Construction of a trans-dominant inhibitor for members of the TGA family of transcription factors conserved in higher plants.

ABSTRACT

Activating sequence factor 1 (ASF-1) is a conserved DNA-binding activity that interacts with sequence elements containing TGACG motifs, some of which have been demonstrated to respond to exogenous application of auxin and salicylic acid. Genes encoding transcription factors with similar DNA-binding specificity to ASF-1 have been cloned from diverse plant species and these factors all contain a distinct basic-leucine-zipper (bZIP) motif. Members of this family of DNA-binding proteins, designated as TGA factors, have been shown to interact with similar DNA sequences and at least seven distinct TGA genes are present in the Arabidopsis genome. To study the roles that this family of factors may play in plant development, a trans-dominant inhibitor of ASF-1 was constructed by deleting the basic portion of the bZIP domain in the tobacco factor TGA1a. In vitro co-expression studies demonstrated that this deletion mutant, named TGA1a-D, suppresses the DNA-binding activity of wild-type tobacco TGA1a, and three different members of the Arabidopsis TGA family. In contrast, co-expression of TGA1a-D with another class of bZIP proteins, the G-Box Binding Factor family, showed no suppression of DNA-binding activity. Over-expression of TGA1a-D in transgenic tobacco significantly decreased nuclear ASF-1 relative to several other known factors, indicating that the proteins comprising ASF-1 activity in vivo are likely TGA family members. Thus, TGA1a-D may be a family-specific inhibitor for the TGA family and should facilitate the study of ASF-1 function in vivo.