Reverse self-splicing of group II intron RNAs in vitro.
Group II introns, which are classed together on the basis of a conserved secondary structure, are found in organellar genes of lower eukaryotes and plants. Like introns in nuclear pre-messenger RNA, they are excised by a two-step splicing reaction to generate branched circular RNAs, the so-called lariats. A remarkable feature of group II introns is their self-splicing activity in vitro. In the absence of a nucleotide cofactor, the intron RNAs catalyse two successive transesterification reactions which lead to autocatalytic excision of the lariat IVS from pre-mRNA and concomitantly to exon ligation. By virtue of its ability to specifically bind the 5' exon, the intron can also catalyse such reactions on exogenous RNA substrates. This sequence-specific attachment could enable group II introns to integrate into unrelated RNAs by reverse splicing, in a process similar to that described for the self-splicing Tetrahymena group I intron. Here we report that group II lariat IVS can indeed reintegrate itself into an RNA composed of the ligated exons in vitro. This occurs by a process of self-splicing that completely reverses both transesterification steps of the forward reaction: it involves a transition of the 2'-5' phosphodiester bond of the lariat RNA into the 3'-5' bond of the reconstituted 5' splice junction.
Augustin
S
S
Institut für Mikrobiologie und Genetik, Universität Wien, Austria.
Müller
M W
MW
Schweyen
R J
RJ
D001059:Apoproteins / Q000235:genetics; D001483:Base Sequence; D003573:Cytochrome b Group / Q000235:genetics; D045303:Cytochromes b; D004951:Esterification; D005091:Exons; D007438:Introns / Q000235:genetics*; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D012313:RNA / Q000235:genetics; D012322:RNA Precursors / Q000235:genetics; D012326:RNA Splicing; D000079962:RNA, Circular; D012331:RNA, Fungal / Q000235:genetics*; D012441:Saccharomyces cerevisiae / Q000235:genetics
D001059:Apoproteins; D003573:Cytochrome b Group; D012322:RNA Precursors; D000079962:RNA, Circular; D012331:RNA, Fungal; D012313:RNA; D045303:Cytochromes b