SEQUENCE-SPECIFIC INITIATOR ELEMENTS FOCUS INITIATION OF TRANSCRIPTION TO DISTINCT SITES IN THE YEAST TRP4 PROMOTER

Transcription from the yeast TRP4 promoter initiates at two basal (i127 and i76) and three GCN4 dependent (i31, i25 and i12) initiator elements. All of these elements contain not more than one deviation from the earlier proposed initiator consensus sequence PuPuPyPuPu, a pyrimidine nucleotide flanked on either side by two purine nucleotides. A point mutation analysis of these elements in various combinations was performed and revealed that the central pyrimidine nucleotide and at least one of the 3' flanking purine nucleotides of the PuPuPyPuPu consensus sequence are essential but alone not sufficient to define a functional initiator element. Multiple cryptic transcription start sites, which function independently whether they are located on the coding or the non-coding strand, can replace the function of mutated initiator elements and therefore the overall level of transcription initiation is not affected. The sequence specificity is identical for basal and GCN4 dependent initiator elements demonstrating that they are functionally homologous. These findings imply that the role of initiator elements is to 'focus' the start point(s) of transcription to distinct sites located in the region between the site(s) of the assembly of the transcriptional complex and the start codon of translation.