Contribution of base-pairing interactions between group II intron fragments during trans-splicing in vivo.
Group II introns are mobile genetic elements that self-splice from pre-mRNA transcripts. Some fragmented group II introns found in chloroplastic and mitochondrial genomes are able to assemble and splice in trans. The Ll.LtrB group II intron from the Gram-positive bacterium Lactococcus lactis was shown to splice in trans when fragmented at various locations throughout its structure. Here we used Ll.LtrB to assess the contribution of base-pairing interactions between intron fragments during trans-splicing in vivo. By comparing closely located fragmentation sites, we show that Ll.LtrB trans-splices more efficiently when base-pairing interactions can occur between the two intron fragments. Disruptions and stepwise restorations of specific base-pairing interactions between intron fragments resulted respectively in significant reductions and recoveries of the Ll.LtrB trans-splicing efficiency. Finally, although we confirm that LtrA is an important co-factor for trans-splicing, its overexpression cannot compensate for the reduction in trans-splicing efficiency when the potential base-pairing interactions between intron fragments are disrupted. These findings demonstrate the important contribution of base-pairing interactions for the assembly of group II intron fragments during trans-splicing and rationalizes why such interactions were evolutionarily conserved in natural trans-splicing group II introns.
Department of Microbiology and Immunology, McGill University, Montréal, Québec, Canada.
Cecilia
C
Quiroga
Lisa
L
Kronstad
Christine
C
Ritlop
Audrey
A
Filion
Benoit
B
Cousineau
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D020029:Base Pairing; D004251:DNA Transposable Elements / Q000235:genetics; D007438:Introns / Q000235:genetics*; D013294:Lactococcus lactis / Q000235:genetics* / Q000378:metabolism; D009690:Nucleic Acid Conformation; D012322:RNA Precursors / Q000235:genetics / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism; D020040:Trans-Splicing / Q000235:genetics*
D001426:Bacterial Proteins; D004251:DNA Transposable Elements; C099786:LtrB protein, Lactococcus lactis; D012322:RNA Precursors; C099785:LtrA protein, Lactococcus lactis; D012194:RNA-Directed DNA Polymerase