A High-Throughput Genome-Integrated Assay Reveals Spatial Dependencies Governing Tcf7l2 Binding

Cell Syst. 2020 Sep 23;11(3):315-327.e5. doi: 10.1016/j.cels.2020.08.004. Epub 2020 Sep 9.


Predicting where transcription factors bind in the genome from their in vitro DNA-binding affinity is confounded by the large number of possible interactions with nearby transcription factors. To characterize the in vivo binding logic for the Wnt effector Tcf7l2, we developed a high-throughput screening platform in which thousands of synthesized DNA phrases are inserted into a specific genomic locus, followed by measurement of Tcf7l2 binding by DamID. Using this platform at two genomic loci in mouse embryonic stem cells, we show that while the binding of Tcf7l2 closely follows the in vitro motif-binding strength and is influenced by local chromatin accessibility, it is also strongly affected by the surrounding 99 bp of sequence. Through controlled sequence perturbation, we show that Oct4 and Klf4 motifs promote Tcf7l2 binding, particularly in the adjacent ∼50 bp and oscillating with a 10.8-bp phasing relative to these cofactor motifs, which matches the turn of a DNA helix.

Keywords: CRISPR-Cas9; DamID; Gaussian process; Tcf7l2; transcription factor.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites
  • High-Throughput Screening Assays / methods*
  • Humans
  • Kruppel-Like Factor 4
  • Transcription Factor 7-Like 2 Protein / metabolism*
  • Transcription Factors / genetics*


  • KLF4 protein, human
  • Klf4 protein, mouse
  • Kruppel-Like Factor 4
  • TCF7L2 protein, human
  • Transcription Factor 7-Like 2 Protein
  • Transcription Factors