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Loss of Tet enzymes compromises proper differentiation of embryonic stem cells

Meelad M Dawlaty1, Achim Breiling2, Thuc Le3

  • 1Whitehead Institute for Biomedical Research, Cambridge, MA 02142, USA.

Developmental Cell
|April 17, 2014
PubMed

Insights

Complete loss of Tet enzymes (Tet1/2/3) impairs mouse embryonic stem cell differentiation and development by depleting 5-hydroxymethylcytosine (5hmC) marks. This highlights the critical role of Tet enzymes and 5hmC in regulating gene expression during development.

Area of Science:

  • Epigenetics and Developmental Biology
  • DNA demethylation mechanisms
  • Mammalian embryogenesis

Background:

  • Tet enzymes (Tet1/2/3) catalyze the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC).
  • While individual or dual Tet deficiencies permit embryogenesis, the impact of complete Tet activity loss on development remains unknown.
  • 5hmC is dynamically regulated during development and its precise role in the absence of Tet activity requires investigation.

Purpose of the Study:

  • To investigate the developmental potential of mouse embryonic stem cells (ESCs) lacking all three Tet enzymes (Tet1/2/3 triple-knockout, TKO).
  • To determine the effect of complete Tet deficiency and absence of 5hmC on ESC differentiation and embryonic development.
  • To elucidate the molecular mechanisms underlying developmental defects in TKO ESCs.

Main Methods:

  • Generation and characterization of Tet1/2/3 TKO mouse ESCs.
  • Assessment of ESC differentiation potential using embryoid bodies (EBs) and teratoma formation.
  • Analysis of chimeric embryo development and contribution of TKO ESCs.
  • Global gene expression and whole-genome bisulfite sequencing (WGBS) for methylome analysis of TKO EBs.

Main Results:

  • TKO ESCs exhibited significantly depleted 5hmC levels.
  • TKO embryoid bodies and teratomas showed impaired differentiation.
  • TKO ESCs contributed poorly to chimeric embryos and could not support embryonic development, a defect rescued by Tet1 reexpression.
  • Global gene expression and methylome analyses revealed promoter hypermethylation and dysregulation of key developmental genes in TKO EBs.

Conclusions:

  • Complete loss of Tet enzyme activity and 5hmC impairs mouse ESC differentiation and embryonic development.
  • Tet- and 5hmC-mediated DNA demethylation is essential for the proper regulation of gene expression during ESC differentiation.
  • These findings underscore the critical role of the Tet-5hmC pathway in mammalian development.

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