Genome-wide mapping of transcription factor Oct4, Sox2 and Nanog binding-sites in mouse embryonic stem cells Genome Institute of Singapore Department of Biological Sciences National 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-PET captures Oct4 binding 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

Oct4Sox2 de novo prediction of motifs Oct4Sox2 The code for targeting Oct4 to the genome

Oct4Sox2 Oct4Sox Sox2 binding sites Sox2 binding sites contain a prominent Sox2-Oct4 motif de novo prediction of motif

Question 1: What are the motifs bound by these TFs? Oct4Sox2Nanog Oct4 Sox2 ? Oct4 and Sox2 are primarily recruited to the genome through the joint sox2 oct4 motifs de novo prediction of motifs using the ChIP-PET data

What are the motifs bound by Nanog?

Refined using ChIP-PET data Sequences 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 region

Co-occupancy of sites by OCt4/Sox2/nanog

Whole genome maps For Oct4 and Nanog

Oct4NanogSox2 Mouse vs Human comparison of Oct4, Nanog and Sox2 TFBS

Comparison between mouse and human Sox2 binding sites Sox2

Summary Oct4 and Sox2 make use of oct-sox joint motifs for binding (increased specificities?) Nanog’s motif is abundant in the genome Co-targeting of genes by the 3 transcription factors  Coordinated regulation The 3 transcription factors can regulate positively and negatively regulate transcription The target genes fall into diverse functional categories  DNA response pathway  Cellular proliferation  Inhibitors of lineage specific genes

Acknowledgement: Huck Hui Ng Qiang Wu Joon-Lin Chew Yuin-Han Loh Xi Chen Weiwei Zhang Cloning & Sequencing Group Yijun Ruan Chia-Lin Wei Stem Cell Biology Group Bing Lim Paul Robson Larry Stanton Informatics Vega Ken Sung Guillaume Bourque Vlad Leonard Funding: Biomedical Research Council A*STAR National University of Singapore Singapore Millennium Foundation

Comparison between mouse and human Oct4 binding sites Oct4