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Mad2 deficiency provokes a wave of hyperkeratosis in neonates, which largely normalizes in adults. (A) H&E staining of epidermal samples at various time points showing epidermal morphology with hyperkeratotic features in neonates. (B) Hyperkeratotic epidermis shows signs of increased differentiation (evidenced by thickened Krt10 positive epidermis).
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The spindle assembly checkpoint (SAC) ensures correct chromosome segregation during mitosis by preventing aneuploidy, an event that is detrimental to the fitness and survival of normal cells but oncogenic in tumor cells. Deletion of SAC genes is incompatible with early mouse development, and RNAi-mediated depletion of SAC components in cultured cel...
Citations
... CIN and aneuploidy often result in a deregulation of metabolic signalling, thereby disrupting metabolic homeostasis [60][61][62][63]. Altered metabolic signalling is strongly associated with oncogenesis, particularly in tumours marked by hypoxia, glycolysis, and altered levels of oncogenic metabolites [64,65]. ...
Chromosomal instability (CIN) is a prevalent characteristic of solid tumours and haematological malignancies. CIN results in an increased frequency of chromosome mis-segregation events, thus yielding numerical and structural copy number alterations, a state also known as aneuploidy. CIN is associated with increased chances of tumour recurrence, metastasis, and acquisition of resistance to therapeutic interventions, and this is a dismal prognosis. In this review, we delve into the interplay between CIN and cancer, with a focus on its impact on the tumour microenvironment—a driving force behind metastasis. We discuss the potential therapeutic avenues that have resulted from these insights and underscore their crucial role in shaping innovative strategies for cancer treatment.
... The results showed that Mad2 was expressed during all stages of meiotic maturation, and the level was relatively constant from GV to MI but increased significantly at MII (Fig. 1A). To explore the function of Mad2, we conditionally depleted Mad2 in the mouse oocyte using previously described Mad2 Flox/Flox mice (Foijer et al., 2013) in which 'LoxP' sequences flank the Mad2 coding sequence (exons 2 to 5). We crossed Mad2 Flox/Flox mice with Gdf9-Cre mice to generate Mad2 Flox/Flox ; Gdf9-Cre+ female mice (hereafter referred to as Mad2 −/− mice) (Fig. 1B). ...
... The species of animal used in the experiments was Mus musculus. Mad2 Flox/Flox mice were obtained from Allan Bradley's laboratory (Foijer et al., 2013). To obtain oocytes with conditional knockout of Mad2, Gdf9-Cre mice, which express Cre recombinase via the Gdf9 promoter, were crossed with Mad2 Flox/Flox mice. ...
Accurate chromosome segregation, monitored by the spindle assembly checkpoint (SAC), is crucial for the production of euploid cells. Previous in vitro studies by us and others showed that Mad2, a core member of the SAC, performs a checkpoint function in oocyte meiosis. Here, through an oocyte-specific knockout approach in mouse, we reconfirmed that Mad2-deficient oocytes exhibit an accelerated metaphase-to-anaphase transition caused by premature degradation of securin and cyclin B1 and subsequent activation of separase in meiosis I. However, it was surprising that the knockout mice were completely fertile and the resulting oocytes were euploid. In the absence of Mad2, other SAC proteins, including BubR1, Bub3 and Mad1, were normally recruited to the kinetochores, which likely explains the balanced chromosome separation. Further studies showed that the chromosome separation in Mad2-null oocytes was particularly sensitive to environmental changes and, when matured in vitro, showed chromosome misalignment, lagging chromosomes, and aneuploidy with premature separation of sister chromatids, which was exacerbated at a lower temperature. We reveal for the first time that Mad2 is dispensable for proper chromosome segregation but acts to mitigate environmental stress in meiotic oocytes.
... The contradiction between aneuploidy's capacity to act as a promoter and suppressor of malignant cell growth is described as the aneuploidy paradox [154]. To explain this anomaly, aneuploidy-coping mechanisms are proposed, which include changes in gene expression and the accommodation of proteotoxicity and metabolic stress [155,156]. Indeed, extra copies of the TP53 gene are reported to reduce cancer incidence in mice whilst ageing normally [157]. ...
An escape from cellular senescence through the development of unlimited growth potential is one of the hallmarks of cancer, which is thought to be an early event in carcinogenesis. In this review, we propose that the molecular effectors of senescence, particularly the inactivation of TP53 and CDKN2A, together with telomere attrition and telomerase activation, all lead to aneuploidy in the keratinocytes from oral potentially malignant disorders (OPMD). Premalignant keratinocytes, therefore, not only become immortal but also develop genotypic and phenotypic cellular diversity. As a result of these changes, certain clonal cell populations likely gain the capacity to invade the underlying connective tissue. We review the clinical implications of these changes and highlight a new PCR-based assay to identify aneuploid cell in fluids such as saliva, a technique that is extremely sensitive and could facilitate the regular monitoring of OPMD without the need for surgical biopsies and may avoid potential biopsy sampling errors. We also draw attention to recent studies designed to eliminate aneuploid tumour cell populations that, potentially, is a new therapeutic approach to prevent malignant transformations in OPMD.
... For this study, we wanted to address if CIN and the resulting aneuploidy influence the developing cerebellum and cause tumor formation. To this end, we generated a transgenic mouse model that allows the spatiotemporally controlled deletion of mitotic spindle checkpoint component Mad2l1 from developing CGNPs using floxed Mad2l1 alleles and the Math1CreER T2 -tdTomato driver/reporter [18,23,[38][39][40][41][42]. To mitigate potential deleterious effects on CGNP viability and promote tumor formation, we also included the conditional deletion of floxed Trp53, a tumor suppressor gene that is frequently mutated in medulloblastoma [6,40]. ...
... To study CIN and aneuploidy in the developing cerebellum, two additional mouse strains were bred into the above-described model: the conditional Mad2l1 deletion mouse strain [38,39,51], and the conditional Trp53 deletion mouse strain [40]. Four days after birth (P4), the neonates received 0.6 mg of Tamoxifen in a total volume of 30 microliters dissolved in peanut oil by oral gavage. ...
Medulloblastoma is a pediatric brain malignancy that consists of four transcriptional subgroups. Structural and numerical aneuploidy are common in all subgroups, although they are particularly profound in Group 3 and Group 4 medulloblastoma and in a subtype of SHH medulloblastoma termed SHHα. This suggests that chromosomal instability (CIN), the process leading to aneuploidy, is an important player in medulloblastoma pathophysiology. However, it is not known if there is ongoing CIN in medulloblastoma or if CIN affects the developing cerebellum and promotes tumor formation. To investigate this, we performed karyotyping of single medulloblastoma cells and demonstrated the presence of distinct tumor cell clones harboring unique copy number alterations, which is suggestive of ongoing CIN. We also found enrichment for processes related to DNA replication, repair, and mitosis in both SHH medulloblastoma and in the highly proliferative compartment of the presumed tumor cell lineage-of-origin, the latter also being sensitive to genotoxic stress. However, when challenging these tumor cells-of-origin with genetic lesions inducing CIN using transgenic mouse modeling, we found no evidence for large chromosomal aberrations in the cerebellum or for medulloblastoma formation. We therefore conclude that without a background of specific genetic mutations, CIN is not tolerated in the developing cerebellum in vivo and, thus, by itself is not sufficient to initiate medulloblastoma.
... While this general ARP pattern seems to hold true for several aneuploid cell strains [13] and is also observed in vivo (e.g., in aneuploid basal epidermal cells) [32], part of this aneuploidy signature is reverted in aneuploid cancers. For instance, while cultured aneuploid cells appear to have decreased expression of ribosomal genes [13,32], expression these genes is upregulated in a model for CIN-driven T-ALL [33]. ...
... While this general ARP pattern seems to hold true for several aneuploid cell strains [13] and is also observed in vivo (e.g., in aneuploid basal epidermal cells) [32], part of this aneuploidy signature is reverted in aneuploid cancers. For instance, while cultured aneuploid cells appear to have decreased expression of ribosomal genes [13,32], expression these genes is upregulated in a model for CIN-driven T-ALL [33]. Interestingly, as discussed above, stable aneuploid yeast strains also exhibit increased expression of ribosome genes [3], suggesting that yeast cell biology represents the biology of an in vivo cancer cell more than the biology of mammalian cell lines, at least in its response to aneuploidy. ...
... Given that ongoing CIN is incompatible with early embryonic development [120], the most suitable CIN predisposition would be a conditional CIN-driving allele that does not efficiently promote cancer by itself. This could for instance be a Mps1 truncation or mutation allele [33,122], a Mad2 deletion allele [32], a hypomorphic BubR1 allele [123], or a Plk4 overexpression allele [24], as well as any other tissue-specific CIN driver. Indeed, in most of the CIN models, the CIN-driving allele alone leads to aneuploidy but not to rapid tumorigenesis. ...
Chromosomal instability is the process of mis-segregation for ongoing chromosomes, which leads to cells with an abnormal number of chromosomes, also known as an aneuploid state. Induced aneuploidy is detrimental during development and in primary cells but aneuploidy is also a hallmark of cancer cells. It is therefore believed that premalignant cells need to overcome aneuploidy-imposed stresses to become tumorigenic. Over the past decade, some aneuploidy-tolerating pathways have been identified through small-scale screens, which suggest that aneuploidy tolerance pathways can potentially be therapeutically exploited. However, to better understand the processes that lead to aneuploidy tolerance in cancer cells, large-scale and unbiased genetic screens are needed, both in euploid and aneuploid cancer models. In this review, we describe some of the currently known aneuploidy-tolerating hits, how large-scale genome-wide screens can broaden our knowledge on aneuploidy specific cancer driver genes, and how we can exploit the outcomes of these screens to improve future cancer therapy.
... Animal experiments were carried out and animal experiment protocols were described according to the ARRIVE 2.0 guidelines 27 . The conditional Mad2 f/f and Mps1 f/f animal models were previously described 10,25,26 . Mice used in OPEN show many apoptotic cells (zoomed-in sections Fig. 2A), indicating that these cells indeed cope very poorly with CIN. ...
... While this phenotype is intriguing www.nature.com/scientificreports/ and suggests that Mad2 loss is particularly toxic to the stem cells residing in the intestinal crypts, similar to what was observed previously for hair follicle stem cells 25 , the severity of the phenotype also precluded assessment of Mad2 loss in other adult tissues. ...
... However, to our knowledge, the consequences of complete systemic SAC alleviation in adult mice have not been reported so far. We have previously shown that tissue-specific inactivation of the SAC can circumvent embryonic lethality associated with Mad2 loss 25,26 , and found that different tissues cope differently with SAC loss. For instance, inactivation in the epidermis revealed that SAC alleviation is not tolerated by hair follicle stem cells, but remarkably well-tolerated by the basal cells of the epidermis 25 . ...
Chromosomal instability (CIN) is a hallmark of cancer, leading to aneuploid cells. To study the role that CIN plays in tumor evolution, several mouse models have been engineered over the last 2 decades. These models have unequivocally shown that systemic high-grade CIN is embryonic lethal. We and others have previously shown that embryonic lethality can be circumvented by provoking CIN in a tissue-specific fashion. In this study, we provoke systemic high-grade CIN in adult mice as an alternative to circumvent embryonic lethality. For this, we disrupt the spindle assembly checkpoint (SAC) by alleviating Mad2 or truncating Mps1, both essential genes for SAC functioning, with or without p53 inactivation. We find that disruption of the SAC leads to rapid villous atrophy, atypia and apoptosis of the epithelia of the jejunum and ileum, substantial weight loss, and death within 2–3 weeks after the start of the CIN insult. Despite this severe intestinal phenotype, most other tissues are unaffected, except for minor abnormalities in spleen, presumably due to the lower proliferation rate in these tissues. We conclude that high-grade CIN in vivo in adult mice is most toxic to the high cell turnover intestinal epithelia.
... and Cdc20 MCC (Foijer et al., 2013;Kollu et al., 2015). where APC/C CDC20 is associated with self-renewal activity of stem cells by interacting with pluripotency-related transcription factor SOX2 (Mao et al., 2015). ...
Damage to the genome can accelerate aging. The percentage of aneuploid cells, that is, cells with an abnormal number of chromosomes, increases during aging; however, it is not clear whether increased aneuploidy accelerates aging. Here, we report an individual showing premature aging phenotypes of various organs including early hair loss, atrophic skin, and loss of hematopoietic stem cells; instability of chromosome numbers known as mosaic variegated aneuploidy (MVA); and spindle assembly checkpoint (SAC) failure. Exome sequencing identified a de novo heterozygous germline missense mutation of c.856C>A (p.R286S) in the mitotic activator CDC20. The mutant CDC20 showed lower binding affinity to BUBR1 during the formation of the mitotic checkpoint complex (MCC), but not during the interaction between MCC and the anaphase‐promoting complex/cyclosome (APC/C)–CDC20 complex. While heterozygous knockout of CDC20 did not induce SAC failure, knock‐in of the mutant CDC20 induced SAC failure and random aneuploidy in cultured cells, indicating that the particular missense mutation is pathogenic probably via the resultant imbalance between MCC and APC/C‐CDC20 complex. We postulate that accelerated chromosome number instability induces premature aging in humans, which may be associated with early loss of stem cells. These findings could form the basis of a novel disease model of the aging of the body and organs.
... The existence of such mechanisms is further supported by in vivo studies. For instance, the loss of Mad2 is tolerated by epidermal cells in mouse skin, but hair follicle stem cells are eliminated as a result of apoptotic cell death, which suggests that epidermal basal cells have an aneuploidy-tolerating mechanism in place, while hair follicle stem cells do not [47]. Furthermore, aneuploid cells have been found within a variety of somatic cell types while aneuploidy appears to be less common in stem cells, further suggesting that stem cells have dedicated mechanisms such as special checkpoints to circumvent the propagation of aneuploid cells [48]. ...
Aneuploidy, an irregular number of chromosomes in cells, is a hallmark feature of cancer. Aneuploidy results from chromosomal instability (CIN) and occurs in almost 90% of all tumours. While many cancers display an ongoing CIN phenotype, cells can also be aneuploid without displaying CIN. CIN drives tumour evolution as ongoing chromosomal missegregation will yield a progeny of cells with variable aneuploid karyotypes. The resulting aneuploidy is initially toxic to cells because it leads to proteotoxic and metabolic stress, cell cycle arrest, cell death, immune cell activation and further genomic instability. In order to overcome these aneuploidy-imposed stresses and adopt a malignant fate, aneuploid cancer cells must develop aneuploidy-tolerating mechanisms to cope with CIN. Aneuploidy-coping mechanisms can thus be considered as promising therapeutic targets. However, before such therapies can make it into the clinic, we first need to better understand the molecular mechanisms that are activated upon aneuploidization and the coping mechanisms that are selected for in aneuploid cancer cells. In this review, we discuss the key biological responses to aneuploidization, some of the recently uncovered aneuploidy-coping mechanisms and some strategies to exploit these in cancer therapy.
... In human cancers, high expression of MAD2 is also a common feature, with conflicting data regarding prognosis [13]. These contradictory data come from the fact that the tolerance of CIN is tissue specific; thus, the absence of MAD2 is incompatible with embryonic development [26] and toxic to hair follicle stem cells [27], but tolerated by basal epidermal cells [27], T cells and hepatocytes [28]. We propose here that there is a threshold of MAD2 levels in the tumor and in GCSCs, and levels that fall below the "minimum" will affect GCSCs stemness characteristics, minimizing their self-renewal and tumorigenic potential. ...
... In human cancers, high expression of MAD2 is also a common feature, with conflicting data regarding prognosis [13]. These contradictory data come from the fact that the tolerance of CIN is tissue specific; thus, the absence of MAD2 is incompatible with embryonic development [26] and toxic to hair follicle stem cells [27], but tolerated by basal epidermal cells [27], T cells and hepatocytes [28]. We propose here that there is a threshold of MAD2 levels in the tumor and in GCSCs, and levels that fall below the "minimum" will affect GCSCs stemness characteristics, minimizing their self-renewal and tumorigenic potential. ...
Rationale: Gastric cancer (GC) is a solid tumor that contains subpopulations of cancer stem cells (CSCs), which are considered drivers of tumor initiation and metastasis; responsible for therapeutic resistance; and promoters of tumor relapse. The balance between symmetric and asymmetric division is crucial for stem cell maintenance. The objective of this study is to evaluate the role of MAD2, a key protein for proper mitotic checkpoint activity, in the tumorigenesis of GC.
Methods: Gastric cancer stem cells (GCSCs) were obtained from MKN45, SNU638 and ST2957 cell lines. Pluripotency and stemness markers were evaluated by RT-qPCR and autofluorescence and membrane markers by flow cytometry. Relevant signal transduction pathways were studied by WB. We analysed cell cycle progression, migration and invasion after modulation of MAD2 activity or protein expression levels in these in vitro models. In vivo assays were performed in a nude mouse subcutaneous xenograft model.
Results: We found that NANOG, CXCR4 and autofluorescence are common and consistent markers for the GCSCs analysed, with other markers showing more variability. The three main signalling pathways (Wnt/β-catenin; Hedgehog and Notch) were activated in GCSCs. Downregulation of MAD2 in MKN45CSCs decreased the expression of markers CXCR4, CD133, CD90, LGR5 and VIM, without affecting cell cycle profile or therapy resistance. Moreover, migration, invasion and tumor growth were clearly reduced, and accordingly, we found that metalloprotease expression decreased. These results were accompanied by a reduction in the levels of transcription factors related with epithelial-to-mesenchymal transition.
Conclusions: We can conclude that MAD2 is important for GCSCs stemness and its downregulation in MKN45CSCs plays a central role in GC tumorigenesis, likely through CXCR4-SNAI2-MMP1. Thus, its potential use in the clinical setting should be studied as its functions appear to extend beyond mitosis.
... In mice predisposed to cancer, CIN is either neutral [30][31][32] , promotes tumor formation 23,26,27,30,[33][34][35][36][37][38][39] , or, in some conditions, suppresses it 29,30,32,37 . Comparisons between these studies is however exceedingly difficult due to the use of different oncogenic backgrounds, to differences in tissues that were examined 40 , and to the manner and time by which the tissues were exposed to CIN. Moreover, technical limitations often precluded direct measurements of CIN in the relevant tissues, and oncogenic effects in the different models cannot be attributed to distinct CIN levels. ...
Most human cancers are aneuploid, due to a chromosomal instability (CIN) phenotype. Despite being hallmarks of cancer, however, the roles of CIN and aneuploidy in tumor formation have not unequivocally emerged from animal studies and are thus still unclear. Using a conditional mouse model for diverse degrees of CIN, we find that a particular range is sufficient to drive very early onset spontaneous adenoma formation in the intestine. In mice predisposed to intestinal cancer (ApcMin/+), moderate CIN causes a remarkable increase in adenoma burden in the entire intestinal tract and especially in the distal colon, which resembles human disease. Strikingly, a higher level of CIN promotes adenoma formation in the distal colon even more than moderate CIN does, but has no effect in the small intestine. Our results thus show that CIN can be potently oncogenic, but that certain levels of CIN can have contrasting effects in distinct tissues.
