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. 2018 Jan 30;46(7):3764–3773. doi: 10.1093/nar/gky052

Figure 5.

Figure 5.

Cwc23 facilitates disassembly of spliceosome intermediates. (A) Splicing reactions performed in Slu7 and Prp22 doubly-depleted extracts were precipitated with anti-Ntc20 antibody (lane 1). The isolated spliceosomes were incubated without (lanes 2 and 3) or with (lanes 4–7) Ntr1–Ntr2 and Prp43, without (lanes 4 and 5) or with (lanes 6 and 7) further addition of Cwc23. Supernatant and pellet fractions were separated for analysis. (B) Splicing reactions performed in Cwc25-depleted extracts were precipitated with anti-Ntc20 antibody (lane 1). The isolated spliceosomes were incubated without (lanes 2 and 3) or with (lanes 4–7) Ntr1–Ntr2 dimer and Prp43, without (lanes 4 and 5) or with (lanes 6 and 7) further addition of Cwc23. Supernatant and pellet fractions were separated for analysis. (C) Quantification of disassembly efficiency in the presence or absence of Cwc23. Amounts of RNA from Figures 4B, 5A and 5B were quantified by a PhosphoImager. Data are presented as mean ± SD of three (dCwc25 and dPrp43) or four (dSlu7/dPrp22) replicates. P values for dCwc25, dSlu7/dPrp22 and dPrp43 are 0.000933, 0.002647 and 0.778647, respectively. 1/2/43, Ntr1–Ntr2 and Prp43; T, total precipitates; P, pellet; S, supernatant.