RECONSTITUTION OF A GROUP-I INTRON SELF-SPLICING REACTION WITH AN ACTIVATOR RNA

The self-splicing rRNA intron of Tetrahymena thermophila belongs to a subgroup of group I introns that contain a conserved extra stem-loop structure termed P5abc. A tetrahymena mutant precursor RNA lacking this P5abc is splicing-defective under standard conditions (5 mM MgCl2/2 mM NH4Cl, pH 7.5) in vitro. However, the mutant precursor RNA by itself is capable of performing the self-splicing reaction without P5abc under different conditions (15 mM MgCl2/2mM spermidine, pH 7.5). We have investigated the functional role of the P5abc in the mechanism of the self-splicing reaction. When an RNA consisting of the P5abc but lacking the rest of the Tetrahymena intron is incubated with the mutant precursor, the self-splicing reaction can be performed accurately by a shortened precursor RNA containing all essential components required in the self-splicing reaction and an activator RNA consisting of the P5abc. Gel-mobility-shift assays suggest that two molecules associate by a direct RNA.RNA interaction during the splicing reaction. The results imply that there might exist other small RNAs whose role is to activate ribozymes.