GCN4P ACTIVATION OF THE YEAST TRP3 GENE IS ENHANCED BY ABF1P AND USESA SUBOPTIMAL TATA ELEMENT

Transcription of the TRP3 gene of Saccharomyces cerevisiae is regulate d by GCN4p from a position proximal to the transcriptional initiation sites. The promoter's apparent lack of a conventional TATA element seq uence has led it to be used as a model for TATA-less promoters. Throug h mutational analysis of the TRP3 promoter, we have identified two add itional regulatory elements required for expression. The first, locate d 57 base pairs (bp) upstream of the GCN4p binding site, binds ABF1p i n vitro. The ABF1p binding site was required for maximal levels of GCN 4p-activated transcription in vivo; however, the -fold activation by G CN4p was not altered by ABF1p. The second element, positioned 23 bp do wnstream of the GCN4p binding site, contained the TATAlike sequence, T ATTAA. This element was required for both basal and activated expressi on and almost certainly functions as a TATA-binding protein interactio n site. Mutations that improved its TATA character for native or an al tered specificity mutant of TATA-binding protein correspondingly impro ved its function. Interestingly, basal expression induced by ABF1p was virtually unchanged in the presence of point mutations in the TATTAA element. Furthermore, unlike the case for HIS3 where only a limited su bset of TATA-like sequences can activate transcription in conjunction with GCN4p, many divergent TATA-like sequences allowed GCN4p activatio n of TRP3. We suggest that the apparent promoter specific use of these TATA elements by GCN4p results from ABF1p amplifying the GCN4p-induce d expression to a detectable level.