Abstract
Antisense long noncoding (aslnc)RNAs are extensively degraded by the nuclear exosome and the cytoplasmic exoribonuclease Xrn1 in the budding yeast Saccharomyces cerevisiae, lacking RNAi. Whether the ribonuclease III Dicer affects aslncRNAs in close RNAi-capable relatives remains unknown. Using genome-wide RNA profiling, here we show that aslncRNAs are primarily targeted by the exosome and Xrn1 in the RNAi-capable budding yeast Naumovozyma castellii, Dicer only affecting Xrn1-sensitive aslncRNAs levels in Xrn1-deficient cells. The dcr1 and xrn1 mutants display synergic growth defects, indicating that Dicer becomes critical in the absence of Xrn1. Small RNA sequencing showed that Dicer processes aslncRNAs into small RNAs, with a preference for Xrn1-sensitive aslncRNAs. Consistently, Dicer localizes into the cytoplasm. Finally, we observed an expansion of the exosome-sensitive antisense transcriptome in N. castellii compared with S. cerevisiae, suggesting that the presence of cytoplasmic RNAi has reinforced the nuclear RNA surveillance machinery to temper aslncRNAs expression. Our data provide fundamental insights into aslncRNAs metabolism and open perspectives into the possible evolutionary contribution of RNAi in shaping the aslncRNAs transcriptome.
Highlights
Considered as by-products of the pervasive transcription of eukaryotic genomes, long noncodingRNAs have been progressively recognized as genuine transcripts playing important roles in the regulation of multiple cellular processes (Mercer et al, 2009; Wery et al, 2011; Rinn & Chang, 2012; Jarroux et al, 2017)
Among all the ≥200-nt segments not overlapping a coding sequence, tRNA, sn(o)RNA or rRNA on the same strand, using a signal threshold and differential expression analysis between each mutant and its corresponding WT control (Fig 1A; see the Materials and Methods section), we identified 146 stable unannotated transcripts (SUTs, i.e., lncRNAs detected in the WT context but not significantly stabilized in any of the mutant), 10 Dicer-sensitive unstable transcripts (DUTs), 1,021 Xrn1sensitive unstable transcripts (XUTs), and 1,280 cryptic unstable transcripts (CUTs) (Figs 1B and S1A–C)
The number of DUTs appears to be dramatically low compared with CUTs and XUTs, indicating than the effect of Dcr1 on the lncRNAs transcriptome of N. castellii is marginal compared with Rrp6 and Xrn1 (Fig 1C–E)
Summary
Considered as by-products of the pervasive transcription of eukaryotic genomes, long noncoding (lnc)RNAs have been progressively recognized as genuine transcripts playing important roles in the regulation of multiple cellular processes (Mercer et al, 2009; Wery et al, 2011; Rinn & Chang, 2012; Jarroux et al, 2017). Pioneer works in S. cerevisiae have revealed that they are extensively degraded by RNA surveillance machineries (Tisseur et al, 2011; Tudek et al, 2015). These “cryptic” aslncRNAs cannot be detected in wild-type (WT) cells but accumulate upon inactivation of the factor responsible for their degradation. Despite some of them are produced from intergenic regions, most CUTs and XUTs are antisense to protein-coding genes, at least partially
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