphorylated normally, even under high phosphatase inhibition, provided that PLK1 activity is simultaneously blocked. These findings indicate that the central mechanism by which kinetochore phosphatases control SAC exit is by restraining and extinguishing the autonomous PLK1 kinase activity, rather than directly acting as the main dephosphorylating enzyme for the SAC scaffold itself, as previously assumed.
Commentary article for Allan et al (2020) in EMBO J: click here
Commentary article for Allan et al (2020) in J Cell Biol: click here
Allan et al (2020) investigates a crucial regulatory mechanism within the spindle assembly checkpoint (SAC), which ensures chromosomes attach correctly to microtubules before cell division. The study focuses on how the vital checkpoint component MAD1 is recruited to the kinetochore corona, a step previously not understood mechanistically. Through a series of biochemical and cellular experiments, the authors demonstrate that the cell cycle regulatory protein Cyclin B1 serves as the direct receptor, binding to the N-terminus of MAD1 and acting as a scaffold. This binding interaction is critical for the stability and robustness of the SAC, enabling the checkpoint to remain active and tolerate reductions in upstream signalling from the MPS1 kinase. In essence, the Cyclin B1 scaffolding function ensures the prolonged and efficient maintenance of SAC machinery necessary for proper chromosome segregation. This was the first identified scaffolding role for the main mitotic cyclin which typically functions in complex with CDK1 to phosphorylate mitotic proteins.
- 2019 -
Commentary article for Smith et al (2019) in prelights click here
Smith et al (2019) examines the functional divergence of two ubiquitously expressed phosphatases, PP1 and PP2A-B56, at the kinetochore, a structure critical for cell division. These phosphatases localise to an almost identical position at the kinetochore but they regulate distinct mitotic processes, with PP2A-B56 regulating microtubule attachment and PP1 controlling checkpoint silencing. The authors establish that this functional specificity is not based on the phosphatases’ catalytic differences or their exact location, but rather on their unique ability to respond to kinase signaling through opposite phosphorylation inputs. They bind to the kinetochore via short linear motifs (SLiMs) and this interaction can be either inhibited (PP1) or enhance (PP2A-B56) by phosphorylation inputs. Mathematical modelling confirms that this inverse regulatory mechanism generates unique cross-regulation and feedback loops, allowing the two otherwise identical phosphatases to govern distinct mitotic processes essential for chromosome stability. Evolutionary analysis suggest these main two phosphatase families have likely evolved to respond to phosphorylation inputs in opposite ways, likely explaining why they are two of the most important and well-conserved phosphatase in eukaryotic cells.
Vallardi et al (2019) examines the function of PP2A-B56 isoforms, crucial serine/threonine phosphatases that regulate key stages of mitosis in human cells. The central finding demonstrates that different B56 isoforms exhibit specific localization patterns, contradicting previous assumptions that they were redundant. For instance, the research revealed that B56α and B56ε localize primarily to the centromere, whereas B56γ and B56δ are found mainly at the kinetochore, establishing a division of labor that governs distinct biological processes. This functional specialization dictates that only B56α can successfully maintain sister chromatid cohesion, while B56γ is essential for proper chromosome alignment and the spindle assembly checkpoint. Mechanistic analysis showed that this specificity is achieved through differential binding to receptors (Sgo2 at the centromere and BubR1 at the kinetochore), which is controlled by a small C-terminal loop within the B56 subunit structure. These results explain how slight structural differences between isoforms lead to major differences in subcellular targeting and mitotic regulation.
- 2018 -
- 2014 -
Skowyra et al (2019) analyses the molecular function of the deubiquitinase USP9X and demonstrates a vital role in regulating cell division. Findings indicate that USP9X enhances the strength of the Spindle Assembly Checkpoint (SAC), the mechanism responsible for delaying anaphase until chromosomes are correctly aligned. Mechanistically, USP9X functions by restraining the ubiquitin ligase APC/C, preventing it from accelerating the turnover and subsequent degradation of its inhibitor, the Mitotic Checkpoint Complex (MCC). When USP9X is lost, the resulting rapid MCC turnover weakens the SAC, which causes frequent chromosome mis-segregations and contributes to chromosomal instability (CIN). This connection between USP9X, SAC failure, and CIN may explain why reduced USP9X is linked to tumorigenesis, proposing that targeting the enzyme could be a therapeutic avenue for certain cancers.
Nijenhuis et al (2014) investigates the Spindle Assembly Checkpoint (SAC) signaling system at the kinetochores, finding that its rapid responsiveness is managed by an internal regulatory mechanism. The authors define a localized negative feedback loop crucial for rapidly switching the SAC signal between its on and off states during cell division. When the SAC is active, the phosphatase PP2A-B56 is recruited, where it works against the kinase Aurora B to enhance cellular response. This antagonism promotes the recruitment of the second phosphatase, PP1, which subsequently functions to silence the checkpoint signal and remove PP2A-B56. This sophisticated cycle ensures that the kinetochore can quickly extinguish the active checkpoint state upon force-producing microtubule attachment, thereby facilitating stable genome stability.
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