THE YEAST HANSENULA-WINGEI U3 SNORNA GENE CONTAINS AN INTRON AND ITS CODING SEQUENCE CO-EVOLVED WITH THE 5'-ETS REGION OF THE PRE-RIBOSOMAL-RNA

The 5' external transcribed spacer (ETS) region of the pre-rRNA in Sac charomyces cerevisiae contains a sequence with 10 bp of perfect comple mentarity to the U3 snoRNA. Base pairing between these sequences has b een shown to be required for 18S rRNA synthesis, although interaction over the full in bp of complementarity is not required. We have identi fied the homologous sequence in the 5' ETS from the evolutionarily dis tant yeast Hansenula wingei; unexpectedly, this shows two sequence cha nges in the region predicted to base pair to U3. By PCR amplification and direct RNA sequencing, a single type of U3 snoRNA coding sequence was identified in H. wingei. As in the S. cerevisiae U3 snoRNA genes, it is interrupted by an intron with features characteristic of introns spliced in a spliceosome. Consequently, this unusual property is not restricted to the yeast genus Saccharomyces. The introns of the H. win gei and S. cerevisiae U3 genes show strong differences in length and s equence, but are located at the same position in the U3 sequence, imme diately upstream of the phylogenetically conserved Box A region. The 3 ' domains of the H. wingei and S. cerevisiae U3 snoRNAs diverge strong ly in primary sequence, but have very similar predicted secondary stru ctures. The 5' domains, expected to play a direct role in pre-ribosoma l RNA maturation, are more conserved. The sequence predicted to base p air to the pre-rRNA contains two nucleotide substitutions in H. wingei that restore in bp of perfect complementarity to the 5' ETS. This is a strong phylogenetic evidence for the importance of the U3/pre-rRNA i nteraction.