RNase treatment or knockdown of specific PB components provokes the simultaneous dissolution of PBs and Pab 1801 puncta.
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A, live U2OS cells were permeabilized with 0.1% TX100 for five minutes and then exposed to CSKB without (Control) or with 100 µg/ml of RNase (RNase) for additional 5 minutes. The percentage of cells with PBs simultaneous stained with Dcp1a and Pab 1801 was evaluated in 100 cells from duplicate experiments for each treatment and diminished from 75% in control cells to 40% in RNAse-treated cells. The remaining Pab 1801 foci have less than half of the size of control cells. B, U2OS and SK-N-SH cells were treated with specific siRNAs against the PB components Hedls, Rck/p54 or 4ET, and stained with the indicated antibodies. The Pab 1801 puncta vanished when PB are disrupted. The proportion of cells with foci is indicated, as evaluated in 100 cells from duplicate experiments for each treatment. As before (Figures 4, 5 and 6), all puncta were double stained for each pair of antibodies and single-stained foci were not present in any of the treatments. Bar, 10 µm.