Octamer and Sox Elements Are Required for Transcriptional cis Regulation of Nanog Gene Expression

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.

Functional analysis of two Sp1/Sp3 binding sites in murine Nanog gene promoter

“…The loss of Nanog does not affect oct-4 gene expression, and the loss of oct-4 does not affect Nanog gene expression in the blastocyst [2]. However, there were evidences showing that Oct-4 is necessary for Nanog gene expression [3], which was confirmed by recent reports that Oct-4 and Sox-2 synergistically activate Nanog gene expression [4,5].…”

“…Like many other genes in early embryo as fgf4, sox-2, utf1 and fbx15, Nanog gene expression is regulated by Oct-4 and Sox-2 synergistically [4,5]. However, overexpression of Oct-4 or Sox-2, or both, could not effectively activate the Nanog gene promoter [5].…”

“…However, overexpression of Oct-4 or Sox-2, or both, could not effectively activate the Nanog gene promoter [5]. Here we further analyzed other putative bindings sites located within the same promoter region, which may be responsible for Nanog promoter activity.…”

Defective Myofibroblast Formation from Mesenchymal Stem Cells in the Aging Murine Heart

“…The same degree of down-regulation was observed using both techniques, which suggested that Nanog expression was controlled on a transcriptional level in those cells, as has been reported by others. 27,28 Because some protein expression [ie, lamin A, GAPDH (glyceraldehyde-3-phosphate dehydrogenase), and actin, all used as loading indicators] is affected by aging (lamin A and GAPDH not shown), we included photographs of membranes stained using Ponceau S with our results of Western blot analysis to ensure equal protein loading and to enable comparison of protein concentrations.…”

β-Catenin up-regulates Nanog expression through interaction with Oct-3/4 in embryonic stem cells

“…The finding that these three factors co-occupy a substantial portion of their target genes suggests that they work collaboratively to maintain ES cell self-renewal [7]. Recent studies have also demonstrated that the Oct-3/4•Sox2 complex regulates Nanog expression through binding to the promoter region of Nanog [8,9].…”