FIG. 10.

Bop1 and Bop1Δ cofractionate with the 50S-80S pre-RNP particles in nuclear extracts. (A to C) The inducible Bop1Δ expression cell line (line 6) was grown in the presence (C) or absence (A and B) of IPTG for 24 h as indicated. Nuclear extracts isolated from these cells were analyzed on 10 to 30% sucrose density gradients, which were fractionated with continuous monitoring of absorbance at 254 nm (A). Individual fractions were electrophoresed on an SDS–10% polyacrylamide gel and subjected to immunoblotting analysis using affinity-purified anti-Bop1 antibodies. Sn, unfractionated soluble nuclear extracts; P, unsoluble pellet. (D) (Left) Immunoblot analysis with anti-Bop1 antibodies detects Bop1 in nuclear RNPs (N) but not cytoplasmic ribosomes (C). (Right) Electrophoretic analysis of RNA in the fractions used for immunoblotting. RNA was extracted from the nucleoprotein complexes and separated by electrophoresis on a formaldehyde-containing agarose gel to demonstrate the presence equivalent amounts of rRNA in both samples.