Self-splicing of the Tetrahymena intron from mRNA in mammalian cells

The Tetrahymena pre-rRNA self-splicing intron is shown to function in the u nnatural context of an mRNA transcribed by RNA polymerase II in mammalian c ells, Mutational analysis supports the conclusion that splicing in cells oc curs by the same RNA-catalyzed mechanism established for splicing in vitro. Insertion of the intron at five positions spanning the luciferase open rea ding frame revealed 10-fold differences in accumulation of ligated exons an d in luciferase activity; thus, the intron self-splices in many exon contex ts, but the context can have a significant effect on activity. In addition, even the best self-splicing constructs, which produced half as much mRNA a s did an uninterrupted luciferase gene, gave similar to 100-fold less lucif erase enzyme activity, revealing an unexpected discontinuity between mRNA p roduction and translation in cells. The finding that production of accurate ly spliced mRNA in cells does not guarantee a corresponding level of protei n production is surprising, and may have implications for the development o f traits-splicing ribozymes as therapeutics.