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Fig. 3
Interaction of Her1 and Her7:Hes6 with her1, her7, and dlc promoter fragments in a yeast one-hybrid assay.
(A, B, C) 18 yeast strains carrying promoter fragments of the dlc (A), her7 (B), or her1 (C) genes coupled to His- and lacZ-reporter genes were transformed with plasmids encoding N-terminally Gal4-AD tagged Her1, Her7, or Hes6. Protein-DNA interactions were detected by staining for β-galactosidase, except for two fragments from the her7 promoter (1647 to 2491) and dlc promoter (719 to 1114), where self-activation precluded use of the β-gal assay and interactions were detected by growth in the presence of 3-aminotriazole. Colonies were arrayed in quadruplicates to aid identification of positives. Numbers indicate start and end of fragments relative to the gene′s start codon in the genome. (D) Comparison of MITOMI and yeast one-hybrid results. Promoter fragments are depicted by grey and green horizontal bars, with start codon to the right. Fragments displaying interaction with Her1 are in green, and fragments without interaction with Her1 are in grey. Vertical bars indicate H-box sites. Relative binding affinity of each H-box to Her1 as determined by MITOMI is color-coded from black (low affinity) to yellow (high affinity). For relative affinities of individual sites, see Table S1. Numbers indicate distance from start codon. (E) Bait strains were transformed with pAD-DIMER vectors encoding two copies of AD-tagged her1, her7, or hes6 or a combination of her7 and hes6 and interactions detected as in (A–C). Only one fragment each from the her1, her7, and the dlc promoter gives signals with Her1 only expressed from pAD-DIMER, and the same fragments interact with a combination of Her7 and Hes6.