Xenopus U3 snoRNA GAC-box A ' and box A sequences play distinct functionalroles in rRNA processing

Mutations in the 5 ' portion of Xenopus U3 snoRNA were tested for function in oocytes. The results revealed a new cleavage site (AO) in the 3 ' region of vertebrate external transcribed spacer sequences. In addition, U3 mutag enesis uncoupled cleavage at sites 1 and 2, flanking the 5 ' and 3 ' ends o f 18S rRNA, and generated novel intermediates: 19S and 18.5S pre-rRNAs. Fur thermore, specific nucleotides in Xenopus U3 snoRNA that are required for c leavages in pre-rRNA were identified: box A is essential for site AO cleava ge, the GAC-box A ' region is necessary for site I cleavage, and the 3 ' en d of box A ' and flanking nucleotides are required for site 2 cleavage. Dif ferences between metazoan and yeast U3 snoRNA-mediated rRNA processing are enumerated. The data support a model where metazoan U3 snoRNA acts as a bri dge to draw together the 5 ' and 3 ' ends of the 18S rRNA coding region wit hin pre-rRNA to coordinate their cleavage.