Figure 5.
Box C of U3 snoRNA is required for U3-55k association in vitro. In vitro translated, 35S-labeled hU3-55k was added to in vitro transcribed, UTP-21-biotin-labeled RNA. The mixture was incubated for 1 h and biotinylated RNAs were selected from the mixture with streptavidin agarose beads. hU3-55k associated with the biotinylated RNA was eluted by boiling in SDS loading buffer and analyzed by SDS–PAGE and fluorography. Biotinylated RNAs used in each reaction are indicated above each lane. (A) Evaluation of U3-55k association with wild-type (lane 1) and mutant (ΔA, ΔC′, ΔB, ΔC and ΔD, lanes 2–6, respectively) U3 snoRNA as well as U8 (lane 7). (B) Evaluation of U3-55k association with U3 snoRNA 3′ domain and Box B/C sub-fragments (lanes 2 and 3). Wild-type U3 and U8 are shown in lanes 1 and 4.