FIG. 5.

Multiple regions on the surface of 15.5K are required for box C/D snoRNP assembly. HeLa nuclear extract was preincubated with either 800 pmol of U4-SL1 (SL1) RNA oligonucleotide (+) or buffer (−). Radiolabeled U14 snoRNA was subsequently added in the presence (lanes 2 to 8) or absence (−) (lanes 1 and 2) of wild-type or mutant 15.5K. The binding of individual snoRNP proteins was then assayed by immunoprecipitation (see Materials and Methods). Bound RNAs were recovered and then separated on an 8% polyacrylamide-7 M urea gel. Antibodies used for immunoprecipitation are indicated on the right. Ten percent of the RNA after incubation in nuclear extract was used as input in the bottom gel. The 15.5K protein used is indicated above each lane (15.5K-Wt, wild-type 15.5K).