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. Author manuscript; available in PMC: 2013 Sep 19.
Published in final edited form as: Mol Cell. 2006 Aug 4;23(3):389–399. doi: 10.1016/j.molcel.2006.05.043

Figure 4.

Figure 4

Spliceosome Disassembly is Promoted and Regulated by the GTPase Snu114p (A) Disassembly of Prp43p-associated spliceosomes is inhibited by snu114 mutants. Spliceosomes were purified from the wild-type strain SNU114 (yJPS1053) or the mutant strains snu114-14 (yJPS1120) and snu114-60 (yJPS1056). Spliceosomes were incubated and analyzed as in Figure 2E. (B) prp43 and snu114 mutants genetically interact. yTB135 was cotransformed with plasmids encoding PRP43 (pJPS1346) or prp43-Q423N (pJPS1347) and SNU114 (pTB106), snu114-14 (pTB108) or snu114-60 (pTB113), grown in liquid media, diluted and grown on solid media containing 5-fluoroorotic acid for 5 days at 30°C. (C) Spliceosome disassembly is repressed specifically by GDP and derepressed specifically by GTP. Prp43p-associated spliceosomes, purified from a wild-type strain (yJPS797), were incubated at 4°C for 60 s and analyzed as in Figure 2B. (D) Spliceosome disassembly is regulated by Snu114p. Prp43p-associated spliceosomes were purified from a wild-type strain (yJPS1053) or a snu114-D271N mutant strain (yJPS1054) that appears to switch the specificity of Snu114p from guanine to xanthine (Bartels et al., 2003). Spliceosomes were incubated and analyzed as in Figure 2B. ATP, GDP and XDP concentrations were 2 mM; GTP and XTP concentrations were 0.5 mM. (E) Spliceosome disassembly does not require GTP hydrolysis. Prp43p-associated spliceosomes, purified from wild-type SNU114 (yJPS1053) or mutant snu114-D271N (yJPS1054), were incubated at 4°C for 60 s and analyzed as in Figure 2B.