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Transcription of the gene encoding yeast ribosomal protein L25 was previously shown to be activated through tandemly arranged upstream sequence elements that most rp-genes in yeast have in common. A single copy of such a conserved element is now demonstrated to restore transcription of an inactivated heterologous gene, which confirms its role as a genuine UAS: UASrpg. Though a single box is sufficient to activate transcription, most rp-genes harbor two neighbouring elements. Northern analysis of mutants of the L25 upstream region lacking either the gene-distal (RPG1) or the gene-proximal (RPG2) box provided evidence that RPG2 is significantly more effective than RPG1 in vivo. Moreover the sum of the effects of the individual boxes as measured separately is significantly lower than their joint effect, supporting cooperative interaction between the two boxes in vivo. Making use of oligomer-insertion experiments several additional features of the UASrpgwere elucidated. First of all we confirmed that the extent of transcription activation by the UASrpg depends upon the orientation of the element. Secondly we show that a certain minimal distance (> 100 n) between UASrpg and the transcription initiation site is required for transcription activation. Finally, internal deletion of the L25-upstream region as well as oligomer-insertion shed some light on the nucleotide requirements of the UASrpg.