Fig. 5. Microinjected U3 and U65 snoRNAs are retained within the nucleus and localized to nucleoli of Xenopus oocytes after injection of T24N Ran. (A,C) Recombinant RanT24N in microinjection buffer (bottom panels) or microinjection buffer alone (top panels) were injected into separate sets of oocytes. One hour later, one fmole of fluorescently and ³²P-labeled U3 or U65 snoRNA was injected into the same set of oocytes. Nuclear spreads were made four hours after injection of the RNA, and slides were analyzed by fluorescence microscopy. Several individual nucleoli are shown in each differential interference contrast (DIC) panel. The fluorescence signals (RNA) show that the injected snoRNAs are targeted to a centrally located subregion of the nucleoli in oocytes, which were injected with the T24N Ran (+T24N). (B,D) Nuclear (N) and cytoplasmic (C) fractions were obtained from the same set of injected cells. RNA was extracted from the N and C fractions and subjected to denaturing PAGE followed by autoradiography. The marker lane (M) indicate samples prior to injection.