Identification of potential regulatory elements in the far-upstream region of the Arabidopsis thaliana plastocyanin promoter

Abstract

The far-upstream region of the Arabidopsis thaliana plastocyanin (Pc) promoter acts positively on transcription. This -1580 to -710 region (relative to the translation start site) has enhancer-like properties since it is also functional when situated downstream of the gene. Using tobacco nuclear extracts, this region was tested for protein-binding sites. Two short binding sequences were identified. The AT-rich sequence separating these binding sites shows extensive homology to the sequences separating the paired GT-1-binding sites of the pea rbcS-3A promoter. The requirements for complex formation strongly suggest that a GT-1-like protein binds to the two identified boxes in the Pc promoter. Sequence comparisons revealed that both boxes fit within the moderate consensus sequence needed for GT-1-binding. This GT-1-like DNA-binding activity is present in light-grown as well as in dark-adapted plants. Therefore, the possible role for GT-1 in light regulation of transcription does not depend upon its de novo synthesis. In some of the gel mobility shift assays, an additional DNA-protein complex was formed. The formation of this complex was only observed if the heteropolymer poly(dAdT).poly(dAdT) was used as a non-specific competitor and was dependent on the CpG density of the probe used.