Regulation of Cyclin E by transcription factors of the naïve pluripotency network in mouse embryonic stem cells

Cell Cycle. 2019 Oct;18(20):2697-2712. doi: 10.1080/15384101.2019.1656475. Epub 2019 Aug 28.

Abstract

Continuous, non-cell cycle-dependent expression of cyclin E is a characteristic feature of mouse embryonic stem cells (mESCs). We studied the 5' regulatory region of Cyclin E, also known as Ccne1, and identified binding sites for transcription factors of the naïve pluripotency network, including Esrrb, Klf4, and Tfcp2l1 within 1 kilobase upstream of the transcription start site. Luciferase assay and chromatin immunoprecipitation-quantitative polymerase chain reaction (ChiP-qPCR) study highlighted one binding site for Esrrb that is essential to transcriptional activity of the promoter region, and three binding sites for Klf4 and Tfcp2l1. Knockdown of Esrrb, Klf4, and Tfcp2l1 reduced Cyclin E expression whereas overexpression of Esrrb and Klf4 increased it, indicating a strong correlation between the expression level of these factors and that of cyclin E. We observed that cyclin E overexpression delays differentiation induced by Esrrb depletion, suggesting that cyclin E is an important target of Esrrb for differentiation blockade. We observed that mESCs express a low level of miR-15a and that transfection of a miR-15a mimic decreases Cyclin E mRNA level. These results lead to the conclusion that the high expression level of Cyclin E in mESCs can be attributed to transcriptional activation by Esrrb as well as to the absence of its negative regulator, miR-15a.

Keywords: Cyclin E; Embryonic stem cell; cell-cycle; pluripotent.

MeSH terms

  • Animals
  • Binding Sites
  • Cell Differentiation / genetics*
  • Cell Line
  • Chromatin Immunoprecipitation
  • Cyclin E / genetics
  • Cyclin E / metabolism*
  • Cyclins / genetics
  • Cyclins / metabolism*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • Gene Knockdown Techniques
  • Kruppel-Like Transcription Factors / genetics
  • Kruppel-Like Transcription Factors / metabolism
  • Mice
  • MicroRNAs / genetics
  • MicroRNAs / metabolism
  • Mouse Embryonic Stem Cells / metabolism*
  • Promoter Regions, Genetic*
  • RNA, Small Interfering
  • Receptors, Estrogen / genetics
  • Receptors, Estrogen / metabolism
  • Transcription Factors / genetics
  • Transcription Factors / metabolism
  • Up-Regulation

Substances

  • Ccne2 protein, mouse
  • Cyclin E
  • Cyclins
  • DNA-Binding Proteins
  • Esrrb protein, mouse
  • GKLF protein
  • Kruppel-Like Transcription Factors
  • MicroRNAs
  • Mirn15a microRNA, mouse
  • RNA, Small Interfering
  • Receptors, Estrogen
  • Tfcp2 protein, mouse
  • Transcription Factors

Grant support

This work was supported by the Agence Nationale de la Recherche [ANR-10-LABX-0061]; Agence Nationale de la Recherche [ANR-11-LABX-0042];Agence Nationale de la Recherche [ANR-10-IBHU-003]; Agence Nationale de la Recherche [ANR-11-INBS-0009]; Agence Nationale de la Recherche [ANR-11-IDEX-0007]; Fondation pour la Recherche Médicale [DEQ20170336757].