Coop-Seq Analysis Demonstrates that Sox2 Evokes Latent Specificities in the DNA Recognition by Pax6

J Mol Biol. 2017 Nov 24;429(23):3626-3634. doi: 10.1016/j.jmb.2017.10.013. Epub 2017 Oct 16.


Sox2 and Pax6 co-regulate genes in neural lineages and the lens by forming a ternary complex likely facilitated allosterically through DNA. We used the quantitative and scalable cooperativity-by-sequencing (Coop-seq) approach to interrogate Sox2/Pax6 dimerization on a DNA library where five positions of the Pax6 half-site were randomized yielding 1024 cooperativity factors. Consensus positions normally required for the high-affinity DNA binding by Pax6 need to be mutated for effective dimerization with Sox2. Out of the five randomized bases, a 5' thymidine is present in most of the top ranking elements. However, this thymidine maps to a region outside of the Pax half site and is not expected to directly interact with Pax6 in known binding modes suggesting structural reconfigurations. Re-analysis of ChIP-seq data identified several genomic regions where the cooperativity promoting sequence pattern is co-bound by Sox2 and Pax6. A highly conserved Sox2/Pax6 bound site near the Sprouty2 locus was verified to promote cooperative dimerization designating Sprouty2 as a potential target reliant on Sox2/Pax6 cooperativity in several neural cell types. Collectively, the functional interplay of Sox2 and Pax6 demands the relaxation of high-affinity binding sites and is enabled by alternative DNA sequences. We conclude that this binding mode evolved to warrant that a subset of target genes is only regulated in the presence of suitable partner factors.

Keywords: Coop-seq; Pax6; Sox2; cooperativity; deep sequencing; gene regulation; transcription factors.

Publication types

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

MeSH terms

  • DNA / chemistry
  • DNA / genetics
  • DNA / metabolism*
  • Humans
  • Models, Molecular
  • PAX6 Transcription Factor / chemistry
  • PAX6 Transcription Factor / genetics
  • PAX6 Transcription Factor / metabolism*
  • Protein Binding
  • Protein Conformation
  • Protein Multimerization
  • SOXB1 Transcription Factors / chemistry
  • SOXB1 Transcription Factors / genetics
  • SOXB1 Transcription Factors / metabolism*
  • Sequence Analysis, DNA / methods*


  • PAX6 Transcription Factor
  • PAX6 protein, human
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • DNA