2 GROUP-I RIBOZYMES WITH DIFFERENT FUNCTIONS IN A NUCLEAR RDNA INTRON

DiSSU1, a mobile intron in the nuclear rRNA gene of Didymium iridis, w as previously reported to contain two independent catalytic RNA elemen ts. We have found that both catalytic elements, renamed GIR1 and GIR2, are group I ribozymes, but with differing functionality. GIR2 carries out the several reactions associated with self-splicing. GIR1 carries out a hydrolysis reaction at an internal processing site (IPS-1). The se conclusions are based on the catalytic properties of RNAs transcrib ed in vitro. Mutation of the P7 pairing segment of GIR2 abrogated self -splicing, while mutation of P7 in GIR1 abrogated hydrolysis at the IP S-1. Much of the P2 stem and all of the associated loop could be delet ed without effect on self-splicing, These results are accounted for by a secondary structure model, in which a long P2 pairing segment bring s the 5' splice site to the GIR2 catalytic core. GIR1 is the smallest natural group I ribozyme yet reported and is the first example of a gr oup I ribozyme whose presumptive biological function is hydrolysis. We hypothesize that GIR1-mediated cleavage of the excised intron RNA fun ctions in the generation and expression of the mRNA for the intron-enc oded endonuclease I-DirI.