A group III intron is formed from domains of two individual group II introns

A 1352-nucleotide intron within the Euglena gracilis chloroplast ycf8 gene has been characterized as a complex twintron with overlapping internal introns and alternative splicing pathways. Partially spliced pre-mRNAs were characterized by a combination of cDNA cloning and sequencing, Northern hybridization, and S1 nuclease protection analyses. In the predominant pathway, two internal group II introns (601 and 392 nucleotides) are spliced from subdomain ID of an external group II intron (359 nucleotides). In an alternative pathway, following excision of the 601-nucleotide intron, splicing of a group III intron occurs. This group III intron is recruited from sequences of the external intron and the 392-nucleotide intron. This is the first evidence that a group III intron can be derived from portions of existing group II introns. The mechanism of group III intron formation may also be relevant to the evolution of nuclear introns from putative group II intron ancestors.