SUMMARYGroup II introns are mobile ribozymes that self-splice from precursor RNAs to yield excised intron lariat RNAs, which then invade new genomic DNA sites by reverse splicing. The introns encode a reverse transcriptase that stabilizes the catalytically active RNA structure for forward and reverse splicing, and afterwards converts the integrated intron RNA back into DNA. The characteristics of group II introns suggest that they or their close relatives were evolutionary ancestors of spliceosomal introns, the spliceosome, and retrotransposons in eukaryotes. Further, their ribozyme-based DNA integration mechanism enabled the development of group II introns into gene targeting vectors ("targetrons"), which have the unique feature of readily programmable DNA target specificity.

Mobile group II introns, found in bacterial and organellar genomes, are both catalytic RNAs and retrotransposable elements. They use an extraordinary mobility mechanism in which the excised intron RNA reverse splices directly into a DNA target site and is then reverse transcribed by the intron-encoded protein. After DNA insertion, the introns remove themselves by protein-assisted, autocatalytic RNA splicing, thereby minimizing host damage. Here we discuss the experimental basis for our current understanding of group II intron mobility mechanisms, beginning with genetic observations in yeast mitochondria, and culminating with a detailed understanding of molecular mechanisms shared by organellar and bacterial group II introns. We also discuss recently discovered links between group II intron mobility and DNA replication, new insights into group II intron evolution arising from bacterial genome sequencing, and the evolutionary relationship between group II introns and both eukaryotic spliceosomal introns and non-LTR-retrotransposons. Finally, we describe the development of mobile group II introns into gene-targeting vectors, "targetrons," which have programmable target specificity.

Catalytic RNAs are often regarded as molecular fossils from the RNA World, yet it is usually difficult to get more specific information about their evolution. Here we have investigated the coevolution of group II intron RNA structures with their intron-encoded reverse transcriptases (RTs). Unlike group I introns, there has been no obvious reshuffling between intron RNA structures and ORFs. Of the six classes of intron structures that encode ORFs, three are conventional forms of group II A1, B1, and B2 secondary structures, whereas the remaining classes are bacterial, are possibly associated with the most primitive ORFs, and have unusual features and hybrid features of group IIA and group IIB intron structures. Based on these data, we propose a new model for the evolution of group II introns, designated the retroelement ancestor hypothesis, which predicts that the major RNA structural forms of group II introns developed through coevolution with the intron-encoded protein rather than as independent catalytic RNAs, and that most ORF-less introns are derivatives of ORF-containing introns. The model is supported by the distribution of ORF-containing and ORF-less introns, and by numerous examples of ORF-less introns that contain ORF remnants.

Bordetella bacteriophages generate diversity in a gene that specifies host tropism. This microevolutionary adaptation is produced by a genetic element that combines the basic retroelement life cycle of transcription, reverse transcription and integration with site-directed, adenine-specific mutagenesis. Central to this process is a reverse transcriptase-mediated exchange between two repeats; one serving as a donor template (TR) and the other as a recipient of variable sequence information (VR). Here we describe the genetic basis for diversity generation. The directionality of information transfer is determined by a 21-base-pair sequence present at the 3' end of VR. On the basis of patterns of marker transfer in response to variant selective pressures, we propose that a TR reverse transcript is mutagenized, integrated into VR as a single non-coding strand, and then partially converted to the parental VR sequence. This allows the diversity-generating system to minimize variability to the subset of bases under selection. Using the Bordetella phage cassette as a signature, we have identified numerous related elements in diverse bacteria. These elements constitute a new family of retroelements with the potential to confer selective advantages to their host genomes.

Group II introns are widely believed to have been ancestors of spliceosomal introns, yet little is known about their own evolutionary history. In order to address the evolution of mobile group II introns, we have compiled 71 open reading frames (ORFs) related to group II intron reverse transcriptases and subjected their derived amino acid sequences to phylogenetic analysis. The phylogenetic tree was rooted with reverse transcriptases (RTs) of non-long terminal repeat retroelements, and the inferred phylogeny reveals two major clusters which we term the mitochondrial and chloroplast-like lineages. Bacterial ORFs are mainly positioned at the bases of the two lineages but with weak bootstrap support. The data give an overview of an apparently high degree of horizontal transfer of group II intron ORFs, mostly among related organisms but also between organelles and bacteria. The Zn domain (nuclease) and YADD motif (RT active site) were lost multiple times during evolution. Differences in domain structures suggest that the oldest ORFs were concise, while the ORF in the mitochondrial lineage subsequently expanded in three locations. The data are consistent with a bacterial origin for mobile group II introns.