Destabilization of pluripotency in the absence of Mad2l2

Cell Cycle. 2015;14(10):1596-610. doi: 10.1080/15384101.2015.1026485.

Abstract

The induction and maintenance of pluripotency requires the expression of several core factors at appropriate levels (Oct4, Sox2, Klf4, Prdm14). A subset of these proteins (Oct4, Sox2, Prdm14) also plays crucial roles for the establishment of primordial germ cells (PGCs). Here we demonstrate that the Mad2l2 (MAD2B, Rev7) gene product is not only required by PGCs, but also by pluripotent embryonic stem cells (ESCs), depending on the growth conditions. Mad2l2(-/-) ESCs were unstable in LIF/serum medium, and differentiated into primitive endoderm. However, they could be stably propagated using small molecule inhibitors of MAPK signaling. Several components of the MAPK cascade were up- or downregulated even in undifferentiated Mad2l2(-/-) ESCs. Global levels of repressive histone H3 variants were increased in mutant ESCs, and the epigenetic signatures on pluripotency-, primitive endoderm-, and MAPK-related loci differed. Thus, H3K9me2 repressed the Nanog promoter, while the promoter of Gata4 lost H3K27me3 and became de-repressed in LIF/serum condition. Promoters associated with genes involved in MAPK signaling also showed misregulation of these histone marks. Such epigenetic modifications could be indirect consequences of mutating Mad2l2. However, our previous observations suggested the histone methyltransferases as direct (G9a) or indirect (Ezh2) targets of Mad2l2. In effect, the intricate balance necessary for pluripotency becomes perturbed in the absence of Mad2l2.

Keywords: MAP kinase; Mad2B; Rev7; differentiation; embryonic stem cells; pluripotency; primitive endoderm.

MeSH terms

  • Animals
  • Benzamides / pharmacology
  • Cell Differentiation / drug effects
  • Cells, Cultured
  • Diphenylamine / analogs & derivatives
  • Diphenylamine / pharmacology
  • Embryoid Bodies / cytology
  • Embryoid Bodies / metabolism
  • Enzyme Inhibitors / pharmacology
  • Epigenesis, Genetic
  • Glycogen Synthase Kinase 3 / metabolism
  • Glycogen Synthase Kinase 3 beta
  • Histones / metabolism
  • Homeodomain Proteins / genetics
  • Homeodomain Proteins / metabolism
  • Leukemia Inhibitory Factor / pharmacology
  • Mad2 Proteins / deficiency
  • Mad2 Proteins / genetics
  • Mad2 Proteins / metabolism*
  • Mice
  • Mitogen-Activated Protein Kinases / metabolism
  • Mouse Embryonic Stem Cells / cytology
  • Mouse Embryonic Stem Cells / metabolism
  • Nanog Homeobox Protein
  • Promoter Regions, Genetic
  • Pyridines / pharmacology
  • Pyrimidines / pharmacology
  • Signal Transduction / drug effects
  • Transcription Factors / genetics
  • Transcription Factors / metabolism

Substances

  • Benzamides
  • Chir 99021
  • Enzyme Inhibitors
  • Histones
  • Homeodomain Proteins
  • Leukemia Inhibitory Factor
  • Mad2 Proteins
  • Mad2l2 protein, mouse
  • Nanog Homeobox Protein
  • Nanog protein, mouse
  • PD 0325901
  • Pyridines
  • Pyrimidines
  • Transcription Factors
  • Diphenylamine
  • Glycogen Synthase Kinase 3 beta
  • Mitogen-Activated Protein Kinases
  • Glycogen Synthase Kinase 3