B Retrohoming

GP II INTRON

Repair via DNA

intron-encoded endonuclease endonuclease ORF

inteiri endonuclease exon/extein DNA intron DNA

inteiri DNA intron RNA

HS homing site intron-encoded RNP

Fig. 2. Mobility of introns and inteins. a DNA-based homing of group I introns and interns. The intron or intein endonuclease cleaves the homing site of a cognate intron- or in-tein-less allele. Gene conversion repairs the break to generate intron- or intein-containing products (Dujon, this Vol.). b Retrohoming of a group II intron. In this case, the homing site DNA is invaded by intron RNA, and the opposite strand is cleaved by an intron-encod-ed protein, which is part of an RNP complex. The intron is copied into cDNA to generate the intron-containing product (Lambowitz et al., this Vol.).

the intron or intein (Fig. 2A). The group II intron-encoded proteins are more complex, forming a ribonucleoprotein (RNP) particle with the intron RNA (Fig. 2B). The intron invades the DNA sense strand (mRNA-like strand) by reverse-splicing, whereas the endonuclease domain of the protein nicks the antisense strand. The intron acquisition event is completed with a cDNA copy of the intron, in a process termed retrohoming (Lambowitz et al., this Vol.).

The homing endonucleases fall within four families, characterized by the sequence motifs LAGLIDADG (Caprara and Waring, this Vol.; Chevalier et al., this Vol.; Dujon, this Vol.; Haber and Wolfe, this Vol), GIY-YIG (Edgell, this Vol.; Van Roey and Derbyshire, this Vol.), His-Cys box (Galburt and Jurica, this Vol.; Keeble et al., this Vol.) and HNH (Keeble et al., this Vol.). However, recent structural data support the hypothesis that the His-Cys box and HNH

enzymes share features at their active sites, and should be considered a single family, called ppa-Me (Keeble et al., this Vol.). All of the homing endonu-cleases recognize lengthy asymmetric or pseudosymmetric DNA sequences, ranging from a 14-bp homing site for I-Dmol, a member of the LAGLIDADG family, to a 40-bp site for I-TevI, a member of the GIY-YIG family (described in Chevalier et al., this Vol.; Van Roey and Derbyshire, this Vol., respectively). In addition, the enzymes exhibit varying degrees of sequence tolerance, with I-TevI again being exceptional, in this case in its promiscuity (Van Roey and Derbyshire, this Vol.). The conserved sequences and substrate-recognition characteristics stand in contrast to the properties of the restriction endonu-cleases, which usually recognize short palindromic DNA sequences with absolute sequence specificity. Thus, while both types of endonuclease cleave DNA, they have evolved independently. A growing understanding of the structures and mechanisms of some of these enzymes is facilitating their engineering for genomic applications (Dujon, this Vol.; Gimble, this Vol.).

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