Self-splicing of a group I intron reveals partitioning of native and misfolded RNA populations in yeast.
Stable RNAs must form specific three-dimensional structures, yet many RNAs become kinetically trapped in misfolded conformations. To understand the factors that control the accuracy of RNA folding in the cell, the self-splicing activity of the Tetrahymena group I intron was compared in different genetic contexts in budding yeast. The extent of splicing was 98% when the intron was placed in its natural rDNA context, but only 3% when the intron was expressed in an exogenous pre-mRNA. Further experiments showed that the probability of forming the active intron structure depends on local sequence context and transcription by Pol I. Pre-rRNAs decayed at similar rates, whether the intron was wild type or inactivated by an internal deletion, suggesting that most of the unreacted pre-rRNA is incompetent to splice. Northern blots and complementation assays showed that mutations that destabilize the intron tertiary structure inhibited self-splicing and processing of internal transcribed spacer 2. The data are consistent with partitioning of pre-rRNAs into active and inactive populations. The misfolded RNAs are sequestered and degraded without refolding to a significant extent. Thus, the initial fidelity of folding can dictate the intracellular fate of transcripts containing this group I intron.
Jackson
Scott A
SA
Department of Chemistry and Biochemistry, University of Maryland, College Park, Maryland 20742, USA.
Koduvayur
Sujatha
S
Woodson
Sarah A
SA
D000818:Animals; D001483:Base Sequence; D004275:DNA, Ribosomal; D005816:Genetic Complementation Test; D007438:Introns; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012326:RNA Splicing; D012331:RNA, Fungal / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012335:RNA, Ribosomal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013696:Temperature; D013768:Tetrahymena / Q000235:genetics*
D004275:DNA, Ribosomal; D012322:RNA Precursors; D012331:RNA, Fungal; D012335:RNA, Ribosomal; C065039:RNA, ribosomal, 25S; D012319:RNA Polymerase II