Splicing of the mitochondrial group-II intron rl1: conserved intron-exon interactions diminish splicing efficiency.
The mitochondrial intron rI1 is a self-splicing group-II intron of algal mitochondria that can be transferred into chloroplasts from the green alga Chlamydomonas reinhardtii for in vivo investigations (Herdenberger et al. 1994). Thus, rI1 is a suitable system to compare in vitro and in vivo RNA processing. Interestingly, rI1 shows correct RNA splicing, although typical cis-acting exon-sequences (IBS2, delta) of group-II introns are lacking. In order to examine the effect of these exon-intron interactions on splicing, we introduced the endogenous mitochondrial IBS2 sequence in order to produce optimal IBS2-EBS2 base pairing. In addition, the first nucleotide of the 3'exon (delta') was substituted to create an optimal delta-delta' interaction. Neither of the two mutations, nor a combination of both, had any effect on the precision of the splice-site selection. Unexpectedly, introduction of IBS2 led to a reduction in the efficiency of the second splicing step in vitro but not in vivo. These findings lead us to conclude that trans-acting factors are present in vivo to optimize splicing efficiency. The possibility is discussed that these factors may, for example, stabilize tertiary intron structures that are a prerequisite for correct RNA processing. Furthermore, our data indicate that similar trans-acting factors promote correct intron splicing in chloroplasts and mitochondria.
Holländer
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Lehrstuhl für Allgemeine Botanik, Ruhr-Universität Bochum, 44780 Bochum, Germany.
Kück
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D017398:Alternative Splicing; D000818:Animals; D016825:Chlamydomonas reinhardtii / Q000235:genetics*; D005091:Exons / Q000235:genetics; D007438:Introns / Q000235:genetics*; D008928:Mitochondria / Q000235:genetics*; D009690:Nucleic Acid Conformation; D018747:RNA, Chloroplast / Q000235:genetics*
D018747:RNA, Chloroplast