THE MINIMAL RIBOSOMAL-RNA GENE PROMOTER OF ARABIDOPSIS-THALIANA INCLUDES A CRITICAL ELEMENT AT THE TRANSCRIPTION INITIATION SITE

The genes encoding the precursor of 18S, 5.8S and 25S ribosomal RNAs a re transcribed in the nucleolus by RNA polymerase I. Unlike rRNA gene promoters in animals which differ substantially across species boundar ies, plant rRNA gene promoters share sequence similarity for several n ucleotides upstream and downstream of the transcription start site (+1 ). The conserved sequence consists of a near-consensus TATA box, a cri tical promoter element of most genes transcribed by RNA polymerase II and certain genes transcribed by RNA polymerase III, followed by a clu ster of four to six guanosines. Using transient expression of cloned p romoter deletions in Arabidopsis thaliana protoplasts, it is shown tha t the 5' boundary of the Arabidopsis rRNA gene promoter is located bet ween -55 and -33 and the 3' promoter boundary is approximately +6. The critical role of the TATA element at the initiation site was demonstr ated by altering this region using tandem point mutations within const ructs containing essentially complete intergenic spacer sequences from -2590 to +6. The initiation site mutations either abolished transcrip tion or dramatically reduced transcript abundance relative to the wild -type promoter. In some mutants, transcription initiation was shifted to a new site, suggesting a role for the TATA-like initiator region in both start site selection and promoter strength. It is suggested that minimal rRNA gene promoters might be similar in design to minimal pro moters of protein-encoding genes, many of which utilize an initiator e lement, or INR, as an important promoter element located directly at t he transcription start site.