BASE MODIFICATION PATTERN AT THE WOBBLE POSITION OF XENOPUS SELENOCYSTEINE TRNA(SEC)

We examined the base modification pattern of Xenopus tRNA(Sec) using m icroinjection into Xenopus oocytes, with particular focus an the wobbl e base U34 at the first position of the anticodon. We found that U34 b ecomes modified to mcm(5)U34 (5-methylcarboxymethyluridine) in the ooc yte cytoplasm in a rather complex manner. When the tRNA(Sec) gene is i njected into Xenopus oocyte nuclei, Psi 55 and m(1)A58 are readily obt ained, but not mcm(5)U34. This will appear only upon cytoplasmic injec tion of the gene product arising from the first nuclear injection. In contrast, tRNA(Sec) produced by in vitro transcription with T7 RNA pol ymerase readily acquires i(6)A37, Psi 55, m(1)A58, and mcm(5)U34. The latter is obtained after direct nuclear or cytoplasmic injections. It has been reported by others that mcm(5)Um, a 2'-O-methylated derivativ e of mcm(5)U34, also exists in rat and bovine tRNA(Sec). With both the gene product and the in vitro transcript, and using the sensitive RNa se T2 assay, we were unable to detect under our conditions the presenc e of a dinucleotide carrying mcm(5)Um and that would be therefore refr actory to hydrolysis. We showed that the unusual mcm(5)U acquisition p athway does not result from impairment of nucleocytoplasmic transport. Rather, these data can be interpreted to mean that the modification i s performed by a tRNA(Sec) specific enzyme, limiting in the oocyte cyt oplasm.