Mapping Gene Circuits Essential for Germ Layer Differentiation via Loss-of-Function Screens in Haploid Human Embryonic Stem Cells

Yilmaz, Atilgan; Braverman-Gross, Carmel; Bialer-Tsypin, Anna; Peretz, Mordecai; Benvenisty, Nissim

doi:10.1016/j.stem.2020.06.023

Cited by 28 publications

(32 citation statements)

References 54 publications

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“…We examined the expression of neuroectoderm-associated essential transcription factors (TFs) between hPSCs and CLNSCs ( Figure 4 C). The specific germ layer TFs selected have been described before [ 49 ]. The neural development transcription factors such as POU3F2 and POU3F3 were significantly up-regulated.…”

Section: Resultsmentioning

confidence: 99%

Global Transcriptional Analyses of the Wnt-Induced Development of Neural Stem Cells from Human Pluripotent Stem Cells

Liu

Gao

et al. 2021

IJMS

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The differentiation of human pluripotent stem cells (hPSCs) to neural stem cells (NSCs) is the key initial event in neurogenesis and is thought to be dependent on the family of Wnt growth factors, their receptors and signaling proteins. The delineation of the transcriptional pathways that mediate Wnt-induced hPSCs to NSCs differentiation is vital for understanding the global genomic mechanisms of the development of NSCs and, potentially, the creation of new protocols in regenerative medicine. To understand the genomic mechanism of Wnt signaling during NSCs development, we treated hPSCs with Wnt activator (CHIR-99021) and leukemia inhibitory factor (LIF) in a chemically defined medium (N2B27) to induce NSCs, referred to as CLNSCs. The CLNSCs were subcultured for more than 40 passages in vitro; were positive for AP staining; expressed neural progenitor markers such as NESTIN, PAX6, SOX2, and SOX1; and were able to differentiate into three neural lineage cells: neurons, astrocytes, and oligodendrocytes in vitro. Our transcriptome analyses revealed that the Wnt and Hedgehog signaling pathways regulate hPSCs cell fate decisions for neural lineages and maintain the self-renewal of CLNSCs. One interesting network could be the deregulation of the Wnt/β-catenin signaling pathway in CLNSCs via the downregulation of c-MYC, which may promote exit from pluripotency and neural differentiation. The Wnt-induced spinal markers HOXA1-4, HOXA7, HOXB1-4, and HOXC4 were increased, however, the brain markers FOXG1 and OTX2, were absent in the CLNSCs, indicating that CLNSCs have partial spinal cord properties. Finally, a CLNSC simple culture condition, when applied to hPSCs, supports the generation of NSCs, and provides a new and efficient cell model with which to untangle the mechanisms during neurogenesis.

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Section: Resultsmentioning

confidence: 99%

Global Transcriptional Analyses of the Wnt-Induced Development of Neural Stem Cells from Human Pluripotent Stem Cells

Liu

Gao

et al. 2021

IJMS

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“…To access the pluripotency by in vitro differentiation, the cell aggregation via EB formation represented an ability of in vitro three-germ layer development. Expressions of germ layer-specific genes increased simultaneously with the loss of pluripotency and fate transition to differentiation stage (Yilmaz et al, 2020). From qRT-PCR results, EBs derived from compact cell type had highly expressed the three germ layers gene markers.…”

Section: Discussionmentioning

confidence: 96%

Molecular signature and colony morphology affect in vitro pluripotency of porcine induced pluripotent stem cells

Setthawong

Phakdeedindan

Techakumphu

et al. 2021

Reprod Domestic Animals

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Pig is a potential animal model for biomedical research as the organ size, physiology and immunology of pigs are resemblance to humans (Ye et al., 2014). Moreover, long lifespan of pigs has advantage for long-term study of human diseases following cell transplantation and experimental surgery (Ezashi et al., 2016;Shiue et al., 2016).The genetic modifications via cell reprogramming technology have also held a great promise to improve the efficiency of regenerative medicine and pig production (Gutierrez et al., 2015). However,

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“…S4A; see table S2G for functional annotation). Given the critical role of trafficking in cell polarity (41) and in pluripotency exit (42), the polarized membrane proteins involved in membrane traffic (e.g., SNX27 and AP1G1) in 3D hPSC systems are of particular interest for further investigation.…”

Section: Cell Polarity Proteomes Reveal New Apically and Basolaterally Polarized Proteinsmentioning

confidence: 99%

Spatially resolved cell polarity proteomics of a human epiblast model

et al. 2021

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Critical early steps in human embryonic development include polarization of the inner cell mass, followed by formation of an expanded lumen that will become the epiblast cavity. Recently described three-dimensional (3D) human pluripotent stem cell–derived cyst (hPSC-cyst) structures can replicate these processes. To gain mechanistic insights into the poorly understood machinery involved in epiblast cavity formation, we interrogated the proteomes of apical and basolateral membrane territories in 3D human hPSC-cysts. APEX2-based proximity bioinylation, followed by quantitative mass spectrometry, revealed a variety of proteins without previous annotation to specific membrane subdomains. Functional experiments validated the requirement for several apically enriched proteins in cyst morphogenesis. In particular, we found a key role for the AP-1 clathrin adaptor complex in expanding the apical membrane domains during lumen establishment. These findings highlight the robust power of this proximity labeling approach for discovering novel regulators of epithelial morphogenesis in 3D stem cell–based models.

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Mapping Gene Circuits Essential for Germ Layer Differentiation via Loss-of-Function Screens in Haploid Human Embryonic Stem Cells

Cited by 28 publications

References 54 publications

Global Transcriptional Analyses of the Wnt-Induced Development of Neural Stem Cells from Human Pluripotent Stem Cells

Global Transcriptional Analyses of the Wnt-Induced Development of Neural Stem Cells from Human Pluripotent Stem Cells

Molecular signature and colony morphology affect in vitro pluripotency of porcine induced pluripotent stem cells

Spatially resolved cell polarity proteomics of a human epiblast model

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