Effect of Group IIC Introns on Host Gene Expression in S. cerevisiae

Group II introns are self-splicing ribozymes believed to be the evolutionary ancestors to eukaryotic spliceosomal introns. They are found in bacteria, archaea, and eukaryotic organelles but not in eukaryotic nuclear genomes. Group IIA and IIB introns introduced to nuclei in Saccharomyces cerevisiae are able to transcribe and splice, but post-transcriptionally silence their host genes. It has been shown that both RNA-RNA interactions and pre-mRNA cytoplasmic mislocalization contribute to the silencing of the genes. RNA-RNA interactions include binding between the exon-binding sequences (EBS) in the intron and intron-binding sequences (IBS) in the ligated exons. Group IIC introns contain reduced EBS-IBS sequences compared to their group IIA and group IIB counterparts; thus it is worthwhile to investigate if IIC introns will function differently in yeast, either causing no silencing or using a distinct silencing mechanism. To this end, a group IIC-reporter cassette was prepared and transformed into yeast. Phenotype screening of the yeast was carried out to determine if silencing occurs. Mechanisms, including the presence of RNA-RNA interactions and their impact on the host genes were investigated. Primer extension assays conducted show that group IIC-reporter constructs can splice in vivo in S. cerevisiae, although exon ligation does not occur. This is a noteworthy distinction in silencing mechanism between the closely related group IIA, IIB, and IIC introns. This information will assist in establishing the evolutionary narrative of the expulsion of group II introns from nuclear genomes and the evolution of group II introns to ancestrally-related spliceosomal introns