Genome-wide mapping of transcription factor Oct4, Sox2 and Nanog

binding-sites in mouse embryonic stem cells

Genome Institute of SingaporeDepartment of Biological SciencesNational University of Singapore

ChIP-PET technology to map ES cell specific transcription factors

Yijun Ruan / Chia-Lin Wei (Cloning & Sequencing, Genome Institute of Singapore)

ChIP-PETcapturesOct4binding profile

Determination of cutoff for ChIP-PET data

cutoff

Oct4 Sox2 Nanog

No. of binding sites ~1000 ~1000 ~3000

Based on 4 PET overlap as a cutoff.

Distribution of Oct4 binding sites

Binding site in gene desert

Sox2

Distribution of Nanog binding sites

Oct4Oct4Sox2Sox2

de novo prediction of motifs

Oct4Sox2

The code for targeting Oct4 to the genome

Oct4Oct4Sox2Sox2

Oct4Sox2

1000 Sox2 binding sites1000 Sox2 binding sites

Sox2 binding sites contain a prominent Sox2-Oct4 motifSox2 binding sites contain a prominent Sox2-Oct4 motif

de novode novopredictionpredictionof motifof motif

Question 1: What are the motifs bound by these TFs?Question 1: What are the motifs bound by these TFs?

Oct4 Sox2 Nanog

Oct4 Oct4Sox2 Sox2

?

Oct4 and Sox2 are primarily recruited to the genomeOct4 and Sox2 are primarily recruited to the genomethrough the joint sox2 oct4 motifsthrough the joint sox2 oct4 motifs

de novo prediction of motifs using the ChIP-PET data

What are the motifs bound by Nanog?What are the motifs bound by Nanog?

What are the motifs bound by Nanog?What are the motifs bound by Nanog?

Refined using ChIP-PET dataRefined using ChIP-PET data

Sequences found in EMSA validated probesSequences found in EMSA validated probes

Oct4 positively regulates the expression of novel target genes

Oct4 negatively regulates the expression of Cdx2(trophectoderm transcription factor)

Nanog

Sox2

microRNAs targeted by Nanog

“Core” Target Genes of Oct4, Sox2 and Nanog in mouse ES cells

Nanog

Sox2

Oct4

Configuration of transcription factors interaction with upstream regulatory regionConfiguration of transcription factors interaction with upstream regulatory region

Co-occupancy of sites by OCt4/Sox2/nanog

Whole genome mapsFor Oct4 and Nanog

Oct4 Nanog Sox2

Mouse vs Human comparison of Oct4, Nanog and Sox2 TFBS

Comparison between mouse and human Sox2 binding sitesSox2

SummarySummary

Oct4 and Sox2 make use of oct-sox joint motifs for binding Oct4 and Sox2 make use of oct-sox joint motifs for binding (increased specificities?)(increased specificities?)

Nanog’s motif is abundant in the genomeNanog’s motif is abundant in the genome

Co-targeting of genes by the 3 transcription factorsCo-targeting of genes by the 3 transcription factors Coordinated regulationCoordinated regulation

The 3 transcription factors can regulate positively andThe 3 transcription factors can regulate positively andnegatively regulate transcriptionnegatively regulate transcription

The target genes fall into diverse functional categoriesThe target genes fall into diverse functional categories DNA response pathwayDNA response pathway Cellular proliferationCellular proliferation Inhibitors of lineage specific genesInhibitors of lineage specific genes

Acknowledgement:

Huck Hui NgQiang WuJoon-Lin ChewYuin-Han LohXi ChenWeiwei Zhang

Cloning & Sequencing GroupYijun RuanChia-Lin Wei

Stem Cell Biology GroupBing LimPaul RobsonLarry Stanton

InformaticsVegaKen SungGuillaume BourqueVladLeonard

Funding:

Biomedical Research CouncilA*STARNational University of SingaporeSingapore Millennium Foundation

Comparison between mouse and human Oct4 binding sitesOct4