Crystal Structure and DNA Binding of the Homeodomain of the Stem Cell Transcription Factor Nanog

The transcription factor Nanog is an upstream regulator in early mammalian development and a key determinant of pluripotency in embryonic stem cells. Nanog binds to promoter elements of hundreds of target genes and regulates their expression by an as yet unknown mechanism. Here, we report the crysta...

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Bibliographic Details
Published in:Journal of molecular biology 2008-02, Vol.376 (3), p.758-770
Main Authors: Jauch, Ralf, Ng, Calista Keow Leng, Saikatendu, Kumar Singh, Stevens, Raymond C., Kolatkar, Prasanna R.
Format: Article
Language:English
Subjects:
AFFINITY
Amino Acid Sequence
AMINO ACIDS
Animals
CRYSTAL STRUCTURE
Crystallography, X-Ray
DNA
DNA binding
DNA-Binding Proteins - chemistry
FUNCTIONALS
GENES
homeodomain
Homeodomain Proteins - chemistry
MATERIALS SCIENCE
Mice
Models, Molecular
Molecular Sequence Data
MUTAGENESIS
Nanog
Nanog Homeobox Protein
pluripotency
Promoter Regions, Genetic
PROMOTERS
Protein Structure, Tertiary
RESIDUES
Sequence Alignment
SPECIFICITY
STEM CELLS
TARGETS
transcription factor
TRANSCRIPTION FACTORS
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Summary:The transcription factor Nanog is an upstream regulator in early mammalian development and a key determinant of pluripotency in embryonic stem cells. Nanog binds to promoter elements of hundreds of target genes and regulates their expression by an as yet unknown mechanism. Here, we report the crystal structure of the murine Nanog homeodomain (HD) and analysis of its interaction with a DNA element derived from the Tcf3 promoter. Two Nanog amino acid pairs, unique among HD sequences, appear to affect the mechanism of nonspecific DNA recognition as well as maintain the integrity of the structural scaffold. To assess selective DNA recognition by Nanog, we performed electrophoretic mobility shift assays using a panel of modified DNA binding sites and found that Nanog HD preferentially binds the TAAT(G/T)(G/T) motif. A series of rational mutagenesis experiments probing the role of six variant residues of Nanog on its DNA binding function establish their role in affecting binding affinity but not binding specificity. Together, the structural and functional evidence establish Nanog as a distant member of a Q50-type HD despite having considerable variation at the sequence level.
ISSN:0022-2836
1089-8638
DOI:10.1016/j.jmb.2007.11.091