BINDING OF MYC PROTEINS TO CANONICAL AND NONCANONICAL DNA-SEQUENCES

Using an in vitro binding-site selection assay, we have demonstrated t hat c-Myc-Max complexes bind not only to canonical CACGTG or CATGTG mo tifs that are flanked by variable sequences but also to noncanonical s ites that consist of an internal CG or TG dinucleotide in the context of particular variations in the CA--TG consensus. None of the selected sites contain an internal TA dinucleotide, suggesting that Myc protei ns necessarily bind asymmetrically in the context of a CAT half-site. The noncanonical sites can all be bound by proteins of the Myc-Max fam ily but not necessarily by the related CACGTG- and CATGTG-binding prot eins USF and TFE3. Substitution of an arginine that is conserved in th ese proteins into MyoD (MyoD-R) changes its binding specificity so tha t it recognizes CACGTG instead of the MyoD cognate sequence (CAGCTG). However, like USF and TFE3, MyoD-R does not bind to all of the noncano nical c-Myc-Max sites. Although this R substitution changes the intern al dinucleotide specificity of MyoD, it does not significantly alter i ts wild-type binding sequence preferences at positions outside of the CA--TG motif, suggesting that it does not dramatically change other im portant amino acid-DNA contacts; this observation has important implic ations for models of basic-helix-loop-helix protein-DNA binding.