A SMALL NUCLEOLAR RNA REQUIREMENT FOR SITE-SPECIFIC RIBOSE METHYLATION OF RIBOSOMAL-RNA IN XENOPUS

Vertebrate cells contain a large number of small nucleolar RNA (snoRNA ) species, the vast majority of which bind fibrillarin. Most of the fi brillarin-associated snoRNAs can form 10- to 21-nt duplexes with rRNA and are thought to guide 2'-O-methylation of selected nucleotides in r RNA. These include mammalian UHG (U22 host gene)-encoded U25-U31 snoRN As. We have characterized two novel human snoRNA species, U62 and U63, which similarly exhibit 15- (with one interruption) and 12-nt complem entarities and are therefore predicted to direct 2'-O-methylation of A 590 in 18S and A4531 in 28S rRNA, respectively. To establish the funct ion of antisense snoRNAs in vertebrates, we exploited the Xenopus oocy te system. Cloning of the Xenopus U25-U31 snoRNA genes indicated that they are encoded within multiple homologs of mammalian UHG. Depletion of U25 from the Xenopus oocyte abolished 2'-O-methylation of G1448 in 18S rRNA; methylation could be restored by injecting either the Xenopu s or human U25 transcript into U25-depleted oocytes. Comparison of Xen opus and human U25 sequences revealed that only boxes C, D, and D', as well as the 18S rRNA complement, were invariant, suggesting that they may be the only elements required for U25 snoRNA stability and functi on.