Group II intron organization in M. acetivorans. (A) A typical group II intron structure (M.a.I1–1). The intron contains six helical domains (indicated by six stem-loop structures) with the ORF encoded in the loop of domain 4 (open box). The exons surrounding the intron are gray boxes, and intron splicing would ligate them together. (B,C,D) Nested organization of group II introns in M. acetivorans. There are four nested introns in B, and three nested introns in C and D. See text for detailed descriptions. Truncated intron copies are indicated by a truncated ORF or the absence of intron RNA domains. The percent identity (e.g., 90%, 232/258) indicates amino acid identity between that particular intron-encoded protein and the reference intron (either M.a.I1–1 or M.a.I5–1). “YADD” and “PQG” are abbreviations for homing sites that contain either the YADD or PQG motif (see F). (E) Comparison of ORF-less (M.a.I6) and ORF-containing (M.a.I5) intron structures. M.a.I5 and M.a.I6 are 82% identical. Their overall secondary structures are shown by the line drawing. Major differences are indicated, but the numerous single base differences are not. A stem-loop is absent from domain I in M.a.I6. There are also sequence differences in EBS1 and EBS2, which are important because they pair with IBS1 and IBS2 in the upstream exon during splicing, and also during reverse splicing into DNA exons in homing reactions. Most importantly, the ORF is deleted from domain 4 of M.a.I6; however, the start and stop codons remain, as well as several amino acid codons, suggesting that M.a.I6 was derived from M.a.I5 by ORF loss. (F) Comparison of insertion sites of ORF-less and ORF-containing introns. Insertion sites are shown for 40 bp upstream and 20 bp downstream of the intron insertion site. IBS1 and IBS2 sequences are shaded, and pair with EBS1 and EBS2 in the intron structure during splicing and reverse splicing reactions. Light shading indicates mispairings in IBS/EBS interactions due to sequence variations. Homing sites of group II introns typically extend from approximately −25 to +10 relative to the intron insertion site. The PQG and YADD amino acid codons are indicated for M.a.I1 and M.a.I5. EBS–IBS pairings between introns and exons are maintained for all four sets of introns. M.a.I1 and M.a.I4 are related introns, but their insertion sites are completely different. Likewise, M.a.I5 and M.a.I6 introns are related but have different insertion sites.
