Octamer and Sox elements are required for transcriptional cis regulation of Nanog gene expression

Mol Cell Biol. 2005 Mar;25(6):2475-85. doi: 10.1128/MCB.25.6.2475-2485.2005.

Abstract

The pluripotential cell-specific gene Nanog encodes a homeodomain-bearing transcription factor required for maintaining the undifferentiated state of stem cells. However, the molecular mechanisms that regulate Nanog gene expression are largely unknown. To address this important issue, we used luciferase assays to monitor the relative activities of deletion fragments from the 5'-flanking region of the gene. An adjacent pair of highly conserved Octamer- and Sox-binding sites was found to be essential for activating pluripotential state-specific gene expression. Furthermore, the 5'-end fragment encompassing the Octamer/Sox element was sufficient for inducing the proper expression of a green fluorescent protein reporter gene even in human embryonic stem (ES) cells. The potential of OCT4 and SOX2 to bind to this element was verified by electrophoretic mobility shift assays with extracts from F9 embryonal carcinoma cells and embryonic germ cells derived from embryonic day 12.5 embryos. However, in ES cell extracts, a complex of OCT4 with an undefined factor preferentially bound to the Octamer/Sox element. Thus, Nanog transcription may be regulated through an interaction between Oct4 and Sox2 or a novel pluripotential cell-specific Sox element-binding factor which is prominent in ES cells.

MeSH terms

  • 5' Flanking Region / genetics
  • Animals
  • Base Sequence
  • Binding Sites / genetics
  • DNA-Binding Proteins / genetics*
  • DNA-Binding Proteins / metabolism
  • DNA-Binding Proteins / physiology*
  • Electrophoretic Mobility Shift Assay
  • Embryo, Mammalian / cytology
  • Gene Expression Regulation*
  • Genes, Reporter / genetics
  • Green Fluorescent Proteins / analysis
  • Green Fluorescent Proteins / genetics
  • HMGB Proteins
  • Homeodomain Proteins / genetics*
  • Humans
  • Luciferases / analysis
  • Luciferases / genetics
  • Mice
  • Molecular Sequence Data
  • Mutation / genetics
  • Nanog Homeobox Protein
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • Nuclear Proteins / physiology*
  • Octamer Transcription Factor-3
  • Response Elements / genetics*
  • SOXB1 Transcription Factors
  • Sequence Deletion / genetics
  • Stem Cells / metabolism*
  • Transcription Factors / genetics
  • Transcription Factors / metabolism
  • Transcription Factors / physiology*
  • Transcription, Genetic

Substances

  • DNA-Binding Proteins
  • HMGB Proteins
  • Homeodomain Proteins
  • NANOG protein, human
  • Nanog Homeobox Protein
  • Nanog protein, mouse
  • Nuclear Proteins
  • Octamer Transcription Factor-3
  • POU5F1 protein, human
  • Pou5f1 protein, mouse
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • Sox2 protein, mouse
  • Transcription Factors
  • enhanced green fluorescent protein
  • Green Fluorescent Proteins
  • Luciferases