Native elongating transcript sequencing reveals global anti-correlation between sense and antisense nascent transcription in fission yeast

Abstract

Antisense transcription can regulate sense gene expression. However, previous annotations of antisense transcription units have been based on detection of mature antisense long noncoding (aslnc)RNAs by RNA-seq and/or microarrays, only giving a partial view of the antisense transcription landscape and incomplete molecular bases for antisense-mediated regulation. Here, we used native elongating transcript sequencing to map genome-wide nascent antisense transcription in fission yeast. Strikingly, antisense transcription was detected for most protein-coding genes, correlating with low sense transcription, especially when overlapping the mRNA start site. RNA profiling revealed that the resulting aslncRNAs mainly correspond to cryptic Xrn1/Exo2-sensitive transcripts (XUTs). ChIP-seq analyses showed that antisense (as)XUT's expression is associated with specific histone modification patterns. Finally, we showed that asXUTs are controlled by the histone chaperone Spt6 and respond to meiosis induction, in both cases anti-correlating with levels of the paired-sense mRNAs, supporting physiological significance to antisense-mediated gene attenuation. Our work highlights that antisense transcription is much more extended than anticipated and might constitute an additional nonpromoter determinant of gene regulation complexity.

Endogenous RNAi pathway evolutionarily shapes the destiny of the antisense lncRNAs transcriptome

“…As previously reported in S. cerevisiae and S. pombe (Wery et al 2016;Atkinson et al 2018;Wery et al 2018b), many lncRNAs are targeted by both Rrp6 and Xrn1 in N. castellii ( Figure 1B).…”

“…To characterize the population of aslncRNAs in N. castellii, we performed genome-wide RNA profiling using RNA-seq data obtained from WT, dcr1, xrn1 and rrp6 cells ( Figure 1A). For the identification of DUTs and XUTs, we performed RNA-Seq in WT, dcr1 and xrn1 strains, followed by segmentation using the algorithm that we previously developed to annotate CUTs (Watts et al 2018) and XUTs (Wery et al 2016;Wery et al 2018b) in other yeast species. For the identification of N.…”

“…Both Rrp6 and Xrn1 are conserved across Eukaryotes (Houseley et al 2006;Nagarajan et al 2013). In this respect, CUTs and XUTs were recently identified in fission yeast (Atkinson et al 2018;Watts et al 2018;Wery et al 2018b), and they are also mainly antisense to protein-coding genes in this species. This indicates that the roles of the nuclear exosome and Xrn1 in restricting aslncRNAs levels have been conserved across the yeast clade.…”

“…S. pombe has a functional RNAi machinery (Volpe et al 2002), but asXUTs are insulated from it (Wery et al 2018b). This is probably explained by different subcellular localization, as Dicer is restricted to the nucleus in fission yeast, mainly contributing in heterochromatin formation at centromeric repeats (Woolcock and Buhler 2013).…”

Fine chromatin-driven mechanism of transcription interference by antisense noncoding transcription

“…Early termination of noncoding transcription is not strict and mostly depends on the number of Nrd1-Nab3 recognition motifs carried by the lncRNA (Schulz et al 2013;Castelnuovo et al 2014). This implies that some long noncoding RNAs will be cleared through early-termination while others will naturally extend into sense promoters, followed by export and degradation in the cytoplasm via Nonsense-Mediated Decay (NMD) (Malabat et al 2015;Wery et al 2018). Importantly, artificial loss of early termination through Nrd1 depletion leads to antisense elongation into paired sense promoters and to a subsequent transcription interference directly correlated with the levels of lncRNA accumulation over the promoters (Schulz et al 2013).…”