FIG. 6.

Prp43p is required for snoRNA-mediated methylation of C₂₃₃₇ in 25S rRNA at the 27S pre-rRNA stage. 2′-O-methylation by the box C/D snoRNA snR64 at C₂₃₃₇ was assessed by a site-specific RNase H cleavage protection assay (58). The unmodified 5.8S residue U₆₈ was assayed as a positive control for cleavage. The first two lanes represent negative controls. RNA from PRP43 or prp43-Q423N strains, shifted 2 h at 15°C, was RNase H digested and analyzed by Northern using probe o6. The schematic shown at the top indicates the position of C₂₃₃₇, U₆₈, and the o6 probe in 35S; the other schematics symbolize the cleavage products, marked by arrows and labeled with the pre-rRNA from which it derives. The asterisk marks nonspecific hybridization.