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Mga safeguards embryonic stem cells from acquiring extraembryonic endoderm fates.

ABSTRACT: Polycomb group (PcG) proteins form multiprotein complexes that affect stem cell identity and fate decisions by still largely unexplored mechanisms. Here, by performing a CRISPR-based loss-of-function screen in embryonic stem cells (ESCs), we identify PcG gene Mga involved in the repression of endodermal transcription factor Gata6 We report that deletion of Mga results in peri-implantation embryonic lethality in mice. We further demonstrate that Mga-null ESCs exhibit impaired self-renewal and spontaneous differentiation to primitive endoderm (PE). Our data support a model in which Mga might serve as a scaffold for PRC1.6 assembly and guide this multimeric complex to specific genomic targets including genes that encode endodermal factors Gata4, Gata6, and Sox17. Our findings uncover an unexpected function of Mga in ESCs, where it functions as a gatekeeper to prevent ESCs from entering into the PE lineage by directly repressing expression of a set of endoderm differentiation master genes.

SUBMITTER: Qin J

PROVIDER: S-EPMC7821913 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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Mga safeguards embryonic stem cells from acquiring extraembryonic endoderm fates.

Qin Jinzhong J Wang Congcong C Zhu Yaru Y Su Ting T Dong Lixia L Huang Yikai Y Hao Kunying K

Science advances 20210120 4

Polycomb group (PcG) proteins form multiprotein complexes that affect stem cell identity and fate decisions by still largely unexplored mechanisms. Here, by performing a CRISPR-based loss-of-function screen in embryonic stem cells (ESCs), we identify PcG gene Mga involved in the repression of endodermal transcription factor Gata6 We report that deletion of Mga results in peri-implantation embryonic lethality in mice. We further demonstrate that Mga-null ESCs exhibit i ...[more]

PMID: 33523934

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Project description:Initial specification of cardiomyocytes in the mouse results from interactions between the extraembryonic anterior visceral endoderm (AVE) and the nascent mesoderm. However the mechanism by which AVE activates cardiogenesis is not well understood, and the identity of specific cardiogenic factors in the endoderm remains elusive. Most mammalian studies of the cardiogenic potential of the endoderm have relied on the use of cell lines that are similar to the heart-inducing AVE. These include the embryonal-carcinoma-derived cell lines, END2 and PYS2. The recent development of protocols to isolate eXtraembryonic ENdoderm (XEN) stem cells, representing the extraembryonic endoderm lineage, from blastocyst stage mouse embryos offers new tools for the genetic dissection of cardiogenesis.Here, we demonstrate that XEN cell-conditioned media (CM) enhances cardiogenesis during Embryoid Body (EB) differentiation of mouse embryonic stem (ES) cells in a manner comparable to PYS2-CM and END2-CM. Addition of CM from each of these three cell lines enhanced the percentage of EBs that formed beating areas, but ultimately, only XEN-CM and PYS2-CM increased the total number of cardiomyocytes that formed. Furthermore, our observations revealed that both contact-independent and contact-dependent factors are required to mediate the full cardiogenic potential of the endoderm. Finally, we used gene array comparison to identify factors in these cell lines that could mediate their cardiogenic potential.These studies represent the first step in the use of XEN cells as a molecular genetic tool to study cardiomyocyte differentiation. Not only are XEN cells functionally similar to the heart-inducing AVE, but also can be used for the genetic dissection of the cardiogenic potential of AVE, since they can be isolated from both wild type and mutant blastocysts. These studies further demonstrate the importance of both contact-dependent and contact-independent factors in cardiogenesis and identify potential heart-inducing proteins in the endoderm.

Project description:BACKGROUND:The development of somatic reprogramming, especially purely chemical reprogramming, has significantly advanced biological research. And chemical-induced extraembryonic endoderm-like (ciXEN) cells have been confirmed to be an indispensable intermediate stage of chemical reprogramming. They resemble extraembryonic endoderm (XEN) cells in terms of transcriptome, reprogramming potential, and developmental ability in vivo. However, the other characteristics of ciXEN cells and the effects of chemicals and bFGF on the in vitro culture of ciXEN cells have not been systematically reported. METHODS:Chemicals and bFGF in combination with Matrigel were used to induce the generation of ciXEN cells derived from mouse embryonic fibroblasts (MEFs). RNA sequencing was utilised to examine the transcriptome of ciXEN cells, and PCR/qPCR assays were performed to evaluate the mRNA levels of the genes involved in this study. Hepatic functions were investigated by periodic acid-Schiff staining and indocyanine green assay. Lactate production, ATP detection, and extracellular metabolic flux analysis were used to analyse the energy metabolism of ciXEN cells. RESULTS:ciXEN cells expressed XEN-related genes, exhibited high proliferative capacity, had the ability to differentiate into visceral endoderm in vitro, and possessed the plasticity allowing for their differentiation into induced hepatocytes (iHeps). Additionally, the upregulated biological processes of ciXEN cells compared to those in MEFs focused on metabolism, but their energy production was independent of glycolysis. Furthermore, without the cocktail of chemicals and bFGF, which are indispensable for the generation of ciXEN cells, induced XEN (iXEN) cells remained the expression of XEN markers, the high proliferative capacity, and the plasticity to differentiate into iHeps in vitro. CONCLUSIONS:ciXEN cells had high plasticity, and energy metabolism was reconstructed during chemical reprogramming, but it did not change from aerobic oxidation to glycolysis. And the cocktail of chemicals and bFGF were non-essential for the in vitro culture of ciXEN cells.

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Mga safeguards embryonic stem cells from acquiring extraembryonic endoderm fates.

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Mga safeguards embryonic stem cells from acquiring extraembryonic endoderm fates.

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