A site-specific endonuclease and co-conversion of flanking exons associated with the mobile td intron of phage T4

Abstract

The product of the td intron open reading frame (ORF) of phage T4 is required for high-frequency transfer of the intervening sequence from intron-plus (In +) to intron-minus (In −) alleles. In vivo studies have demonstrated that the td ORF product targets cleavage of td In − DNA, and that cleavage is correlated with intron inheritance [Quirk et al., Cell 56 (1989) 455–465]. In the present study we show by in vitro synthesis of the td intron ORF product, that the protein possesses endonuclease activity and efficiently cleaves double-stranded DNA at or near the site of intron integration. In addition, we demonstrate that intron insertion is accompanied by co-conversion of the flanking exon sequences. Co-conversion of markers within 50 nt surrounding the site of intron insertion occurred at a high frequency (80–100%), and decreased at greater distance from the intervening sequence. Co-conversion may provide a mechanism for maintaining exon-intron RNA contacts required for accurate splicing of the relocated intron. Cleavage of target DNA by an intron endonuclease and co-conversion of flanking exon sequences are both features associated with mobile introns of eukaryotes, indicating a common mechanism for intron transfer in the eukaryotic and prokaryotic kingdoms.