Regulatory elements within the agropine synthase promoter of T-DNA

DNA sequences involved in the expression of the agropine synthase gene (ags) of T-DNA were identified by analysis of transcriptional activity of promoter mutants in crown gall tumors of sunflower. Precise quantification of activity was achieved using a homologous reference gene as an internal standard. Analysis of 5'-deletion mutants demonstrated the requirement of 314 base pairs of upstream DNA sequences for optimal activity. Five regions involved in transcriptional regulation were identified in the 5'-flanking sequences between positions -74 and -314. Four of these regions make a positive contribution to promoter activity, and the fifth exerts a negative influence. The TATA motif (-26 to -33) and the TATA proximal domain (-74 to -105), which contains two sequences similar to the mammalian CCAAT box, are the major determinants of promoter activity. The two TATA distal domains A and B are separated by a negative element (-166 to -205) which may attenuate promoter strength by distancing the TATA distal domain B (-206 to -314 base pairs) from downstream components of the promoter. The TATA distal domain B contains the a/b repeat first described in the nopaline synthase (nos) promoter and was unable to support transcription in the absence of elements within the TATA proximal domain.