A single residue substitution causes a switch from the dual DNA binding specificity of plant transcription factor MYB.Ph3 to the animal c-MYB specificity.
Transcription factor MYB.Ph3 from Petunia binds to two types of sequences, MBSI and MBSII, whereas murine c-MYB only binds to MBSI, and Am305 from Antirrhinum only binds to MBSII. DNA binding studies with hybrids of these proteins pointed to the N-terminal repeat (R2) as the most involved in determining binding to MBSI and/or MBSII, although some influence of the C-terminal repeat (R3) was also evident. Furthermore, a single residue substitution (Leu71 --> Glu) in MYB.Ph3 changed its specificity to that of c-MYB, and c-MYB with the reciprocal substitution (Glu132 --> Leu) essentially gained the MYB.Ph3 specificity. Molecular modeling and DNA binding studies with site-specific MYB.Ph3 mutants strongly supported the notion that the drastic changes in DNA binding specificity caused by the Leu --> Glu substitution reflect the fact that certain residues influence this property both directly, through base contacts, and indirectly, through interactions with other base-contacting residues, and that a single residue may establish alternative base contacts in different targets. Additionally, differential effects of mutations at non-base-contacting residues in MYB.Ph3 and c-MYB were observed, reflecting the importance of protein context on DNA binding properties of MYB proteins.
Solano
R
R
Centro Nacional de Biotecnología-CSIC, Campus Cantoblanco, Carretera de Colmenar Km 15.5, Madrid 28049, Spain.
Fuertes
A
A
Sánchez-Pulido
L
L
Valencia
A
A
Paz-Ares
J
J
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D051379:Mice; D008961:Models, Structural; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism; D010944:Plants / Q000378:metabolism*; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D017433:Protein Structure, Secondary; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D020598:Proto-Oncogene Proteins c-myb; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D015534:Trans-Activators / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D014157:Transcription Factors / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*
D004268:DNA-Binding Proteins; D009838:Oligodeoxyribonucleotides; D010940:Plant Proteins; D011518:Proto-Oncogene Proteins; D020598:Proto-Oncogene Proteins c-myb; D015534:Trans-Activators; D014157:Transcription Factors; D004247:DNA