Figure 4.

Capped ³²P-labeled snRNA transcripts were injected into the cytoplasm of stage VI Xenopus oocytes. After 18 h incubation, GVs and cytoplasm were isolated and immunoprecipitated with either mAb H1 (anti-coilin) or mAb Y12 (anti-Sm). RNA was isolated from the immunoprecipitates and from untreated control fractions and separated on a polyacrylamide gel. (A) Autoradiograph of entire gel from experiment in which wild-type U7 snRNA was injected. Lane 1 contains a sample of the injected snRNA. Lane 2 contains a sample of total cytoplasmic RNA before immunoprecipitation. Lanes 3 and 4 contain RNA immunoprecipitated from cytoplasm by mAbs H1 and Y12, respectively. Lanes 5, 6, and 7 contain a sample of total GV RNA and the RNA immunoprecipitated by mAbs H1 and Y12. (B) Autoradiograph of relevant portion of gels from experiments in which various constructs were injected; lanes as in panel A. U1 and U2 are wild-type U1 and U2 snRNA. U7, U7(U2), and U7(mut) are wild-type U7, U7 with its Sm site replaced by that of U2, and U7 with an unrelated sequence at the Sm site (wild-type U7 lane is the same as in panel A). Note that all constructs except U7(mut) are immunoprecipitated from the GV by mAb Y12, demonstrating that they exist as Sm complexes. However, only wild-type U7 and, to a lesser extent, U7(U2) are immunoprecipitated from the GV by mAb H1, indicating an association with coilin. (C) The lanes in panel B were quantitated with a phosphorimager, and lanes 3, 4, 6, and 7 were plotted as % immunoprecipitated.