Regulation of in vitro and in vivo differentiation of mouse embryonic stem cells, embryonic germ cells and teratocarcinoma cells by TGFβ family signaling factors

The DiGeorge syndrome critical region gene 8 (Dgcr8) knockout strategy has been widely used to study the function of canonical microRNAs (miRNAs) in vitro and in vivo. However, primary miRNA (pri-miRNA) transcripts are accumulated in Dgcr8 knockout cells due to interrupted processing. Whether abnormally accumulated pri-miRNAs have any function is unknown. Here, using clustered regularly interspaced short palindromic repeats system/CRISPR-associated protein 9 (CRISPR/Cas9), we successfully knocked out the primary microRNA-290~295 (pri-miR-290~295) cluster, the most highly expressed miRNA cluster in mouse embryonic stem cells (ESCs), in Dgcr8 knockout background. We found that the major defects associated with Dgcr8 knockout in mouse ESCs, including higher expression of epithelial-to-mesenchymal transition (EMT) markers, slower proliferation, G1 accumulation, and defects in silencing self-renewal, were not affected by the deletion of pri-miR-290~290 cluster. Interestingly, the transcription of neighboring gene nucleotide-binding oligomerization domain, leucine rich repeat and pyrin domain containing 12(Nlrp12) was upregulated upon the deletion of the pri-miR-290~295 cluster. Together, our results suggested that the major defects in Dgcr8 knockout ESCs were not due to the accumulation of pri-miR-290~295, and the deletion of miRNA genes could affect the transcription of neighboring DNA elements.

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SIRT1 regulates sphingolipid metabolism and neural differentiation of mouse embryonic stem cells through c-Myc- SMPDL3B

10.1101/2021.02.24.432815 ◽

2021 ◽

Author(s):

Wei Fan ◽

Shuang Tang ◽

Xiaojuan Fan ◽

Yi Fang ◽

Xiaojiang Xu ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Neural Differentiation ◽

Transcriptional Control ◽

Molecular Mechanisms ◽

Embryonic Stem ◽

Mouse Embryonic Stem Cells ◽

Sphingolipid Metabolism

AbstractSphingolipids are important structural components of cell membranes and prominent signaling molecules controlling cell growth, differentiation, and apoptosis. Sphingolipids are particularly abundant in the brain, and defects in sphingolipid degradation are associated with several human neurodegenerative diseases. However, molecular mechanisms governing sphingolipid metabolism remain unclear. Here we report that sphingolipid degradation is under transcriptional control of SIRT1, a highly conserved mammalian NAD+-dependent protein deacetylase, in mouse embryonic stem cells (mESCs). Deletion of SIRT1 results in accumulation of sphingomyelin in mESCs, primarily due to reduction of SMPDL3B, a GPI-anchored plasma membrane bound sphingomyelin phosphodiesterase. Mechanistically, SIRT1 regulates transcription of Smpdl3b through c-Myc. Functionally, SIRT1 deficiency-induced accumulation of sphingomyelin increases membrane fluidity and impairs neural differentiation in vitro and in vivo. Our findings discover a key regulatory mechanism for sphingolipid homeostasis and neural differentiation, further imply that pharmacological manipulation of SIRT1-mediated sphingomyelin degradation might be beneficial for treatment of human neurological diseases.

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Mouse Germ Cell Development in-vivo and in-vitro

Biomarker Insights ◽

10.1177/117727190700200024 ◽

2007 ◽

Vol 2 ◽

pp. 117727190700200 ◽

Cited By ~ 15

Author(s):

Deshira Saiti ◽

Orly Lacham-Kaplan

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Germ Cell ◽

Germ Cells ◽

Expression Patterns ◽

Embryonic Stem ◽

Cell Lineage ◽

Initial Population

In mammalian development, primordial germ cells (PGCs) represent the initial population of cells that are committed to the germ cell lineage. PGCs segregate early in development, triggered by signals from the extra-embryonic ectoderm. They are distinguished from surrounding cells by their unique gene expression patterns. Some of the more common genes used to identify them are Blimp1, Oct3/4, Fragilis, Stella, c-Kit, Mvh, Dazl and Gcna1. These genes are involved in regulating their migration and differentiation, and in maintaining the pluripotency of these cells. Recent research has demonstrated the possibility of obtaining PGCs, and subsequently, mature germ cells from a starting population of embryonic stem cells (ESCs) in culture. This phenomenon has been investigated using a variety of methods, and ESC lines of both mouse and human origin. Embryonic stem cells can differentiate into germ cells of both the male and female phenotype and in one case has resulted in the birth of live pups from the fertilization of oocytes with ESC derived sperm. This finding leads to the prospect of using ESC derived germ cells as a treatment for sterility. This review outlines the evolvement of germ cells from ESCs in vitro in relation to in vivo events.

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Glutamatergic Neuronal Differentiation of Mouse Embryonic Stem Cells after Transient Expression of Neurogenin 1 and Treatment with BDNF and GDNF: In Vitro and In Vivo Studies

Journal of Neuroscience ◽

10.1523/jneurosci.0563-08.2008 ◽

2008 ◽

Vol 28 (48) ◽

pp. 12622-12631 ◽

Cited By ~ 83

Author(s):

J. H. Reyes ◽

K. S. O'Shea ◽

N. L. Wys ◽

J. M. Velkey ◽

D. M. Prieskorn ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Neuronal Differentiation ◽

Transient Expression ◽

Embryonic Stem ◽

Mouse Embryonic Stem Cells ◽

In Vivo Studies

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SIRT1 regulates sphingolipid metabolism and neural differentiation of mouse embryonic stem cells through c-Myc- SMPDL3B

eLife ◽

10.7554/elife.67452 ◽

2021 ◽

Vol 10 ◽

Author(s):

Wei Fan ◽

Shuang Tang ◽

Xiaojuan Fan ◽

Yi Fang ◽

Xiaojiang Xu ◽

...

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Neural Differentiation ◽

Transcriptional Control ◽

Molecular Mechanisms ◽

Embryonic Stem ◽

Mouse Embryonic Stem Cells ◽

Sphingolipid Metabolism

Sphingolipids are important structural components of cell membranes and prominent signaling molecules controlling cell growth, differentiation, and apoptosis. Sphingolipids are particularly abundant in the brain, and defects in sphingolipid degradation are associated with several human neurodegenerative diseases. However, molecular mechanisms governing sphingolipid metabolism remain unclear. Here we report that sphingolipid degradation is under transcriptional control of SIRT1, a highly conserved mammalian NAD+-dependent protein deacetylase, in mouse embryonic stem cells (mESCs). Deletion of SIRT1 results in accumulation of sphingomyelin in mESCs, primarily due to reduction of SMPDL3B, a GPI-anchored plasma membrane bound sphingomyelin phosphodiesterase. Mechanistically, SIRT1 regulates transcription of Smpdl3b through c-Myc. Functionally, SIRT1 deficiency-induced accumulation of sphingomyelin increases membrane fluidity and impairs neural differentiation in vitro and in vivo. Our findings discover a key regulatory mechanism for sphingolipid homeostasis and neural differentiation, further imply that pharmacological manipulation of SIRT1-mediated sphingomyelin degradation might be beneficial for treatment of human neurological diseases.

Download Full-text