The circle to lariat ratio of the Ll.LtrB group II intron from Lactococcus lactis is greatly influenced by a variety of biological determinants in vivo

Abstract

Bacterial group II introns mostly behave as versatile retromobile genetic elements going through distinct cycles of gain and loss. These large RNA molecules are also ribozymes splicing autocatalytically from their interrupted pre-mRNA transcripts by two different concurrent pathways, branching and circularization. These two splicing pathways were shown to release in bacterial cells significant amounts of branched intron lariats and perfect end-toend intron circles respectively. On one hand, released intron lariats can invade new sites in RNA and/or DNA by reverse branching while released intron circles are dead end spliced products since they cannot reverse splice through circularization. The presence of two parallel and competing group II intron splicing pathways in bacteria led us to investigate the conditions that influence the overall circle to lariat ratio in vivo. Here we unveil that removing a prominent processing site within the Ll.LtrB group II intron, raising growth temperature of Lactococcus lactis host cells and increasing the expression level of the intron-interrupted gene all increased the relative amount of released intron circles compared to lariats. Strengthening and weakening the base pairing interaction between the intron and its upstream exon respectively increased and decreased the overall levels of released intron circles in comparison to lariats. Host environment was also found to impact the circle to lariat ratio of the Ll.LtrB and Ll.RlxA group II introns from L. lactis and the Ef.PcfG intron from Enterococcus faecalis. Overall, our data show that multiple factors significantly influence the balance between released intron circles and lariats in bacterial cells.

MatK impacts Differential Chloroplast Translation of Ribosomal and Photosynthetic genes by limiting spliced tRNA-K(UUU) abundance

“…In the absence of functional evidence for the alternative splicing products, we consider the alternative lariat as aberrant and detrimental, possibly arising due to misfolding of the RNA that is enhanced at higher temperatures. This would be reminiscent of the finding that the circle-to-lariat ratio of the Ll.LtrB group II intron from Lactococcus lactis is significantly influenced by temperature changes (61) and the general sensitivity of splicing to heat and cold in plants (62). MatK is fostering the canonical splicing pathways as suggested by the correlation of its expression with tRNA-K(UUU) levels in the over-expressor.…”

Identification of Group II Intron RmInt1 Binding Sites in a Bacterial Genome

“…Under physiological conditions, both the RT and X domains form contacts with several intron RNA domains (DIV, DII, DIII and DVI) to promote intron folding and splicing (Wank et al, 1999;Zhao and Pyle, 2017). Intron excision occurs by means of two transesterification reactions resulting in ligated exons and several forms of excised introns (lariat, circular, and linear, depending on biological determinants and host environment, Monat and Cousineau, 2020). After splicing, the IEP remains bound to the excised lariat RNA, forming the RNP complex, which performs the mobility reaction via an RNA intermediate (Cousineau et al, 1998;Martínez-Abarca et al, 2004).…”