Open AccessReconstitution of a group I intron self-splicing reaction with an activator RNA.
Author(s) -
G. Horst,
Alice Christian,
Tan Inoue
Publication year - 1991
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.88.1.184
Subject(s) - intron , rna splicing , ribozyme , tetrahymena , group ii intron , rna , biology , mammalian cpeb3 ribozyme , group i catalytic intron , microbiology and biotechnology , chemistry , genetics , gene
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 MgCl2/200 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 MgCl2/2 mM 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 proceeds highly efficiently under standard conditions (5 mM MgCl2/200 mM NH4Cl, pH 7.5). Two steps of the bimolecular 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.