Facilitation of group I splicing in vivo: misfolding of the Tetrahymena IVS and the role of ribosomal RNA exons.
Self-splicing of the group I IVS from Tetrahymena thermophila rDNA is limited by the time required for the RNA to reach its active conformation. In vitro, folding is slow because the pre-rRNA becomes kinetically trapped in inactive structures. In vivo, splicing is 50 times more rapid, implying that misfolding of the pre-rRNA is corrected. Exon mutations that inhibit self-splicing 100-fold in vitro were fully rescued when the pre-rRNA containing the IVS was expressed in E. coli. In contrast, IVS mutations that cause misfolding were only partially suppressed at 42 degrees C, and doubled the activation energy of splicing. These results suggest that intracellular folding of the pre-rRNA involves metastable intermediates similar to those observed in vitro. Precursors with natural rRNA exons were more active and less cold-sensitive than those with non-rRNA exons. This shows that the rRNA reduces misfolding of the IVS, thereby facilitating splicing of the pre-rRNA in vivo.
Nikolcheva
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T
Program in Molecular and Cell Biology, University of Maryland, College Park, MD 20742-2021, USA.
Woodson
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SA
D000818:Animals; D001483:Base Sequence; D004926:Escherichia coli; D005091:Exons / Q000235:genetics; D007438:Introns; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D012313:RNA / Q000235:genetics*; D012322:RNA Precursors / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics*; D012335:RNA, Ribosomal / Q000235:genetics*; D013696:Temperature; D013768:Tetrahymena / Q000235:genetics*
D012322:RNA Precursors; D012335:RNA, Ribosomal; D012313:RNA