Figure 6

Acetylated ISWI may reside in the NURF complex, but not in CHRAC/ACF in SL2 cells. (A) Ablation of proteins by RNA interference. Drosophila SL2 cells were treated with double-stranded RNA (RNAi) to interfere with the expression of ISWI, ACF1, NURF301, GCN5 or GST (to control for non-specific effects of dsRNA treatment). After 10 days of incubation whole cell lysates were prepared and probed by Western blotting (WB) for the presence of NURF301, ACF1, ISWI, GCN5 and p55 (a subunit of NURF that serves as a loading control). (B) SL2 cells were treated for 12 days with dsRNAs as indicated. Localization of ISWI^K753ac or bulk ISWI was monitored by immunofluorescence staining using the indicated antibodies. DNA was counterstained with Hoechst 33258. (C) SL2 whole cell extract (upper panel) or nuclear extract from 8–12 hr embryos (lower panel) were subjected to immunoprecipitation with antibodies against ISWI, ACF1 or NURF301. The presence of bulk ISWI (left panel) or ISWI^K753ac (right panel) in the immunoprecipitates was analysed by Western blotting.