FIG. 4.
The association of CYPH/hPRP4/hPRP3 to the U4/U6 snRNA requires multiple regions on the surface of 15.5K. Purified CYPH/hPRP4/hPRP3 complex was incubated with U4/U6 snRNA duplex (U4 snRNA base paired with 32P-labeled U6 nucleotides 58 to 87 [Fig. 1]) in the presence (lanes 2 to 7) or absence (lane 1) of wild-type or mutant 15.5K. The CYPH/hPRP4/hPRP3 complex was then immunoprecipitated using anti-hPRP4 antibody (α-hPRP4) and the coprecipitating RNA and the RNA present in the supernatant (5% Sup.) were analyzed on a denaturing 8% polyacrylamide-7 M urea gel and visualized by autoradiography. The identity of the 15.5K protein used is indicated above each lane (−, no 15.5K; 15.5K-Wt, wild-type 15.5K).