DUAL DNA-BINDING SPECIFICITY OF A PETAL EPIDERMIS-SPECIFIC MYB TRANSCRIPTION FACTOR (MYB.PH3) FROM PETUNIA-HYBRIDA

The MYB.Ph3 protein recognized two DNA sequences that resemble the two known types of MYB DNA binding site: consensus I (MBSI), aaaAaaC(G/C) -GTTA, and consensus II (MBSII), aaaAGTTAGTTA. Optimal MBSI was recogn ized by animal c-MYB and not by Am305 from Antirrhinum, whereas MBSII showed the reverse behaviour. Different constraints on MYB.Ph3 binding to the two classes of sequences were demonstrated. DNA binding studie s with mutated MBSI and MBSII and hydroxyl radical footprinting analys is, pointed to the N-terminal MYB repeat (R2) as the most involved in determining the dual DNA binding specificity of MYB.Ph3 and supported the idea that binding to MBSI and MBSII does not involve alternative o rientations of the two repeats of MYB.Ph3. Minimal promoters containin g either MBSI and MBSII were activated to the same extent by MYB.Ph3 i n yeast, indicating that both types of binding site can be functionall y equivalent. MYB.Ph3 binding sites are present in the promoter of fla vonoid biosynthetic genes, such as the Petunia chsJ gene, which was tr anscriptionally activated by MYB.Ph3 in tobacco protoplasts. MYB.Ph3 w as immunolocalized in the epidermal cell layer of petals, where flavon oid biosynthetic genes are actively expressed, This strongly suggests a role for MYB.Ph3 in the regulation of flavonoid biosynthesis.