Flexibility and Interchangeability of Polyadenylation Signals in Saccharomyces cerevisiae

Abstract

Various signal motifs have been reported to be essential for proper mRNA 3′-end formation in the yeast Saccharomyces cerevisiae. However, none of these motifs has been shown to be sufficient to direct 3′-end processing and/or transcription termination. Therefore, several structural motifs have to act in concert for efficient 3′-end formation. In the region upstream of the three polyadenylation sites of the yeast gene for alcohol dehydrogenase I (ADH1), we have identified a hitherto unknown signal sequence contained within the octamer AAAAAAAA. This motif, located 11 nucleotides upstream of the first ADH1 polyadenylation site, is responsible for the utilization of this site in vitro and in vivo, since mutational alteration drastically reduced 3′-end formation at this position. Insertion of 38 ADH1 -derived nucleotides encompassing the (A)₈ motif into the 3′-end formation-deficient cycl-512 deletion mutant restored full processing capacity in vitro. Insertion of the octamer alone did not restore 3′-end formation, although mutation of the (A)₈ motif in the functional construct had abolished 3′-end processing activity almost completely. This demonstrates that the sequence AAAAAAAA is a necessary, although not sufficient, signal for efficient mRNA 3′-end formation in S. cerevisiae.