Regulated Formation of lncRNA-DNA Hybrids Enables Faster Transcriptional Induction and Environmental Adaptation

Abstract

Summary Long non-coding (lnc)RNAs, once thought to merely represent noise from imprecise transcription initiation, have now emerged as major regulatory entities in all eukaryotes. In contrast to the rapidly expanding identification of individual lncRNAs, mechanistic characterization has lagged behind. Here we provide evidence that the GAL lncRNAs in the budding yeast S. cerevisiae promote transcriptional induction in trans by formation of lncRNA-DNA hybrids or R-loops. The evolutionarily conserved RNA helicase Dbp2 regulates formation of these R-loops as genomic deletion or nuclear depletion results in accumulation of these structures across the GAL cluster gene promoters and coding regions. Enhanced transcriptional induction is manifested by lncRNA-dependent displacement of the Cyc8 co-repressor and subsequent gene looping, suggesting that these lncRNAs promote induction by altering chromatin architecture. Moreover, the GAL lncRNAs confer a competitive fitness advantage to yeast cells as expression of these non-coding molecules correlates with faster adaptation in response to an environmental switch.

LncRNAs: Bridging environmental sensing and gene expression

“…Consistently, the GAL lncRNAs have no effect on induction from the derepressed (Craffinose) state, but rather confer a specific fitness advantage to yeast cells over those lacking the GAL lncRNAs during a glucose to galactose switch. 48,53 GAL lncRNA-dependent induction occurs when the GAL lncRNAs are encoded in trans, indicating that lncRNA-dependent transcriptional induction and repression are mechanistically distinct. This result is reminiscent of Air (Airn), which also functions in mechanistically distinct cis and trans roles to regulate gene expression.…”

“…74 Strikingly, nuclear depletion of Dbp2 using the "anchor away" strategy 75 resulted in time-dependent accumulation of R-loops beginning at the 5 0 end of GAL1 and spreading across GAL1, GAL10 and through the 3 0 end of GAL7. 53 Removal of the lncRNAs through genomic deletion reduced R-loops at the 5 0 end of GAL1 and abolished detection of these structures across the rest of the GAL cluster locus. This suggested that Dbp2 regulates formation of lncRNA R-loops at the GAL genes (Fig.…”

“…In support of a role for R-loops in depression of the GAL cluster, we found that ectopic expression of RNase H1 reduced induction of the GAL genes to wild type levels in dbp2D cells. 53 The fact that INO1, an inducible gene without an overlapping, annotated lncRNA, induced at the same rate regardless of the presence of RNAse H1 argues against a general reduction in transcription. 53 Because we observed R-loop accumulation across the entire GAL cluster, this leads one to wonder how these structures could specifically prevent Cyc8 association without impacting subsequent steps of transcriptional activation by Gal4, coactivators, and RNAPII (Fig.…”

“…1C). One possibility is that these R-loops are transient, allowing recruitment of transcriptional 74 and prevents the accumulation of R-loops at the GAL gene cluster 53 via the assembly of Dbp2-dependent lncRNA-protein complexes. 56 This allows the successful docking of the Cyc8/Tup1 co-repressor complex and subsequent repression.…”

“…(C) Galactose Induction In the presence of galactose, the Gal4 transcriptional activator is released from the Gal80 inhibitor (not pictured), 96 enabling recruitment of co-activators and RNA P II. 97 This results in altered chromatin structure, as evidenced by looping of the GAL10 promoter and terminator, 53 which may enhance transcriptional induction. It is currently unknown how the GAL lncRNAs are cleared from chromatin or how Dbp2 is re-imported following long-term growth in galactose.…”