SEQUENCE-SPECIFIC DNA RECOGNITION BY THE THYROID TRANSCRIPTION FACTOR-I HOMEODOMAIN

The molecular basis for the DNA binding specificity of the thyroid tra nscription factor 1 homeodomain (TTF-1HD) has been investigated. Methy lation and ethylation interference experiments show that the TTF-1HD a lone recapitulates the DNA binding properties of the entire protein. S tudies carried out with mutant derivatives of TTF-1HD indicate a preci se correspondence of some of its amino acid residues with specific bas es in its binding site, allowing a crude orientation of the TTF-1HD wi thin the protein - DNA complex. TTF-1HD shows an overall geometry of i nteraction with DNA similar to that previously observed for Antennaped ia class HDs, even though the binding specificities of these two types of HDs are distinct. We demonstrate that the crucial difference betwe en the binding sites of Antennapedia class and TTF-1 HDs is in the mot ifs 5'-TAAT-3', recognized by Antennapedia, and 5'-CAAG-3', preferenti ally bound by TTF-1. Furthermore, the binding of wild type and mutants TTF-1 HD to oligonucleotides containing either 5'-TAAT-3' or 5'-CAAG- 3' indicate that only in the presence of the latter motif the Gin,, in TTF-1 HD is utilized for DNA recognition. Since the Gin at position 5 0 is an essential determinant for DNA binding specificity for several other HDs that bind to 5'-TAAT-3' containing sequences, we suggest tha t utilization by different HDs of key residues may depend on the seque nce context and probably follows a precise hierarchy of contacts.