Fig. 5

Pou5f3 and Nanog bind in a mutually exclusive manner to overlapping motifs.

a–c Gel-retardation assays with the indicated oligos.*, ** - genomic locations oligo 3 and oligo 5 are shown in Fig. 4e, f. a Pou5f3-binding oligos from 2.P + N− antagonistic enhancers. Weaker Nanog binding is also detectable for oligo 3 (magenta arrow); see Fig. S6b with longer exposure time for Nanog-HA supershift with oligo 3 and Nanog binding to oligo 9. b Pou5f3-binding oligos from 1.P + N+ synergistic enhancers (oligo 1 is from mych⁵⁵ enhancer, Nanog binding was not detectable). See Fig. S6c with longer exposure time for Nanog binding to oligo 8 and oligo 7. c Nanog- binding oligos from 3.N + P- antagonistic enhancers. Weaker Pou5f3 binding is also detectable for oligo 6 (magenta arrow). See Fig. S6d with longer exposure times for Nanog-HA supershifts. d Aligned Pou- and Nanog- strong consensus binding sequences from our assays (in bold); homeodomain-binding part is underlined. e Left: Pou5f3 and Nanog binding motifs share common part, recognized by homeodomains (black box). Right: in oligo 3 and oligo 6, pou and nanog motifs overlap in homeodomain-recognition part, so that only Pou5f3 or only Nanog can contact DNA at the same time. SoxB1 binding part of sox:pou motif²⁰ is indicated in red. n.s – non-specific band. Source data are provided as a Source Data file 4.