In meiosis, two specialized cell divisions allow the separation of paired chromosomes first, then of sister chromatids. Separase removes the cohesin complex holding sister chromatids together in a stepwise manner from chromosome arms in meiosis I, then from the centromere region in meiosis II. Using mouse oocytes, our study reveals that cyclin A2 promotes entry into meiosis, as well as an additional unexpected role; namely, its requirement for separase-dependent sister chromatid separation in meiosis II. Untimely cyclin A2-associated kinase activity in meiosis I leads to precocious sister separation, whereas inhibition of cyclin A2 in meiosis II prevents it. Accordingly, endogenous cyclin A is localized to kinetochores throughout meiosis II, but not in anaphase I. Additionally, we found that cyclin B1, but not cyclin A2, inhibits separase in meiosis I. These findings indicate that separase-dependent cohesin removal is differentially regulated by cyclin B1 and A2 in mammalian meiosis.

Mounting evidence indicates that vitamin C has the potential to be a potent anti-cancer agent when administered intravenously and in high doses (high-dose IVC). Early phase clinical trials have confirmed safety and indicated efficacy of IVC in eradicating tumour cells of various cancer types. In recent years, the multi-targeting effects of vitamin C were unravelled, demonstrating a role as cancer-specific, pro-oxidative cytotoxic agent, anti-cancer epigenetic regulator and immune modulator, reversing epithelial-to-mesenchymal transition, inhibiting hypoxia and oncogenic kinase signalling and boosting immune response. Moreover, high-dose IVC is powerful as an adjuvant treatment for cancer, acting synergistically with many standard (chemo-) therapies, as well as a method for mitigating the toxic side-effects of chemotherapy. Despite the rationale and ample evidence, strong clinical data and phase III studies are lacking. Therefore, there is a need for more extensive awareness of the use of this highly promising, non-toxic cancer treatment in the clinical setting. In this review, we provide an elaborate overview of pre-clinical and clinical studies using high-dose IVC as anti-cancer agent, as well as a detailed evaluation of the main known molecular mechanisms involved. A special focus is put on global molecular profiling studies in this respect. In addition, an outlook on future implications of high-dose vitamin C in cancer treatment is presented and recommendations for further research are discussed.

The universal triggering event of eukaryotic chromosome segregation is cleavage of centromeric cohesin by separase. Prior to anaphase, most separase is kept inactive by association with securin. Protein phosphatase 2A (PP2A) constitutes another binding partner of human separase, but the functional relevance of this interaction has remained enigmatic. We demonstrate that PP2A stabilizes separase-associated securin by dephosphorylation, while phosphorylation of free securin enhances its polyubiquitylation by the ubiquitin ligase APC/C and proteasomal degradation. Changing PP2A substrate phosphorylation sites to alanines slows degradation of free securin, delays separase activation, lengthens early anaphase, and results in anaphase bridges and DNA damage. In contrast, separase-associated securin is destabilized by introduction of phosphorylation-mimetic aspartates or extinction of separase-associated PP2A activity. G2-or prometaphase-arrested cells suffer from unscheduled activation of separase when endogenous securin is replaced by aspartate-mutant securin. Thus, PP2A-dependent stabilization of separase-associated securin prevents precocious activation of separase during checkpointmediated arrests with basal APC/C activity and increases the abruptness and fidelity of sister chromatid separation in anaphase.

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Exosomes are extracellular vesicles (EVs) released from cells under both physiological and pathological conditions, and may, thus, be present in biofluids. Urine is one of the most accessible biofluids implemented in clinical diagnostics. Recent mass spectrometry (MS)‐based proteomic analyses have enabled high‐throughput, deep proteome profiling of urinary EVs for the discovery, quantification and characterization of cancer‐specific exosome biomarkers. The protein cargo of urine exosomes is emerging as an attractive source for biomarkers, not only for urological cancers, such as prostate, bladder and kidney cancer, but potentially also for nonurological cancers, including gastric, lung, oesophageal and colorectal cancer. More recently, exosome proteomics dissected protein cargo in the lumen and at the surface of EVs, and unexpectedly indicated that RNA‐ and DNA‐binding proteins might also be present on vesicular surfaces. Here, we analyse MS‐based proteomic data on urinary exosomes from cancer patients, and discuss the potential of urinary exosome‐derived biomarkers in cancer.

Summary Small-cell lung cancer is the most aggressive type of lung cancer, characterized by a remarkable response to chemotherapy followed by development of resistance. Here, we describe SCLC subtypes in Mycl- and Nfib-driven GEMM that include CDH1-high peripheral primary tumor lesions and CDH1-negative, aggressive intrapulmonary metastases. Cisplatin treatment preferentially eliminates the latter, thus revealing a striking differential response. Using a combined transcriptomic and proteomic approach, we find a marked reduction in proliferation and metabolic rewiring following cisplatin treatment and present evidence for a distinctive metabolic and structural profile defining intrinsically resistant populations. This offers perspectives for effective combination therapies that might also hold promise for treating human SCLC, given the very similar response of both mouse and human SCLC to cisplatin.

The onset of anaphase is triggered by the activation of a sitespecific protease called separase. Separase cleaves the chromosomal cohesins holding the duplicated sister chromatids together, allowing sisters to simultaneously separate and segregate to opposite ends of the cell before division. Activated separase cleaves not only cohesin, but also itself; however, the biological significance of separase self-cleavage has remained elusive. Before anaphase, separase is inhibited by at least two mechanisms. The first involves the binding of securin, whereas the second requires the phosphorylation-dependent binding of cyclin-dependent kinase 1 (Cdk1)/cyclin B1. Because securin and Cdk1/cyclin B1 interact with separase in a mutually exclusive manner, the degradation of both these inhibitors plays an important role in activating separase at anaphase. Here we identify a new separase interacting partner, a specific subtype of the heterotrimeric protein phosphatase 2A (PP2A). PP2A associates with separase through the B (B56) regulatory subunit and does so independently of securin and cyclin B1 binding. The association of PP2A with separase requires a 55-amino acid domain closely juxtaposed to separase autocleavage sites. Strikingly, mutation of these cleavage sites increases PP2A binding, suggesting that separase cleavage disrupts the interaction of PP2A with separase. Furthermore, expression of a non-cleavable separase, but not a non-cleavable mutant that cannot bind PP2A, causes a premature loss of centromeric cohesion. Together these observations provide a new mechanistic insight into a physiological function for separase self-cleavage.Each time a cell divides it must first faithfully duplicate its chromosomes and then accurately distribute the sister chromatids between the two daughter cells. After their generation during S phase, sister chromatids are held together through the action of the multisubunit chromosomal cohesion complex (1, 2). During mitosis, cohesin is dissolved, and sister chromatids are segregated toward opposite ends of the cell. In all eukaryotes studied chromosome segregation is initiated by the activation of a site-specific protease known as separase (3, 4). Separase cleaves the cohesins linking each duplicated chromatid pair, allowing sisters to separate simultaneously at anaphase. Because the separation of sister chromatids is an irreversible event, it is clearly vital that separase activation is tightly regulated; activation before the correct attachment of all chromosomes on the microtubule spindle risks chromosome missegregation and the production of aneuploid daughter cells (5).Curiously, upon activation, vertebrate separase cleaves not only cohesin but also itself. Cleavage occurs at three closely spaced sites in between separase conserved C-terminal catalytic domain and its large N-terminal portion (6, 7). Once cleaved, the N-and C-terminal fragments of separase remain associated and are catalytically active. However, separase cleavage is not required for separase activation in vitro. Thus,...

Background: Separase, the trigger protease of eukaryotic anaphase, remains regulated in the absence of its inhibitor, securin. Results: Cdk1-cyclin B1 triggers precipitation of separase by phosphorylation but stabilizes it by inhibitory binding. Conclusion: Only separase that is first complexed by Cdk1-cyclin B1 can later be activated by cyclin B1 degradation. Significance: These minimal requirements of separase regulation could explain the faithful execution of anaphase in the absence of securin.