Nuclear Processing of the 3′-Terminal Nucleotides of Pre-U1 RNA in Xenopus laevis Oocytes

Abstract

U1 small nuclear RNA is synthesized as a precursor with several extra nucleotides at its 3′ end. We show that in Xenopus laevis oocytes, removal of the terminal two nucleotides occurs after the RNA has transited through the cytoplasm and returned to the nucleus. The activity is controlled by an inhibitor of processing, which we call TPI, for 3′-terminal processing inhibitor. This inhibitor is sensitive to both micrococcal nuclease and trypsin treatment, indicating that it is a nucleoprotein. TPI inhibits the 3′ processing of pre-U1 RNAs that have 5′ ends containing m⁷G caps but not mature m^2,2,7G caps; this finding suggests that TPI interacts directly or indirectly with the 5′ end of pre-U1 RNA. The inhibition of processing by TPI, almost complete at 19°C, is reversibly inactivated at slightly higher temperatures. TPI activity is solely in the soluble fraction of oocyte nuclear extracts, in contrast to the 3′-terminal processing activity, which is present in both the particulate and soluble fractions. We propose that the differential processing of the 3′-terminal nucleotides of pre-Ul RNA after its return from the cytoplasm, but not before its exit from the nucleus, may be due to the association of TPI with the m⁷G cap on the newly synthesized pre-U1 RNA.