Abstract
SummaryHuman pluripotent stem cells (hPSCs) are susceptible to numerical and structural chromosomal alterations during long-term culture. We show that mitotic errors occur frequently in hPSCs and that prometaphase arrest leads to very rapid apoptosis in undifferentiated but not in differentiated cells. hPSCs express high levels of proapoptotic protein NOXA in undifferentiated state. Knocking out NOXA by CRISPR or upregulation of the anti-apoptosis gene BCL-XL significantly reduced mitotic cell death, allowing the survival of aneuploid cells and the formation of teratomas significantly larger than their wild-type parental hPSCs. These results indicate that the normally low threshold of apoptosis in hPSCs can safeguard their genome integrity by clearing cells undergoing abnormal division. The amplification of BCL2L1 on chromosome 20q11.21, a frequent mutation in hPSCs, although not directly oncogenic, reduces the sensitivity of hPSCs to damage caused by erroneous mitosis and increases the risk of gaining aneuploidy.
Highlights
For the safe use of human pluripotent stem cells in regenerative medicine, they should be devoid of mutations that could render them or their differentiated progeny malignant upon transplantation into a patient (Goldring et al, 2011)
Undifferentiated human pluripotent stem cells (hPSCs) Are Highly Sensitive to Abnormal Mitosis-Induced Apoptosis, which Is Independent of TP53 hPSCs have been reported to escape mitotic arrest due to uncoupling of the mitotic checkpoint from apoptosis (Mantel et al, 2007)
To test whether hPSCs undergo apoptosis following perturbation of mitosis, we exposed hPSCs to nocodazole (NOC), which prevents the formation of the mitotic spindle
Summary
For the safe use of human pluripotent stem cells (hPSCs) in regenerative medicine, they should be devoid of mutations that could render them or their differentiated progeny malignant upon transplantation into a patient (Goldring et al, 2011). Most of the hPSC lines have normal diploid karyotype, the incidence of aneuploidy increases significantly with passage number, and gains of the whole or parts of chromosomes 1, 12, 17, and 20 are substantially more common than other changes (Amps et al, 2011; Taapken et al, 2011) Most likely, these genetic changes are selected because they confer a growth advantage (Olariu et al, 2010), which may be attributed to their ability to evade the bottlenecks that restrict the expansion of wild-type cells in culture, including mass cell death following plating, failure to re-enter the cell cycle, and the high death rate of daughter cells in incipient colonies (Barbaric et al, 2014). HPSCs subjected to DNA-replication stress in S phase rapidly commit to apoptosis rather than initiate DNA repair mechanisms (Desmarais et al, 2012)
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