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Turner syndrome (TS) is an X monosomy-related disorder caused by X chromosome nondisjunction during embryonic development. Patients with TS have only one intact X chromosome, with the other either completely or partially lost. TS affects various tissues, including the liver, kidneys, brain, cardiovascular system, and ovaries. These abnormalities are suggested to involve an altered dosage of escape genes that evade X chromosome inactivation. However, the mechanisms and roles of these escape genes in the TS phenotype remain unclear. We hypothesized that the expression levels of escape genes differ between wild-type (WT) and TS cell lines. In this study, we generated induced pluripotent stem cell (iPSC) lines from WT and TS fibroblasts and examined the expression levels of escape genes in both undifferentiated fibroblasts and reprogrammed iPSCs from WT and TS samples. The reprogrammed WT and TS iPSCs exhibited general characteristics of pluripotency, including the expression of pluripotency markers and the potential to differentiate into all three germ layers. Forty-five escape genes were differentially expressed between the WT and TS cell lines. Among these, five genes (ATP7A, PHKA1, EBP, ZFX, and SMC1A) were suggested to be implicated in the TS phenotype. However, further studies using additional cell lines are necessary to clarify the correlation between TS and escape genes. [ABSTRACT FROM AUTHOR]

Brain organoids have been considered as an advanced platform for in vitro disease modeling and drug screening, but numerous roadblocks exist, such as lack of large-scale production technology and lengthy protocols with multiple manipulation steps, impeding the industrial translation of brain organoid technology. Here, we describe the high-speed and large-scale production of midbrain organoids using a high-throughput screening-compatible platform within 30 days. Micro midbrain organoids (µMOs) exhibit a highly uniform morphology and gene expression pattern with minimal variability. Notably, µMOs show dramatically accelerated maturation, resulting in the generation of functional µMOs within only 30 days of differentiation. Furthermore, individual µMOs display highly consistent responsiveness to neurotoxin, suggesting their usefulness as an in vitro high-throughput drug toxicity screening platform. Collectively, our data indicate that µMO technology could represent an advanced and robust platform for in vitro disease modeling and drug screening for human neuronal diseases. [ABSTRACT FROM AUTHOR]

Mitochondria are crucial for cellular energy metabolism and are involved in signaling, aging, and cell death. They undergo dynamic changes through fusion and fission to adapt to different cellular states. In this study, we investigated the effect of knocking out the dynamin 1-like protein (Dnm1l) gene, a key regulator of mitochondrial fission, in neural stem cells (NSCs) differentiated from Dnm1l knockout embryonic stem cells (Dnm1l−/− ESCs). Dnm1l−/− ESC-derived NSCs (Dnm1l−/− NSCs) exhibited similar morphology and NSC marker expression (Sox2, Nestin, and Pax6) to brain-derived NSCs, but lower Nestin and Pax6 expression than both wild-type ESC-derived NSCs (WT-NSCs) and brain-derived NSCs. In addition, compared with WT-NSCs, Dnm1l−/− NSCs exhibited distinct mitochondrial morphology and function, contained more elongated mitochondria, showed reduced mitochondrial respiratory capacity, and showed a metabolic shift toward glycolysis for ATP production. Notably, Dnm1l−/− NSCs exhibited impaired self-renewal ability and accelerated cellular aging during prolonged culture, resulting in decreased proliferation and cell death. Furthermore, Dnm1l−/− NSCs showed elevated levels of inflammation and cell stress markers, suggesting a connection between Dnm1l deficiency and premature aging in NSCs. Therefore, the compromised self-renewal ability and accelerated cellular aging of Dnm1l−/− NSCs may be attributed to mitochondrial fission defects. [ABSTRACT FROM AUTHOR]

Mouse primordial germ cells (PGCs) first appear at embryonic day (E)7.25 on the base of the allantois, and then migrate through the hindgut to the genital ridge (3). During migration [...], Meiosis is a germ-cell-specific cell division process through which haploid gametes are produced for sexual reproduction (1). Before the initiation of meiosis, mouse primordial germ cells undergo a series of epigenetic reprogramming steps (2,3), including the global erasure of DNA methylation at the 5-position of cytosine (5mC) in CpG-rich DNA (4,5). Although several epigenetic regulators, such as Dnmt3l and the histone methyltransferases G9a and Prdm9, have been reported to be crucial for meiosis (6), little is known about how the expression of meiotic genes is regulated and how their expression contributes to normal meiosis. Using a loss-of-function approach in mice, here we show that the 5mC-specific dioxygenase Tet1 has an important role in regulating meiosis in mouse oocytes. Tet1 deficiency significantly reduces female germ-cell numbers and fertility. Univalent chromosomes and unresolved DNA double-strand breaks are also observed in Tet1-deficient oocytes. Tet1 deficiency does not greatly affect the genome-wide demethylation that takes place in primordial germ cells, but leads to defective DNA demethylation and decreased expression of a subset of meiotic genes. Our study thus establishes a function for Tet1 in meiosis and meiotic gene activation in female germ cells.

Author(s): Shinsuke Ito [sup.1] [sup.2] , Ana C. D'Alessio [sup.1] [sup.2] , Olena V. Taranova [sup.1] [sup.2] , Kwonho Hong [sup.1] [sup.2] , Lawrence C. Sowers [sup.3] , Yi Zhang [...], Tet proteins and DNA demethylation The enzymes responsible for methylating DNA, a ubiquitous epigenetic modification, have been extensively studied. Less is known about demethylation. Here, Tet1, an enzyme that catalyses the conversion of 5-methylcytosine of DNA to 5-hydroxyl-methylcytosine, is shown to have a role in mouse embryonic stem-cell maintenance by maintaining the expression of Nanog and preserving the Nanog promoter in a hypomethylated state. This suggests that Tet1 has a role in regulating DNA methylation, as well as in embryonic stem-cell self-renewal and inner cell mass specification. TET1 is an enzyme that catalyses the conversion of 5-methylcytosine of DNA to 5-hydroxymethylcytosine, raising the possibility that it is involved in mediating DNA demethylation. These authors show that Tet1 is involved in mouse embryonic stem cell maintenance and specification of the inner cell mass. It is required to maintain both the expression of Nanog in embryonic stem cells and the Nanog promoter in a hypomethylated state, supporting a role for Tet1 in regulating DNA methylation. DNA methylation is one of the best-characterized epigenetic modifications.sup.1,2,3,4. Although the enzymes that catalyse DNA methylation have been characterized, enzymes responsible for demethylation have been elusive.sup.5. A recent study indicates that the human TET1 protein could catalyse the conversion of 5-methylcytosine (5mC) of DNA to 5-hydroxymethylcytosine (5hmC), raising the possibility that DNA demethylation may be a Tet1-mediated process.sup.6. Here we extend this study by demonstrating that all three mouse Tet proteins (Tet1, Tet2 and Tet3) can also catalyse a similar reaction. Tet1 has an important role in mouse embryonic stem (ES) cell maintenance through maintaining the expression of Nanog in ES cells. Downregulation of Nanog via Tet1 knockdown correlates with methylation of the Nanog promoter, supporting a role for Tet1 in regulating DNA methylation status. Furthermore, knockdown of Tet1 in pre-implantation embryos results in a bias towards trophectoderm differentiation. Thus, our studies not only uncover the enzymatic activity of the Tet proteins, but also demonstrate a role for Tet1 in ES cell maintenance and inner cell mass cell specification.

Precise regulation of the cell cycle of embryonic stem cells (ESCs) is critical for their self-maintenance and differentiation. The cell cycle of ESCs differs from that of somatic cells and is different depending on the cell culture conditions. However, the cell cycle regulation in ESCs via epigenetic mechanisms remains unclear. Here, we showed that the ATP-dependent chromatin remodeler Ino80 regulates the cell cycle genes in ESCs under primed conditions. Ino80 loss led to a significantly extended length of the G1-phase in ESCs grown under primed culture conditions. Ino80 directly bound to the transcription start site and regulated the expression of cell cycle-related genes. Furthermore, Ino80 loss induced cell apoptosis. However, the regulatory mechanism of Ino80 in differentiating ESC cycle slightly differed; an extended S-phase was detected in differentiating inducible Ino80 knockout ESCs. RNA-seq analysis of differentiating ESCs revealed that the expression of genes associated with organ development cell cycle is persistently altered in Ino80 knockout cells, suggesting that cell cycle regulation by Ino80 is not limited to undifferentiated ESCs. Therefore, our study establishes the function of Ino80 in ESC cycle via transcriptional regulation, at least partly. Moreover, this Ino80 function may be universal to other cell types. [ABSTRACT FROM AUTHOR]

In mice, zygotic genome activation (ZGA) occurs in two steps: minor ZGA at the one–cell stage and major ZGA at the two–cell stage. Regarding the regulation of gene transcription, minor ZGA is known to have unique features, including a transcriptionally permissive state of chromatin and insufficient splicing processes. The molecular characteristics may originate from extremely open chromatin states in the one–cell stage zygotes, yet the precise underlying mechanism has not been well studied. Recently, the R-loop, a triple–stranded nucleic acid structure of the DNA/RNA hybrid, has been implicated in gene transcription and DNA replication. Therefore, in the present study, we examined the changes in R-loop dynamics during mouse zygotic development, and its roles in zygotic transcription or DNA replication. Our analysis revealed that R-loops persist in the genome of metaphase II oocytes and preimplantation embryos from the zygote to the blastocyst stage. In particular, zygotic R-loop levels dynamically change as development proceeds, showing that R-loop levels decrease as pronucleus maturation occurs. Mechanistically, R-loop dynamics are likely linked to ZGA, as inhibition of either DNA replication or transcription at the time of minor ZGA decreases R-loop levels in the pronuclei of zygotes. However, the induction of DNA damage by treatment with anticancer agents, including cisplatin or doxorubicin, does not elicit genome-wide changes in zygotic R-loop levels. Therefore, our study suggests that R-loop formation is mechanistically associated with the regulation of mouse ZGA, especially minor ZGA, by modulating gene transcription and DNA replication. [ABSTRACT FROM AUTHOR]

As a novel cell type from eight-cell-stage embryos, extended pluripotent stem cells (EPSCs) are known for diverse differentiation potency in both extraembryonic and embryonic lineages, suggesting new possibilities as a developmental research model. Although various features of EPSCs have been defined, their ability to directly transfer extended pluripotency to differentiated somatic cells by cell fusion remains to be elucidated. Here, we derived EPSCs from eight-cell mouse embryos and confirmed their extended pluripotency at the molecular level and extraembryonic differentiation ability. Then, they were fused with OG2+/− ROSA+/− neural stem cells (NSCs) by the polyethylene-glycol (PEG)-mediated method and further analyzed. The resulting fused hybrid cells exhibited pluripotential markers with upregulated EPSC-specific gene expression. Furthermore, the hybrid cells contributed to the extraembryonic and embryonic lineages in vivo and in vitro. RNA sequencing analysis confirmed that the hybrid cells showed distinct global expression patterns resembling EPSCs without parental expression of NSC markers, indicating the complete acquisition of extended pluripotency and the erasure of the somatic memory of NSCs. Furthermore, ultrastructural observation and metabolic analysis confirmed that the hybrid cells rearranged the mitochondrial morphology and bivalent metabolic profile to those of EPSCs. In conclusion, the extended pluripotency of EPSCs could be transferred to somatic cells through fusion-induced reprogramming. [ABSTRACT FROM AUTHOR]

The dynamics of uterine endometrium is important for successful establishment and maintenance of embryonic implantation and development, along with extensive cell differentiation and proliferation. The tissue event is precisely and complicatedly regulated as several signaling pathways are involved including two main hormones, estrogen and progesterone signaling. We previously showed a novel signaling molecule, Serine/threonine protein kinase 3/4 (STK3/4), which is responded to hormone in the mouse uterine epithelium. However, the role and regulation of its target, YES-associated protein (YAP) remains unknown. In this study, we investigated the expression and regulation of YAP in mouse endometrium. We found that YAP was periodically expressed in the endometrium during the estrous cycle. Furthermore, periodic expression of YAP was shown to be related to the pathway under hormone treatment. Interestingly, estrogen was shown to positively modulate YAP via endometrial epithelial receptors. In addition, the knockdown of YAP showed that YAP regulated various target genes in endometrial cells. The knockdown of YAP down-regulated numerous targets including ADAMTS1, AMOT, AMOTL1, ANKRD1, CTNNA1, MCL1. On the other hand, the expressions of AREG and AXL were increased by its knockdown. These findings imply that YAP responds via Hippo signaling under various intrauterine signals and is considered to play a role in the expression of factors important for uterine endometrium dynamic regulation. [ABSTRACT FROM AUTHOR]

The lymphatic system is critical for maintaining the homeostasis of lipids and interstitial fluid and regulating the immune cell development and functions. Developmental anomaly-induced lymphatic dysfunction is associated with various pathological conditions, including lymphedema, inflammation, and cancer. Most lymphatic endothelial cells (LECs) are derived from a subset of endothelial cells in the cardinal vein. However, recent studies have reported that the developmental origin of LECs is heterogeneous. Multiple regulatory mechanisms, including those mediated by signaling pathways, transcription factors, and epigenetic pathways, are involved in lymphatic development and functions. Recent studies have demonstrated that the epigenetic regulation of transcription is critical for embryonic LEC development and functions. In addition to the chromatin structures, epigenetic modifications may modulate transcriptional signatures during the development or differentiation of LECs. Therefore, the understanding of the epigenetic mechanisms involved in the development and function of the lymphatic system can aid in the management of various congenital or acquired lymphatic disorders. Future studies must determine the role of other epigenetic factors and changes in mammalian lymphatic development and function. Here, the recent findings on key factors involved in the development of the lymphatic system and their epigenetic regulation, LEC origins from different organs, and lymphatic diseases are reviewed. [ABSTRACT FROM AUTHOR]

The uterus is essential for embryo implantation and fetal development. During the estrous cycle, the uterine endometrium undergoes dramatic remodeling to prepare for pregnancy. Angiogenesis is an essential biological process in endometrial remodeling. Steroid hormones regulate the series of events that occur during such remodeling. Researchers have investigated the potential factors, including angiofactors, involved in endometrial remodeling. The Hippo signaling pathway discovered in the 21st century, plays important roles in various cellular functions, including cell proliferation and cell death. However, its role in the endometrium remains unclear. In this review, we describe the female reproductive system and its association with the Hippo signaling pathway, as well as novel Hippo pathway genes and potential target genes. [ABSTRACT FROM AUTHOR]

Luminal breast cancer, an etiologically heterogeneous disease, is characterized by high steroid hormone receptor activity and aberrant gene expression profiles. Endocrine therapy and chemotherapy are promising therapeutic approaches to mitigate breast cancer proliferation and recurrence. However, the treatment of therapy-resistant breast cancer is a major challenge. Recent studies on breast cancer etiology have revealed the critical roles of epigenetic factors in luminal breast cancer tumorigenesis and drug resistance. Tumorigenic epigenetic factor-induced aberrant chromatin dynamics dysregulate the onset of gene expression and consequently promote tumorigenesis and metastasis. Epigenetic dysregulation, a type of somatic mutation, is a high-risk factor for breast cancer progression and therapy resistance. Therefore, epigenetic modulators alone or in combination with other therapies are potential therapeutic agents for breast cancer. Several clinical trials have analyzed the therapeutic efficacy of potential epi-drugs for breast cancer and reported beneficial clinical outcomes, including inhibition of tumor cell adhesion and invasiveness and mitigation of endocrine therapy resistance. This review focuses on recent findings on the mechanisms of epigenetic factors in the progression of luminal breast cancer. Additionally, recent findings on the potential of epigenetic factors as diagnostic biomarkers and therapeutic targets for breast cancer are discussed. [ABSTRACT FROM AUTHOR]

Objectives: The genetic instability and DNA damage arise during transcription factor‐mediated reprogramming of somatic cells, and its efficiency may be reduced due to abnormal chromatin remodelling. The efficiency in somatic cell nuclear transfer (SCNT)‐mediated reprogramming is also very low, and it is caused by development arrest of most reconstituted embryos. Materials and Methods: Whether the repair of genetic instability or double‐strand breaks (DSBs) during SCNT reprogramming may play an important role in embryonic development, we observed and analysed the effect of Rad 51, a key modulator of DNA damage response (DDR) in SCNT‐derived embryos. Results: Here, we observed that the activity of Rad 51 is lower in SCNT eggs than in conventional IVF and found a significantly lower level of DSBs in SCNT embryos during reprogramming. To address this difference, supplementation with RS‐1, an activator of Rad51, during the activation of SCNT embryos can increase RAD51 expression and DSB foci and thereby increased the efficiency of SCNT reprogramming. Through subsequent single‐cell RNA‐seq analysis, we observed the reactivation of a large number of genes that were not expressed in SCNT‐2‐cell embryos by the upregulation of DDR, which may be related to overcoming the developmental block. Additionally, there may be an independent pathway involving histone demethylase that can reduce reprograming‐resistance regions. Conclusions: This technology can contribute to the production of comparable cell sources for regenerative medicine. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMP) are innate immune molecules with antimicrobial activity that are important to defend against pathogenic infection. We performed genome‐level analyses of the AMP family gene repertories for six terrestrial (Homo sapiens, Bos taurus, Sus scrofa, Capra hircus, Canis lupus, and Loxodonta africana) and five aquatic (Balaenoptera acutorostrata, Tursiops truncatus, Lipotes vexillifer, Odobenus rosmarus, and Trichechus manatus) mammals and identified 59 (30 partial or pseudogenes) previously unannotated β‐defensin (BD) sequences from those aquatic species. Extreme reduction in BD genes was evident in Odontoceti. NK‐lysin and granulysin among AMP family genes were absent from aquatic mammals, except O. rosmarus. A phylogenetic tree constructed with 296 amino acid sequences of BD from 11 species suggested a broad expansion of this gene family with extensive genetic diversity in mammals. We determined a pair of evolutionarily conserved flanking genes for each of four BD containing intervals and defined them as PGK2‐TFAP2D, BCL2L1‐ZCCHC3, AGPAT5‐GPM6A, and ADAM29‐CTSB intervals. The characterization of BD syntenic regions revealed conservation of homologous synteny blocks and reuse of evolutionary breakpoint regions. Phylogenetic reconstruction of BD gene evolution showed that the major expansion events of BD genes occurred four times in the mammalian lineage. Taken together, the findings of this study show the occurrence of significant differences in the number of AMP family genes in diverse mammalian species, including aquatic species, likely due to their environmental adaptation. [ABSTRACT FROM AUTHOR]

During embryonic development, cells undergo changes in gene expression, signaling pathway activation/inactivation, metabolism, and intracellular organelle structures, which are mediated by mitochondria. Mitochondria continuously switch their morphology between elongated tubular and fragmented globular via mitochondrial fusion and fission. Mitochondrial fusion is mediated by proteins encoded by Mfn1, Mfn2, and Opa1, whereas mitochondrial fission is mediated by proteins encoded by Fis1 and Dnm1L. Here, we investigated the expression patterns of mitochondria-related genes during the differentiation of mouse embryonic stem cells (ESCs). Pluripotent ESCs maintain stemness in the presence of leukemia inhibitory factor (LIF) via the JAK-STAT3 pathway but lose pluripotency and differentiate in response to the withdrawal of LIF. We analyzed the expression levels of mitochondrial fusion- and fission-related genes during the differentiation of ESCs. We hypothesized that mitochondrial fusion genes would be overexpressed while the fission genes would be downregulated during the differentiation of ESCs. Though the mitochondria exhibited an elongated morphology in ESCs differentiating in response to LIF withdrawal, only the expression of Mfn2 was increased and that of Dnm1L was decreased as expected, the other exceptions being Mfn1, Opa1, and Fis1. Next, by comparing gene expression and mitochondrial morphology, we proposed an index that could precisely represent mitochondrial changes during the differentiation of pluripotent stem cells by analyzing the expression ratios of three fusion- and two fission-related genes. Surprisingly, increased Mfn2/Dnm1L ratio was correlated with elongation of mitochondria during the differentiation of ESCs. Moreover, application of this index to other specialized cell types revealed that neural stems cells (NSCs) and mouse embryonic fibroblasts (MEFs) showed increased Mfn2/Dnm1L ratio compared to ESCs. Thus, we suggest that the Mfn2/Dnm1L ratio could reflect changes in mitochondrial morphology according to the extent of differentiation. [ABSTRACT FROM AUTHOR]

Human‐induced pluripotent stem cells (hiPSCs) have facilitated studies on organ development and differentiation into specific lineages in in vitro systems. Although numerous studies have focused on cellular differentiation into neural lineage using hPSCs, most studies have initially evaluated embryoid body (EB) formation, eventually yielding terminally differentiated neurons with limited proliferation potential. This study aimed to establish human primitive neural stem cells (pNSCs) from exogene‐free hiPSCs without EB formation. To derive pNSCs, we optimized N2B27 neural differentiation medium through supplementation of two inhibitors, CHIR99021 (GSK‐3 inhibitor) and PD0325901 (MEK inhibitor), and growth factors including basic fibroblast growth factor (bFGF) and human leukemia inhibitory factor (hLIF). Consequently, pNSCs were efficiently derived and cultured over a long term. pNSCs displayed differentiation potential into neurons, astrocytes, and oligodendrocytes. These early NSC types potentially promote the clinical application of hiPSCs to cure human neurological disorders. [ABSTRACT FROM AUTHOR]

Neuronal expression of beta‐secretase 1 (BACE1) has been implicated in the progression of Alzheimer's disease. However, the mechanisms that regulate BACE1 expression are unclear. Here, we show that peroxisome proliferator‐activated receptor delta (PPARδ) decreases BACE1 expression by up‐regulating suppressor of cytokine signaling 1 (SOCS1) in SH‐SY5Y neuroblastoma cells. The activation of PPARδ by GW501516, a specific PPARδ agonist, inhibited expression of BACE1. This effect was abrogated by shRNA‐mediated knockdown of PPARδ and by treatment with the PPARδ antagonist GSK0660, indicating that PPARδ is involved in GW501516‐mediated suppression of BACE1 expression. On the other hand, GW501516‐activated PPARδ induced expression of SOCS1, which is a negative regulator of cytokine signal transduction, at the transcriptional level by binding to a PPAR response element in its promoter. This GW501516‐mediated induction of SOCS1 expression led to down‐regulation of BACE1 expression via inactivation of signal transducer and activator of transcription 1. GW501516‐activated PPARδ suppressed the generation of neurotoxic amyloid beta (Aβ) in accordance with the decrease in BACE1 expression. Taken together, these results indicate that PPARδ attenuates BACE1 expression via SOCS1‐mediated inhibition of signal transducer and activator of transcription 1 signaling, thereby suppressing BACE1‐associated generation of neurotoxic Aβ. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are active regulators of numerous biological and physiological processes including most of the events of mammalian reproduction. Understanding the biological functions of miRNAs in the context of mammalian reproduction will allow a better and comparative understanding of fertility and sterility in male and female mammals. Herein, we summarize recent progress in miRNA‐mediated regulation of mammalian reproduction and highlight the significance of miRNAs in different aspects of mammalian reproduction including the biogenesis of germ cells, the functionality of reproductive organs, and the development of early embryos. Furthermore, we focus on the gene expression regulatory feedback loops involving hormones and miRNA expression to increase our understanding of germ cell commitment and the functioning of reproductive organs. Finally, we discuss the influence of miRNAs on male and female reproductive failure, and provide perspectives for future studies on this topic. [ABSTRACT FROM AUTHOR]

N6-methyladenosine (m6A) modification in RNA has been implicated in diverse biological processes including the maintenance of embryonic stem cells, early development and diseases. Although the m6A modification was discovered several decades ago, its biological function remained unclear. The recent discovery of enzymes responsible for 'writing' or 'erasing' the modification and single-nucleotide resolution mapping by next-generation sequencing technology have revealed its function in biological processes. Its enrichment pattern is conserved in mammalian transcriptomes, and the level of m6A is tightly regulated by methyltransferases (writers), demethylases (erasers) and binding proteins (readers). Furthermore, accumulating evidence suggests that the aberrant regulation of m6A turnover is associated with multiple types of cancer including acute myeloid leukemia, breast cancer, glioblastoma, lung cancer and liver cancer. Studies have demonstrated that factors involved in m6A metabolism serve either oncogenic or tumor-suppressor roles in different contexts. The previous studies of the role of m6A in cancer biology are discussed in the present review. [ABSTRACT FROM AUTHOR]

Pluripotential reprogramming has been examined using various technologies, including nuclear transfer, cell fusion, and direct reprogramming. Many studies have used differentiated cells for reprogramming experiments, and nearly all type of somatic cells can acquire pluripotency. However, within the embryo, other cells types are present in addition to somatic cells. The blastocyst stage embryo consists of two main types of cells, inner cell mass and trophectoderm (TE). TE cells are the first differentiated form of the totipotent zygote and differ from epiblast cells. Thus, we examined whether extraembryonic cells can be reprogrammed using a cell–cell fusion method. Trophoblast stem cells (TSCs), which can be obtained from the TE, are known to acquire pluripotency by transcription factor Oct4 overexpression or somatic cell nuclear transfer. In this study, we demonstrated that TSCs can acquire pluripotent properties by cell fusion with embryonic stem cells (ESCs). TSC-ESC hybrids reactivated Oct4-GFP and displayed self-renewal properties. They expressed the pluripotency markers Oct4 and Nanog, whereas the expression of Cdx2 and Tead4, trophoblast lineage markers, was diminished. Moreover, these cells developed into three germ layers similarly to other pluripotent stem cells. RNA-seq analysis showed that global gene expression patterns of TSC-ESC hybrids are more similar to ESCs than TSCs. Thus, we demonstrated that TSCs successfully complete reprogramming and acquire pluripotency by cell fusion-induced reprogramming. [ABSTRACT FROM AUTHOR]