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Platelet transfusions are critical for severe thrombocytopenia but depend on blood donors. The shortage of donors and the potential of universal HLA-null platelet products have stimulated research on the ex vivo differentiation of human pluripotent stem cells (hPSCs) to platelets. We recently established expandable immortalized megakaryocyte cell lines (imMKCLs) from hPSCs by transducing MYC, BMI1, and BCL-XL (MBX). imMKCLs can act as cryopreservable master cells to supply platelet concentrates. However, the proliferation rates of the imMKCLs vary with the starting hPSC clone. In this study, we reveal from the gene expression profiles of several MKCL clones that the proliferation arrest is correlated with the expression levels of specific cyclin-dependent kinase inhibitors. Silencing CDKN1A and p53 with the overexpression of MBX was effective at stably inducing imMKCLs that generate functional platelets irrespective of the hPSC clone. Collectively, this improvement in generating imMKCLs should contribute to platelet industrialization and platelet biology.

Mammalian hibernators endure severe and prolonged hypothermia that is lethal to non-hibernators, including humans and mice. The mechanisms responsible for the cold resistance remain poorly understood. Here, we found that hepatocytes from a mammalian hibernator, the Syrian hamster, exhibited remarkable resistance to prolonged cold culture, whereas murine hepatocytes underwent cold-induced cell death that fulfills the hallmarks of ferroptosis such as necrotic morphology, lipid peroxidation and prevention by an iron chelator. Unexpectedly, hepatocytes from Syrian hamsters exerted resistance to cold- and drug-induced ferroptosis in a diet-dependent manner, with the aid of their superior ability to retain dietary alpha -tocopherol (alpha T), a vitamin E analog, in the liver and blood compared with those of mice. The liver phospholipid composition is less susceptible to peroxidation in Syrian hamsters than in mice. Altogether, the cold resistance of the hibernator's liver is established by the ability to utilize alpha T effectively to prevent lipid peroxidation and ferroptosis. Daisuke Anegawa et al. investigated the mechanisms responsible for cold resistance in the Syrian hamster's hepatocytes, which exhibited remarkable resistance to prolonged cold culture. Their results suggest that hepatocytes exhibit diet-dependent resistance to cold, which is linked to the retention of alpha -tocopherol in the liver.

Platelet transfusions are critical for severe thrombocytopenia but depend on blood donors. The shortage of donors and the potential of universal HLA-null platelet products have stimulated research on the ex vivo differentiation of human pluripotent stem cells (hPSCs) to platelets. We recently established expandable immortalized megakaryocyte cell lines (imMKCLs) from hPSCs by transducing MYC, BMI1, and BCL-XL (MBX). imMKCLs can act as cryopreservable master cells to supply platelet concentrates. However, the proliferation rates of the imMKCLs vary with the starting hPSC clone. In this study, we reveal from the gene expression profiles of several MKCL clones that the proliferation arrest is correlated with the expression levels of specific cyclin-dependent kinase inhibitors. Silencing CDKN1A and p53 with the overexpression of MBX was effective at stably inducing imMKCLs that generate functional platelets irrespective of the hPSC clone. Collectively, this improvement in generating imMKCLs should contribute to platelet industrialization and platelet biology.

Clear cell sarcoma (CCS) is a rare soft tissue sarcoma caused by the EWS/ATF1 fusion gene. Here, we established induced pluripotent stem cells (iPSCs) from EWS/ATF1-controllable murine CCS cells harboring sarcoma-associated genetic abnormalities. Sarcoma-iPSC mice develop secondary sarcomas immediately after EWS/ATF1 induction, but only in soft tissue. EWS/ATF1 expression induces oncogene-induced senescence in most cell types in sarcoma-iPSC mice but prevents it in sarcoma cells. We identify Tppp3-expressing cells in peripheral nerves as a cell-of-origin for these sarcomas. We show cell type-specific recruitment of EWS/ATF1 to enhancer regions in CCS cells. Finally, epigenetic silencing at these enhancers induces senescence and inhibits CCS cell growth through altered EWS/ATF1 binding. Together, we propose that distinct responses to premature senescence are the basis for the cell type-specificity of cancer development.

Higher-order chromatin organization controls transcriptional programs that govern cell properties and functions. In order for pluripotent stem cells (PSCs) to appropriately respond to differentiation signals, developmental gene loci should be structurally and spatially regulated to be readily available for immediate transcription, even though these genes are hardly expressed in PSCs. Here, we show that both chromatin interaction profiles and nuclear positions at developmental gene loci differ between human somatic cells and hPSCs, and that changes in the chromatin interactions are closely related to the nuclear repositioning. Moreover, we also demonstrate that developmental gene loci, which have bivalent histone modifications, tend to colocalize in PSCs. Furthermore, this colocalization requires PRC1, PRC2, and TrxG complexes, which are essential regulatory factors for the maintenance of transcriptionally poised developmental genes. Our results indicate that higher-order chromatin regulation may be an integral part of the differentiation capacity that defines pluripotency.

The role of secreted molecules in cellular reprogramming has been poorly understood. Here we identify a truncated form of ephrin receptor A7 (EPHA7) as a key regulator of reprogramming. Truncated EPHA7 is prominently upregulated and secreted during reprogramming. EPHA7 expression is directly regulated by OCT3/4. EphA7 knockdown results in marked reduction of reprogramming efficiency, and the addition of truncated EPHA7 is able to restore it. ERK activity is markedly reduced during reprogramming, and the secreted, truncated EPHA7 is responsible for ERK activity reduction. Remarkably, treatment of EphA7-knockdown MEFs with the ERK pathway inhibitor restores reprogramming efficiency. Analyses show that truncated EPHA7-induced ERK activity reduction plays an important role in the middle phase of reprogramming. Thus, our findings uncover the importance of secreted EPHA7-induced ERK activity reduction in reprogramming.