Conserved sequence elements upstream of the gene encoding yeast ribosomal protein L25 are involved in transcription...

Previous studies have revealed the occurrence of two closely linked conserved sequence elements, designated as HOMOL 1 and RPG box, in front of most yeast ribosomal protein genes examined. To investigate whether these conserved nucleotide elements play a role in the regulation of ribosomal protein gene expression, we performed deletion analysis of the DNA region upstream of the gene encoding ribosomal protein L25. To that end we constructed a hybrid gene consisting of the pertinent 5'-flanking sequence and the Escherichia coli galK marker gene. The effects on the transcription of this fusion gene of Bal31-generated deletions were measured by Northern analysis of RNA isolated from the respective transformed yeast cells. The results demonstrate that removal of one box has a detrimental effect on the level of transcription, whereas after the deletion of both boxes hardly any transcription can be observed. Subsequently we inserted synthetic oligonucleotides in the upstream region of an L25 gene from which the original boxes had been removed. Expression of the inactivated hybrid gene turned out to be restored even by insertion of one RPG element. Moreover, the RPG box functions in both orientations, though not with equal efficiency.