The intron of the mitochondrial 21S rRNA gene of Saccharomyces cerevisiae possesses a long internal reading frame (ORF) that is conserved in various yeast species. In crosses between intron-plus and intron-minus variants, this intron determines a specific gene conversion phenomenon, which results in the integration of the intron sequence within all previously intron-minus copies of the gene. We show, from a frameshift mutant within the intron ORF and from the need of mitochondrial protein synthesis, that ORF encodes a protein active in the gene conversion that spreads the intron within populations of interbreeding strains. This new intron function is reminiscent of the XtransposaseX encoded by mobile genetic elements and is discussed in relation to other intron functions.